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Sample records for psychrotrophic facultative intracellular

  1. Complete genome of Phenylobacterium zucineum – a novel facultative intracellular bacterium isolated from human erythroleukemia cell line K562

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    Sun Jie

    2008-08-01

    Full Text Available Abstract Background Phenylobacterium zucineum is a recently identified facultative intracellular species isolated from the human leukemia cell line K562. Unlike the known intracellular pathogens, P. zucineum maintains a stable association with its host cell without affecting the growth and morphology of the latter. Results Here, we report the whole genome sequence of the type strain HLK1T. The genome consists of a circular chromosome (3,996,255 bp and a circular plasmid (382,976 bp. It encodes 3,861 putative proteins, 42 tRNAs, and a 16S-23S-5S rRNA operon. Comparative genomic analysis revealed that it is phylogenetically closest to Caulobacter crescentus, a model species for cell cycle research. Notably, P. zucineum has a gene that is strikingly similar, both structurally and functionally, to the cell cycle master regulator CtrA of C. crescentus, and most of the genes directly regulated by CtrA in the latter have orthologs in the former. Conclusion This work presents the first complete bacterial genome in the genus Phenylobacterium. Comparative genomic analysis indicated that the CtrA regulon is well conserved between C. crescentus and P. zucineum.

  2. Psychrotrophic Pseudomonas mandelii CBS-1 produces high levels of poly-β-hydroxybutyrate.

    Science.gov (United States)

    Li, Rongpeng; Jiang, Yuji; Wang, Xinfeng; Yang, Jingjing; Gao, Yuan; Zi, Xiaoli; Zhang, Xia; Gao, Haofeng; Hu, Nan

    2013-01-01

    A novel facultative psychrotroph (strain CBS-1), which accumulates poly-β-hydroxybutyrate (PHB), was isolated from soil samples taken from Changbai Mountain, China. Phylogenetic analysis based on 16S rRNA sequence data and Biolog analysis identified strain CBS-1 as Pseudomonas mandelii. Transmission electron micrographs revealed abundant electron-transparent intracellular granules. (1)H-nuclear magnetic resonance analysis revealed that the granules were composed of PHB. P. mandelii CBS-1 grew optimally at 20°C. When cultured aerobically for 48 h with sucrose as the sole carbon source, strain CBS-1 yielded a maximum cell density of 29.3 g/L cell dry weight and synthesized 22.3 g/L of PHB. The ability of strain CBS-1 to grow at a low temperature and rapidly synthesize high levels of PHB may reduce the costs of industrial PHB production.

  3. Vibrio cholerae O1 strains are facultative intracellular bacteria, able to survive and multiply symbiotically inside the aquatic free-living amoeba Acanthamoeba castellanii.

    Science.gov (United States)

    Abd, Hadi; Saeed, Amir; Weintraub, Andrej; Nair, G Balakrish; Sandström, Gunnar

    2007-04-01

    Vibrio cholerae species are extracellular, waterborne, gram-negative bacteria that are overwhelmed by predators in aquatic environments. The unencapsulated serogroup V. cholerae O1 and encapsulated V. cholerae O139 cause epidemic and pandemic outbreaks of cholera. It has recently been shown that the aquatic and free-living amoeba Acanthamoeba castellanii is not a predator to V. cholerae O139; rather, V. cholerae O139 has shown an intracellular compatibility with this host. The aim of this study was to examine the ability of V. cholerae O1 classical and El Tor strains to grow and survive in A. castellanii. The interaction between A. castellanii and V. cholerae O1 strains was studied by means of amoeba cell counts and viable counts of the bacteria in the absence or presence of amoebae. The viable count of intracellularly growing bacteria was estimated by utilizing gentamicin assay. Confocal microscopy and electron microscopy were used to determine the intracellular localization of V. cholerae in A. castellanii. The results showed that V. cholerae O1 classical and El Tor strains grew and survived intracellularly in the cytoplasm of trophozoites, and that the bacteria were also found in the cysts of A. castellanii. The interaction showed a facultative intracellular behaviour of V. cholerae O1 classical and El Tor strains and a possible role of A. castellanii as an environmental host of V. cholerae species.

  4. Additional evidence of the facultative intracellular nature of the fish bacterial pathogen Piscirickettsia salmonis Evidencia adicional de la naturaleza intracelular facultativa del patógeno bacteriano de peces Piscirickettsia salmonis

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    F Gómez

    2009-01-01

    Full Text Available Piscirickettsia salmonis is a highly virulent and contagious microorganism that affects net pen-reared salmonid fish worldwide since the last third of the past century, with little knowledge about its intracellular survival mechanisms. Following a number of recent and non-conclusive reports which questioned its obligate intracellular condition, we decided to show additional evidence to challenge this well-established paradigm carrying on both basic biology as well as classical molecular experiments that confirm its facultative intracellular nature. In this report, we unequivocally demonstrate that the bacteria recovered from tissue culture amplification, in vitro grown agar plates or grown in liquid cultures, were the same organism and all of these isolates equally fulfils Koch’s postulates. In addition, genomic and proteomic analyses confirmed that bacteria from different growth source conditions belonged to the same LF89 prototype strain as originally described by Fryer in 1992. Notwithstanding, growth of the bacteria both in cell-free media as well as in tissue culture cell lines were definitively suboptimal, and much more analysis are required to fully understand the biology of the pathogen. Interestingly, we were able to grow the bacteria in liquid media but at very slow rate. We conclude that, in addition to previous reports, our results confirmed the facultative intracellular nature of this fish pathogen.Piscirickettsia salmonis es un microorganismo altamente contagioso y virulento que afecta a la salmonicultura mundial desde el último tercio del siglo pasado y del cual sus mecanismos de sobrevivencia intracelular son completamente desconocidos. Después de algunos reportes recientes en donde se cuestiona su condición de intracelular obligado, hemos decidido mostrar evidencia adicional para cambiar este paradigma, llevando a cabo experimentos tanto clásicos como moleculares que confirman su naturaleza de intracelular facultativo. En

  5. Psychrotrophic bacteria and their negative effects on milk and dairy products quality

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    Šimun Zamberlin

    2012-06-01

    Full Text Available The characteristics of bacterial populations in raw milk at the time of processing has a significant influence on shelf-life, organoleptic quality, spoilage and yields of raw milk, processed milk as well as on the other dairy products. Unfortunately, cold and extended storage of raw milk, as a common practice in dairy sector today, favour the growth of psychrotrophic bacteria. Therefore, their count in the refrigerated milk is more than the ideal limit of 10 % of the mesophilic count. Psychrotrophic bacteria are generally able to form extracellular or intracellular thermo-resistant enzymes (proteases, lipases, phospolipases which can contribute to milk and dairy products spoilage. In addition, besides exhibiting spoilage features, some species belonging to the psychrotrops are considered as emerging pathogens that carry innate resistance to antibiotics or produce toxins. In sense of quality, psychrotrophic bacteria have become major problem for today’s dairy industry as leading cause in spoilage of cold-storage milk and dairy products. This review article focuses on the impact of psychrotrops on quality problems associated with raw milk as well as on th final dairy products. Means of controlling the dominant psychrotrophic species responsible for undesirable activities in milk and dairy products were also discussed.

  6. Facultative thermogenesis induced by carbohydrate

    DEFF Research Database (Denmark)

    Astrup, A; Bülow, J; Christensen, N J

    1986-01-01

    In addition to the obligatory thermogenesis due to processing and storage, carbohydrate ingestion is accompanied by a facultative thermogenesis mediated by catecholamines via beta-adrenoceptors. The anatomical origin of facultative thermogenesis has hitherto not been determined. The possible invo....... The facultative thermogenesis in skeletal muscle may be of importance for the regulation of body weight in humans....

  7. Psychrotrophic bacteria in milk: How much do we really know?

    Science.gov (United States)

    de Oliveira, Gislene B.; Favarin, Luciana; Luchese, Rosa H.; McIntosh, Douglas

    2015-01-01

    The occurrence of psychrotrophic bacteria in raw milk is studied worldwide due to the difficulties associated with controlling their growth during cold storage and the consequent negative effects upon fluid milk or dairy products. Among the psychrotrophic bacteria, the genus Pseudomonas (represented primarily by P. fluorescens) has been highlighted as the cause of numerous defects in dairy products. In light of its perceived predominance, this species has frequently been chosen as a model organism to assess the effects of psychrotrophic bacteria on milk or to evaluate the efficacy of control measures. However, recent findings derived from the application of molecular biological techniques have exposed a number of deficiencies in our knowledge of the biology of milk-associated psychrotrophs. Furthermore, it has been revealed that microbe to microbe communication plays a significant role in determining both the identities and the extent to which different groups of microbes develop during cold storage. The application of molecular identification methods has exposed errors in the classification of members of the genus Pseudomonas isolated from cold stored milk and has stimulated a reevaluation of the presumed status of P. fluorescens as the predominant milk-associated psychrotrophic species. This article presents a succinct review of data from studies on psychrotrophic bacteria in milk, some of which contest established theories in relation to the microbiology of cold stored raw milk, and poses the question: how much do we really know? PMID:26273245

  8. Psychrotrophic bacteria in milk: How much do we really know?

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    Gislene B. de Oliveira

    2015-06-01

    Full Text Available The occurrence of psychrotrophic bacteria in raw milk is studied worldwide due to the difficulties associated with controlling their growth during cold storage and the consequent negative effects upon fluid milk or dairy products. Among the psychrotrophic bacteria, the genus Pseudomonas (represented primarily by P. fluorescens has been highlighted as the cause of numerous defects in dairy products. In light of its perceived predominance, this species has frequently been chosen as a model organism to assess the effects of psychrotrophic bacteria on milk or to evaluate the efficacy of control measures. However, recent findings derived from the application of molecular biological techniques have exposed a number of deficiencies in our knowledge of the biology of milk-associated psychrotrophs. Furthermore, it has been revealed that microbe to microbe communication plays a significant role in determining both the identities and the extent to which different groups of microbes develop during cold storage. The application of molecular identification methods has exposed errors in the classification of members of the genus Pseudomonas isolated from cold stored milk and has stimulated a reevaluation of the presumed status of P. fluorescens as the predominant milk-associated psychrotrophic species. This article presents a succinct review of data from studies on psychrotrophic bacteria in milk, some of which contest established theories in relation to the microbiology of cold stored raw milk, and poses the question: how much do we really know?

  9. Psychrotrophic metal tolerant bacteria for mobilisation of metals in Antarctic waters

    Digital Repository Service at National Institute of Oceanography (India)

    Gonsalves, M.J.B.D.

    Cold tolerant psychrotrophic bacteria abound in the Antarctic waters. While Antarctic krills are known to concentrate heavy metals at ppm levels, psychrotrophic bacteria from Antarctic fresh and marine waters have been reported to tolerate them...

  10. Psychrotrophic ~Iydrolytic Bacteria from Antarctica &,Other I. Low Temperature Habitats

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    K.V. Ramana

    2000-04-01

    Full Text Available Samples of water, soil,llake sediments and blue-green algal mats from Antarctica were'processed for enumeration, isolation and screening of psychrotrophic hydrolytic bacteria. Amylolytic bacteria were preponderant (75 per cent in the blue-green algal htat samples. Protease, lipase, an1ylase and urerse producing/bacteria were also isolated from the samples. ,Biochemical characteristics indicated that the isolates ;mainly comprised Pseudomonas and Bacillus species. Proteases and lipases of antarctic bacterial strains preferably hydrolysed denatured protein substrate and water soluble monomeric synthetic lipid substrates, respectively.

  11. Culture and manipulation of insect facultative symbionts.

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    Pontes, Mauricio H; Dale, Colin

    2006-09-01

    Insects from many different taxonomic groups harbor maternally transmitted bacterial symbionts. Some of these associations are ancient in origin and obligate in nature whereas others originated more recently and are facultative. Previous research focused on the biology of ancient obligate symbionts with essential nutritional roles in their insect hosts. However, recent important advances in understanding the biology of facultative associations have been driven by the development of techniques for the culture, genetic modification and manipulation of facultative symbionts. In this review, we examine these available experimental techniques and illustrate how they have provided fascinating new insight into the nature of associations involving facultative symbionts. We also propose a rationale for future research based on the integration of genomics and experimentation.

  12. Facultative parthenogenesis discovered in wild vertebrates

    National Research Council Canada - National Science Library

    Booth, Warren; Smith, Charles F; Eskridge, Pamela H; Hoss, Shannon K; Mendelson, 3rd, Joseph R; Schuett, Gordon W

    2012-01-01

    Facultative parthenogenesis (FP)-asexual reproduction by bisexual species-has been documented in a variety of multi-cellular organisms but only recently in snakes, varanid lizards, birds and sharks...

  13. Detection of carotenoids in psychrotrophic bacteria by spectroscopic approach

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    Kirti Kushwaha

    2014-12-01

    Full Text Available The combination of Raman and Infrared spectroscopic signatures were used to find the different vibrational modes of individual carotenoid as their spectral fingerprint. Both have been previously demonstrated to be highly useful methodology for the identification and/or typing of microorganisms. In this study, we set out to evaluate whether these technologies could be applied to detect the presence of carotenoids in psychrotrophic bacterial isolates. FTIR and Raman spectra of four psychrotrophic bacteria viz. Kocuria rosea, K. turfanensis, Sanguibacter suarezii and Planococcus maritimus were examined during the investigation. FTIR spectra bands at 1653-1661cm-1 in different samples were assigned as part of chlorophyll, 1424-1426 cm-1 as -C-H- (CH2 bending vibration from methylene of carotenoids or lycopene, 1366-1367 cm-1 band as the -ionone ring of β-carotene due to the C-H, (–CH3 symmetrical bending. Interestingly, Raman spectra revealed intense Raman bands in the range of 1511-1530, 1153-1159 and 1003-1010 cm-1 representing bacterial carotenoids. We hypothesize the biosynthesis of carotenoid as adaptive strategy to cope up inhospitable cold environments of Leh and Ladakh. The strong, scattering bands by different isolates attributable to ν(C=C phase stretching, ν(C-C and δ(C-CH3 methyl components systems, which could be probably membrane-associated C50 carotenoids. Their high intensities are due to resonance enhancement. It can be concluded that Raman spectroscopy is a sensitive and convenient detection tool for typing of the bacterial biomarkers with less time consumption.

  14. Process of Waste Sludge Facultative Metabolism

    Institute of Scientific and Technical Information of China (English)

    李茵

    2001-01-01

    Laboratory- scale experiments were conducted to determine new technology of waste sludge facultative metabolism . 10-L laboratory-scale facultative reactors were operated during 24-hour sludge residence time (SRT) and in room temperature . Results show that the organic matter in waste sludge after hydrolysis acidification will be reduced by 75.39%, the removal rate of CODer above 85% . Advantage of the process is hydrolysis-acidification in ambient air temperature as there is no need for facilities to be sealed or heated. In addition, the sludge will be recycled into the wastewater treatment system and finally towards zero-discharge.

  15. The Pasteur effect in facultative anaerobic metazoa.

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    Schmidt, H; Kamp, G

    1996-05-15

    The existence and the regulatory mechanisms of the Pasteur effect in facultative anaerobic metazoa are discussed. There are three reasons for the controversy surrounding this phenomenon. 1) The different definitions of the Pasteur effect, 2) the antagonistic effect of metabolic depression and its species specific response to hypoxia, as well as 3) the laboratory-specific differences in the experimental procedures for analyzing the Pasteur effect and its regulation. This review aims to clarify the confusion about the existence of the Pasteur effect in facultative anaerobic metazoa and to offer possible molecular mechanisms.

  16. Psychrotrophic clostridia causing spoilage in cooked meat and poultry products.

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    Kalinowski, R M; Tompkin, R B

    1999-07-01

    Certain types of commercially produced noncured turkey breast and roast beef are precooked in situ, stored at 4 degrees C or below, and typically given use by dates of greater than 50 days. While of rare, sporadic occurrence, an unpleasant spoilage characterized by strong H2S odor and gas production has been observed in these products. This spoilage is due to the growth of psychrotrophic anaerobic sporeformers. Isolates from roast beef resemble Clostridium laramie while isolates from uncured turkey have been designated C. ctm for cooked turkey meat. The turkey breast isolates were characterized by temperature growth ranges, carbohydrate fermentations, and other biochemical reactions. Growth of all isolates was inhibited in broth media by 3.0% NaCl, 100 ppm nitrite, 2.0% sodium lactate, or 0.2% sodium diacetate. Inoculated studies were performed with three isolates in cooked turkey product. All three isolates grew and spoiled product at 10 and 3.3 degrees C, and one isolate grew at 0.5 and -3 degrees C. Some differences in growth were observed with the lactate and diacetate treatments in turkey meat among the three isolates. One isolate appeared to utilize the lactate, two were inhibited. Overall, 0.1% diacetate consistently delayed growth, although to different degrees, for all isolates.

  17. Facultative hyperaccumulation of heavy metals and metalloids.

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    Pollard, A Joseph; Reeves, Roger D; Baker, Alan J M

    2014-03-01

    Approximately 500 species of plants are known to hyperaccumulate heavy metals and metalloids. The majority are obligate metallophytes, species that are restricted to metalliferous soils. However, a smaller but increasing list of plants are "facultative hyperaccumulators" that hyperaccumulate heavy metals when occurring on metalliferous soils, yet also occur commonly on normal, non-metalliferous soils. This paper reviews the biology of facultative hyperaccumulators and the opportunities they provide for ecological and evolutionary research. The existence of facultative hyperaccumulator populations across a wide edaphic range allows intraspecific comparisons of tolerance and uptake physiology. This approach has been used to study zinc and cadmium hyperaccumulation by Noccaea (Thlaspi) caerulescens and Arabidopsis halleri, and it will be instructive to make similar comparisons on species that are distributed even more abundantly on normal soil. Over 90% of known hyperaccumulators occur on serpentine (ultramafic) soil and accumulate nickel, yet there have paradoxically been few experimental studies of facultative nickel hyperaccumulation. Several hypotheses suggested to explain the evolution of hyperaccumulation seem unlikely when most populations of a species occur on normal soil, where plants cannot hyperaccumulate due to low metal availability. In such species, it may be that hyperaccumulation is an ancestral phylogenetic trait or an anomalous manifestation of physiological mechanisms evolved on normal soils, and may or may not have direct adaptive benefits. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  18. 46 CFR 308.544 - Facultative binder, Form MA-315.

    Science.gov (United States)

    2010-10-01

    ... 46 Shipping 8 2010-10-01 2010-10-01 false Facultative binder, Form MA-315. 308.544 Section 308.544 Shipping MARITIME ADMINISTRATION, DEPARTMENT OF TRANSPORTATION EMERGENCY OPERATIONS WAR RISK INSURANCE War Risk Cargo Insurance Iii-Facultative War Risk Cargo Insurance § 308.544 Facultative binder, Form...

  19. Identification and biotechnological potential of psychrotrophic marine isolates

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    Swanandi Pote

    2014-06-01

    Full Text Available Background: Importance of microbial metabolites in food, detergent, pharmaceutical, nutraceutical and cosmetic industries has now been widely established. To fulfill the requirement of these industries, psychrophilic/psychrotrophic marine microbes are being explored. These microbes help in the production of metabolites that are active and stable at extreme physiological conditions. In correlation with this scenario, the present study reports identification of 14 bacterial isolates (BRI 32- BRI 45 from marine water samples (out of which 4 are Antarctic with emphasis on their biotechnologically important characters. Materials and Methods: Bacterial isolates were identified using 16S rRNA gene sequencing. Growth of the isolates under different physiological conditions of temperature (10°C to 45°C, pH (3-10 and concentration of NaCl (0-20% was studied. Further, the isolates were examined for their ability to produce i polyunsaturated fatty acids, ii industrially important enzymes and their potential to produce bio-surfactant. Results: 16S rRNA gene sequence analysis revealed that the isolates belonged to Halomonas, Brevibacillus, Kocuria and Oceanobacillus genera. Our results indicated that the isolates could grow over a wide range of physiological conditions of pH (3-10, temperature (10-45°C and NaCl concentration. Eight out of 14 isolates showed the presence of omega-3 and omega-6 fatty acids. BRI 34 was found to produce significant amounts of eicosapentaenoic acid (39.66%. Most of the isolates exhibited the ability to produce 3 or 4 enzymes. Only BRI 35 showed potential for biosurfactant production.

  20. Interaction of the psychrotroph Pseudomonas fluorescens In5 with phytopathogens in cold soils

    DEFF Research Database (Denmark)

    Hennessy, Rosanna Catherine; Olsson, Stefan; Stougaard, Peter

    is disease suppressive owing to the presence of diverse antimicrobial microorganisms. Using culture-based approaches, the psychrotroph Pseudomonas fluorescens In5 was previously isolated from the soil microbiome. The aim of this study is to unravel key biocontrol traits underpinning the antagonistic activity...... of this cold-active bacterium against phytopathogens. Method: A combination of different technologies including genomics, transcriptomics and metabolomics are being used to explore the interaction of the psychrotroph P. fluorescens In5 in dual-culture with diverse plant pathogens. To date, molecular genetics...

  1. Isolation and characterization of a novel violacein-like pigment producing psychrotrophic bacterial species Janthinobacterium svalbardensis sp. nov.

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    Ambrožič Avguštin, Jerneja; Žgur Bertok, Darja; Kostanjšek, Rok; Avguštin, Gorazd

    2013-04-01

    A bacterial strain designated JA-1, related to Janthinobacterium lividum, was isolated from glacier ice samples from the island Spitsbergen in the Arctic. The strain was tested for phenotypic traits and the most prominent appeared to be the dark red brown to black pigmentation different from the violet pigment of Janthinobacterium, Chromobacterium and Iodobacter. Phylogenetic analysis based on 16S rRNA gene sequences and DNA-DNA hybridization tests showed that strain JA-1 belongs to the genus Janthinobacterium but represents a novel lineage distinct from the two known species of this genus, J. lividum and Janthinobacterium agaricidamnosum. The DNA G + C content of strain JA-1 was determined to be 62.3 mol %. The isolate is a psychrotrophic Gram negative bacterium, rod-shaped with rounded ends, containing intracellular inclusions and one polar flagellum. On the basis of the presented results strain JA-1 is proposed as the type strain of a novel species of the genus Janthinobacterium, for which the name Janthinobacterium svalbardensis sp. nov. is proposed (JA-1(T) = DSM 25734, ZIM B637).

  2. The Francisella intracellular life cycle: towards molecular mechanisms of intracellular survival and proliferation

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    Audrey eChong

    2010-12-01

    Full Text Available The tularemia-causing bacterium Francisella tularensis is a facultative intracellular organism with a complex intracellular lifecycle that ensures its survival and proliferation in a variety of mammalian cell types, including professional phagocytes. Because this cycle is essential to Francisella pathogenesis and virulence, much research has focused on deciphering the mechanisms of its intracellular survival and replication and characterizing both bacterial and host determinants of the bacterium’s intracellular cycle. Studies of various strains and host cell models have led to the consensual paradigm of Francisella as a cytosolic pathogen, but also to some controversy about its intracellular cycle. In this review, we will detail major findings that have advanced our knowledge of Francisella intracellular survival strategies and also attempt to reconcile discrepancies that exist in our molecular understanding of the Francisella-phagocyte interaction.

  3. Facultative parthenogenesis discovered in wild vertebrates

    Science.gov (United States)

    Booth, Warren; Smith, Charles F.; Eskridge, Pamela H.; Hoss, Shannon K.; Mendelson, Joseph R.; Schuett, Gordon W.

    2012-01-01

    Facultative parthenogenesis (FP)—asexual reproduction by bisexual species—has been documented in a variety of multi-cellular organisms but only recently in snakes, varanid lizards, birds and sharks. Unlike the approximately 80 taxa of unisexual reptiles, amphibians and fishes that exist in nature, FP has yet to be documented in the wild. Based on captive documentation, it appears that FP is widespread in squamate reptiles (snakes, lizards and amphisbaenians), and its occurrence in nature seems inevitable, yet the task of detecting FP in wild individuals has been deemed formidable. Here we show, using microsatellite DNA genotyping and litter characteristics, the first cases of FP in wild-collected pregnant females and their offspring of two closely related species of North American pitviper snakes—the copperhead (Agkistrodon contortrix) and cottonmouth (Agkistrodon piscivorus). Our findings support the view that non-hybrid origins of parthenogenesis, such as FP, are more common in squamates than previously thought. With this confirmation, FP can no longer be viewed as a rare curiosity outside the mainstream of vertebrate evolution. Future research on FP in squamate reptiles related to proximate control of induction, reproductive competence of parthenogens and population genetics modelling is warranted. PMID:22977071

  4. Hydrolytic potential of a psychrotrophic Pseudomonas isolated from refrigerated raw milk

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    Ana Paula F. Corrêa

    2011-12-01

    Full Text Available The production of extracellular hydrolases by a psychrotrophic bacterium isolated from refrigerated raw milk, and identified as a Pseudomonas sp. belonging to the Pseudomonas jenssenii group, was studied. This bacterium produced proteolytic and lipolytic enzymes in all media investigated (skim milk, cheese whey, casein broth, and tryptone soy broth. High levels of α-glucosidase were produced in skim milk broth. Hydrolytic enzymes detected in skim milk broth are of particular concern, indicating that these enzymes could be produced by Pseudomonas sp. during the cold storage of raw milk, contributing to the spoilage problem in milk and dairy products.

  5. Biodegradation of Variable-Chain-Length Alkanes at Low Temperatures by a Psychrotrophic Rhodococcus sp.

    OpenAIRE

    Whyte, Lyle G; Hawari, Jalal; Zhou, Edward; Bourbonnière, Luc; Inniss, William E.; Charles W Greer

    1998-01-01

    The psychrotroph Rhodococcus sp. strain Q15 was examined for its ability to degrade individual n-alkanes and diesel fuel at low temperatures, and its alkane catabolic pathway was investigated by biochemical and genetic techniques. At 0 and 5°C, Q15 mineralized the short-chain alkanes dodecane and hexadecane to a greater extent than that observed for the long-chain alkanes octacosane and dotriacontane. Q15 utilized a broad range of aliphatics (C10 to C21 alkanes, branched alkanes, and a substi...

  6. Mesophilic and psychrotrophic bacteria from meat and their spoilage potential in vitro and in beef.

    Science.gov (United States)

    Ercolini, Danilo; Russo, Federica; Nasi, Antonella; Ferranti, Pasquale; Villani, Francesco

    2009-04-01

    Mesophilic and psychrotrophic populations from refrigerated meat were identified in this study, and the spoilage potential of microbial isolates in packaged beef was evaluated by analyzing the release of volatile organic compounds (VOC) by gas chromatography-mass spectrometry (GC/MS). Fifty mesophilic and twenty-nine psychrotrophic isolates were analyzed by random amplified polymorphic DNA-PCR, and representative strains were identified by 16S rRNA gene sequencing. Carnobacterium maltaromaticum and C. divergens were the species most frequently found in both mesophilic and psychrotrophic populations. Acinetobacter baumannii, Buttiauxella spp. and Serratia spp. were identified among the mesophilic isolates, while Pseudomonas spp. were commonly identified among the psychrotrophs. The isolates were further characterized for their growth at different temperatures and their proteolytic activity in vitro on meat proteins extracts at 7 degrees C. Selected proteolytic strains of Serratia proteamaculans, Pseudomonas fragi, and C. maltaromaticum were used to examine their spoilage potential in situ. Single strains of these species and mixtures of these strains were used to contaminate beef chops that were packed and stored at 7 degrees C. At time intervals up to 1 month, viable counts were determined, and VOC were identified by GC/MS. Generally, the VOC concentrations went to increase during the storage of the contaminated meats, and the profiles of the analyzed meat changed dramatically depending on the contaminating microbial species. About 100 volatiles were identified in the different contaminated samples. Among the detected volatiles, some specific molecules were identified only when the meat was contaminated by a specific microbial species. Compounds such as 2-ethyl-1-hexanol, 2-buten-1-ol, 2-hexyl-1-octanol, 2-nonanone, and 2-ethylhexanal were detectable only for C. maltaromaticum, which also produced the highest number of aldehydes, lactones, and sulfur compounds. The

  7. Effect of lactic acid concentration on growth on meat of Gram-negative psychrotrophs from a meatworks.

    OpenAIRE

    Gill, C O; Newton, K G

    1982-01-01

    The inhibitory effect of the lactic acid in meat on gram-negative psychrotrophs appears to be due mainly to the decrease in pH, not to action of the undissociated acid. Species of Pseudomonas were essentially unaffected by the pH of normal meat. Other gram-negative psychrotrophs isolated from a meatworks included a large number of strains which would not grow on meat of normal pH at chill temperatures. Raising either the pH or the incubation temperature allowed many of the pH-sensitive strain...

  8. Production of acylated homoserine lactones by psychrotrophic members of the Enterobacteriaceae isolated from foods

    DEFF Research Database (Denmark)

    Gram, Lone; Christensen, A.B.; Flodgaard, Lars

    1999-01-01

    Bacteria are able to communicate and gene regulation can be mediated through the production of acylated homoserine lactone (AHL) signal molecules. These signals play important roles in several pathogenic and symbiotic bacteria. The following study was undertaken to investigate whether AHLs...... are produced by bacteria found in food at temperatures and NaCl conditions commercially used for food preservation and storage. A minimum of 116 of 154 psychrotrophic Enterobacteriaceae strains isolated from cold-smoked salmon or vacuum-packed chilled meat produced AHLs. Analysis by thin-layer chromatography...... of indigenous Enterobacteriaceae reached 106 CFU/g. This level of Enterobacteriaceae is often found in lightly preserved foods, and AHL-mediated gene regulation may play a role in bacteria associated with food spoilage or food toxicity...

  9. Palatal Actinomycosis and Kaposi Sarcoma in an HIV-Infected Subject with Disseminated Mycobacterium avium-intracellulare Infection

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    Yuria Ablanedo-Terrazas

    2012-01-01

    Full Text Available Actinomyces and Mycobacterium avium-intracellulare are facultative intracellular organisms, members of the bacterial order actinomycetales. Although Actinomyces can behave as copathogen when anatomic barriers are compromised, its coinfection with Mycobacterium avium-intracellulare has not previously been reported. We present the first reported case of palatal actinomycosis co-infection with disseminated MAC, in an HIV-infected subject with Kaposi sarcoma and diabetes. We discuss the pathogenesis of the complex condition of this subject.

  10. Optimization of nonribosomal peptides production by a psychrotrophic fungus: Trichoderma velutinum ACR-P1.

    Science.gov (United States)

    Sharma, Richa; Singh, Varun P; Singh, Deepika; Yusuf, Farnaz; Kumar, Anil; Vishwakarma, Ram A; Chaubey, Asha

    2016-11-01

    Trichoderma is an anamorphic filamentous fungal genus with immense potential for production of small valuable secondary metabolites with indispensable biological activities. Microbial dynamics of a psychrotrophic strain Trichoderma velutinum ACR-P1, isolated from unexplored niches of the Shiwalik region, bestowed with rich biodiversity of microflora, was investigated for production of nonribosomal peptides (NRPs) by metabolite profiling by intact-cell mass spectrometry (ICMS) employing matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometer. Being the first report on NRPs production by T. velutinum, studies on optimization of growth conditions by Response Surface Methodology (RSM) for production of NRPs by ACR-P1 was carried out strategically. Multifold enhancement in the yield of NRPs belonging to subfamily SF4 with medium chain of amino acid residues having m/z 1437.9, 1453.9, and 1452.0 at pH 5.9 at 20 °C and of subfamily SF1 with long-chain amino acid residues having m/z 1770.2, 1784.2, 1800.1, 1802.1, and 1815.1 was achieved at pH 7.0 at 25 °C. Complexities of natural mixtures were thus considerably reduced under respective optimized culture conditions accelerating the production of novel microbial natural products by saving time and resources.

  11. Characterization of a cloned subtilisin-like serine proteinase from a psychrotrophic Vibrio species.

    Science.gov (United States)

    Arnórsdottir, Jóhanna; Smáradóttir, Rúna B; Magnússon, Olafur Th; Thorbjarnardóttir, Sigrídur H; Eggertsson, Gudmundur; Kristjánsson, Magnús M

    2002-11-01

    The gene encoding a subtilisin-like serine proteinase in the psychrotrophic Vibrio sp. PA44 has been successfully cloned, sequenced and expressed in Escherichia coli. The gene is 1593 basepairs and encodes a precursor protein of 530 amino acid residues with a calculated molecular mass of 55.7 kDa. The enzyme is isolated, however, as an active 40.6-kDa proteinase, without a 139 amino acid residue N-terminal prosequence. Under mild conditions the enzyme undergoes a further autocatalytic cleavage to give a 29.7-kDa proteinase that retains full enzymatic activity. The deduced amino acid sequence of the enzyme has high homology to proteinases of the proteinase K family of subtilisin-like proteinases. With respect to the enzyme characteristics compared in this study the properties of the wild-type and recombinant proteinases are the same. Sequence analysis revealed that especially with respect to the thermophilic homologues, aqualysin I from Thermus aquaticus and a proteinase from Thermus strain Rt41A, the cold-adapted Vibrio-proteinase has a higher content of polar/uncharged amino acids, as well as aspartate residues. The thermophilic enzymes had a higher content of arginines, and relatively higher number of hydrophobic amino acids and a higher aliphatic index. These factors may contribute to the adaptation of these proteinases to different temperature conditions.

  12. Bioprospecting of plant growth promoting psychrotrophic Bacilli from the cold desert of north western Indian Himalayas.

    Science.gov (United States)

    Yadav, Ajar Nath; Sachan, Shashwati Ghosh; Verma, Priyanka; Saxena, Anil Kumar

    2016-02-01

    The plant growth promoting psychrotrophic Bacilli were investigated from different sites in north western Indian Himalayas. A total of 247 morphotypes were obtained from different soil and water samples and were grouped into 43 clusters based on 16S rDNA-RFLP analysis with three restriction endonucleases. Sequencing of representative isolates has revealed that these 43 Bacilli belonged to different species of 11 genera viz., Desemzia, Exiguobacterium, Jeotgalicoccus, Lysinibacillus, Paenibacillus, Planococcus, Pontibacillus, Sinobaca, Sporosarcina, Staphylococcus and Virgibacillus. With an aim to develop microbial inoculants that can perform efficiently at low temperatures, all representative isolates were screened for different plant growth promoting traits at low temperatures (5-15 degrees C). Among the strains, variations were observed for production (%) of indole-3-acetic acid (20), ammonia (19), siderophores (11), gibberellic acid (4) and hydrogen cyanide (2); solubilisation (%) of zinc (14), phosphate (13) and potassium (7); 1-aminocyclopropane-1-carboxylate deaminase activity (6%) and biocontrol activity (4%) against Rhizoctonia solani and Macrophomina phaseolina. Among all the strains, Bacillus licheniformis, Bacillus muralis, Desemzia incerta, Paenibacillus tylopili and Sporosarcina globispora were found to be potent candidates to be developed as inoculants as they exhibited multiple PGP traits at low temperature.

  13. Cultivation conditions and properties of extracellular crude lipase from the psychrotrophic fungus Penicillium chrysogenum 9'.

    Science.gov (United States)

    Bancerz, R; Ginalska, G; Fiedurek, J; Gromada, A

    2005-06-01

    Among 97 fungal strains isolated from soil collected in the arctic tundra (Spitsbergen), Penicillium chrysogenum 9' was found to be the best lipase producer. The maximum lipase activity was 68 units mL(-1) culture medium on the fifth day of incubation at pH 6.0 and 20 degrees C. Therefore, P. chrysogenum 9' was classified as a psychrotrophic microorganism. The non-specific extracellular lipase showed a maximum activity at 30 degrees C and pH 5.0 for natural oils or at pH 7.0 for synthetic substrates. Tributyrin was found to be the best substrate for lipase, among those tested. The Km and Vmax were calculated to be 2.33 mM and 22.1 units mL(-1), respectively, with tributyrin as substrate. The enzyme was inhibited more by EDTA than by phenylmethylsulfonyl fluoride and was reactivated by Ca2+. The P. chrysogenum 9' lipase was very stable in the presence of hexane and 1,4-dioxane at a concentration of 50%, whereas it was unstable in presence of xylene.

  14. Description of Pseudomonas gregormendelii sp. nov., a Novel Psychrotrophic Bacterium from James Ross Island, Antarctica.

    Science.gov (United States)

    Kosina, Marcel; Švec, Pavel; Černohlávková, Jitka; Barták, Miloš; Snopková, Kateřina; De Vos, Paul; Sedláček, Ivo

    2016-07-01

    During the microbiological research performed within the scope of activities of Czech expeditions based at the Johann Gregor Mendel Station at James Ross Island, Antarctica, two psychrotrophic gram-stain negative non-fluorescent strains CCM 8506T and CCM 8507 from soil were extensively characterized using genotypic and phenotypic methods. Initial characterization using ribotyping with HindIII restriction endonuclease and phenotyping implies that both isolates belong to a single Pseudomonas species. Sequencing of rrs, rpoB, rpoD and glnA genes of strain CCM 8506(T) confirmed affiliation of investigated strains within the genus Pseudomonas. Further investigation using automated ribotyping with EcoRI (RiboPrinter(®) Microbial Characterisation System), whole-cell protein profiling using the Agilent 2100 Bioanalyzer system, extensive biochemical testing and DNA-DNA hybridization experiments confirmed that both investigated strains are members of a single taxon which is clearly separated from all hitherto described Pseudomonas spp. Based on all findings, we describe a novel species Pseudomonas gregormendelii sp. nov. with the type strain CCM 8506(T) (=LMG 28632T).

  15. Microbiological Hazards Assessment of Psychrotrophic Microflora in Bovine Carcasses Slaughtered in North West Romania

    Directory of Open Access Journals (Sweden)

    Silviu Vida

    2016-11-01

    Full Text Available Gram negative bacteria have the greatest capacity to spoil the meat if kept under aerobic conditions, therefor the members of the genera Pseudomonas, Acinetobater, Psychrobacter and Moraxella, will form the dominant microflora. The purpose of this study was to assess the contamination level with psychrotropic microflora of bovine carcasses in order to prove how the results could be used to improve the slaughter process. The research was carried out between January-December 2015, in two slaughterhouses from North West Romania.  The research material was represented by 144 meat samples (slaughterhouse A, n=72, slaughterhouse B, n=72. Weekly, 3 samples were collected from refrigerated carcasses, and examined for total psychrotrophic counts (TPC, Enterobacteriaceae, Pseudomonas, Aeromonas and Yersinia. From statistical processing of recorded data was established that mean log TPC from the surface of refrigerated carcasses has presented different values, ranged between 3.70±0.20 log CFU/cm2 and 6.90±0.43 log CFU/cm2. Initial surface microflora of bovine carcasses was represented by germs from the following genera: Staphylococcus, Micrococcus, Aeromonas, Acinetobacter, Pseudomonas, Yersinia, Serratia, Hafnia, Proteus and Escherichia. Microbiological hazard assessment reveals the key role of psychotropic microorganisms in the spoilage of meat, if the monitoring system of the slaughtering process is not functioning properly.

  16. Biodegradation of variable-chain-length alkanes at low temperatures by a psychrotrophic Rhodococcus sp.

    Energy Technology Data Exchange (ETDEWEB)

    Whyte, L.G.; Hawari, J.; Zhou, E.; Bourbonniere, L.; Greer, C.W. [NRC-Biotechnology Research Inst., Montreal, Quebec (Canada); Inniss, W.E. [Univ. of Waterloo, Ontario (Canada). Dept. of Biology

    1998-07-01

    The psychrotroph Rhodococcus sp. strain Q15 was examined for its ability to degrade individual n-alkanes and diesel fuel at low temperatures, and its alkane catabolic pathway was investigated by biochemical and genetic techniques. At 0 and 5 C, Q15 mineralized the short-chain alkanes dodecane and hexadecane to a greater extent than that observed for the long-chain alkanes octacosane and dotriacontane. Q15 utilized a broad range of aliphatics (C{sub 10} to C{sub 21} alkanes, branched alkanes, and a substituted cyclohexane) present in diesel fuel at 5 C. Mineralization of hexadecane at 5 C was significantly greater in both hydrocarbon-contaminated and pristine soil microcosms seeded with Q15 cells than in uninoculated control soil microcosms. The detection of hexadecane and dodecane metabolic intermediates (1-hexadecanol and 2-hexadecanol and 1-do-decanol and 2-dodecanone, respectively) by solid-phase microextraction-gas chromatography-mass spectrometry and the utilization of potential metabolic intermediates indicated that Q15 oxidizes alkanes by both the terminal oxidation pathway and the subterminal oxidation pathway. Genetic characterization by PCR and nucleotide sequence analysis indicated that Q15 possesses an aliphatic aldehyde dehydrogenase gene highly homologous to the Rhodococcus erythropolis thcA gene. Rhodococcus sp. strain Q15 possessed two large plasmids of approximately 90 and 115 kb (shown to mediate Cd resistance) which were not required for alkane mineralization, although the 90-kb plasmid enhanced mineralization of some alkanes and growth on diesel oil at both 5 and 25 C.

  17. Facultative parthenogenesis in a critically endangered wild vertebrate.

    Science.gov (United States)

    Fields, Andrew T; Feldheim, Kevin A; Poulakis, Gregg R; Chapman, Demian D

    2015-06-01

    Facultative parthenogenesis - the ability of sexually reproducing species to sometimes produce offspring asexually - is known from a wide range of ordinarily sexually reproducing vertebrates in captivity, including some birds, reptiles and sharks [1-3]. Despite this, free-living parthenogens have never been observed in any of these taxa in the wild, although two free-living snakes were recently discovered each gestating a single parthenogen - one copperhead (Agkistrodon contortrix) and one cottonmouth (Agkistrodon piscivorus) [1]. Vertebrate parthenogens are characterized as being of the homogametic sex (e.g., females in sharks, males in birds) and by having elevated homozygosity compared to their mother [1-3], which may reduce their viability [4]. Although it is unknown if either of the parthenogenetic snakes would have been carried to term or survived in the wild, facultative parthenogenesis might have adaptive significance [1]. If this is true, it is reasonable to hypothesize that parthenogenesis would be found most often at low population density, when females risk reproductive failure because finding mates is difficult [5]. Here, we document the first examples of viable parthenogens living in a normally sexually reproducing wild vertebrate, the smalltooth sawfish (Pristis pectinata). We also provide a simple approach to screen any microsatellite DNA database for parthenogens, which will enable hypothesis-driven research on the significance of vertebrate parthenogenesis in the wild.

  18. Facultative social parasitism in the allodapine bee Macrogalea berentyensis

    Institute of Scientific and Technical Information of China (English)

    JACLYN A. SMITH

    2007-01-01

    Previous research on social parasitism has largely ignored allodapine socialparasites, which is surprising given the huge potential of these bees to provide a betterunderstanding of social parasitism. Macrogalea berentyensis, a species that was previouslysuggested to be a social parasite, was collected in nests of M. ellioti, and also in nestsconsisting of only M. berentyensis. These findings, along with morphological and phyloge-netic evidence, show that this species is a facultative social parasite. In the independentlyliving M. berentyensis nests, brood were present that had been reared to an advanced stage,suggesting that: (i) these parasites may be effective at foraging and caring for their brood;or (ii) these nests may be colonies where all the hosts had died, and these parasites had yetto disperse. Macrogalea berentyensis is the closest relative of the facultative social parasite,M. antanosy, and both these species represent the most recent evolutionary origin of socialparasitism within the allodapines. Further behavioral research on both these parasitic specieswould therefore have important implications for the understanding of the evolution of socialparasitism.

  19. 46 CFR 308.545 - Facultative cargo policy, Form MA-316.

    Science.gov (United States)

    2010-10-01

    ... 46 Shipping 8 2010-10-01 2010-10-01 false Facultative cargo policy, Form MA-316. 308.545 Section 308.545 Shipping MARITIME ADMINISTRATION, DEPARTMENT OF TRANSPORTATION EMERGENCY OPERATIONS WAR RISK... policy, Form MA-316. The standard form of War Risk Facultative Cargo Policy, Form MA-316, may be...

  20. Spores from mesophilic Bacillus cereus strains germinate better and grow faster in simulated gastro-intestinal conditions than spores from psychrotrophic strains.

    NARCIS (Netherlands)

    Wijnands, L M; Dufrenne, J B; Zwietering, M H; Leusden, F M van

    2006-01-01

    The species Bacillus cereus, known for its ability to cause food borne disease, consists of a large variety of strains. An important property for discrimination of strains is their growth temperature range. Psychrotrophic strains can grow well at refrigerator temperatures but grow at 37 degrees C wi

  1. Spores from mesophilic Bacillus cereus strains germinate better and grow faster in simulated gastro-intestinal conditions than spores from psychrotrophic strains

    NARCIS (Netherlands)

    Wijnands, L.M.; Dufrenne, J.B.; Zwietering, M.H.; Leusden, van F.M.

    2006-01-01

    The species Bacillus cereus, known for its ability to cause food borne disease, consists of a large variety of strains. An important property for discrimination of strains is their growth temperature range. Psychrotrophic strains can grow well at refrigerator temperatures but grow at 37 °C with diff

  2. The Evolution of Facultative Conformity Based on Similarity

    Science.gov (United States)

    Efferson, Charles; Lalive, Rafael; Cacault, Maria Paula; Kistler, Deborah

    2016-01-01

    Conformist social learning can have a pronounced impact on the cultural evolution of human societies, and it can shape both the genetic and cultural evolution of human social behavior more broadly. Conformist social learning is beneficial when the social learner and the demonstrators from whom she learns are similar in the sense that the same behavior is optimal for both. Otherwise, the social learner’s optimum is likely to be rare among demonstrators, and conformity is costly. The trade-off between these two situations has figured prominently in the longstanding debate about the evolution of conformity, but the importance of the trade-off can depend critically on the flexibility of one’s social learning strategy. We developed a gene-culture coevolutionary model that allows cognition to encode and process information about the similarity between naive learners and experienced demonstrators. Facultative social learning strategies that condition on perceived similarity evolve under certain circumstances. When this happens, facultative adjustments are often asymmetric. Asymmetric adjustments mean that the tendency to follow the majority when learners perceive demonstrators as similar is stronger than the tendency to follow the minority when learners perceive demonstrators as different. In an associated incentivized experiment, we found that social learners adjusted how they used social information based on perceived similarity, but adjustments were symmetric. The symmetry of adjustments completely eliminated the commonly assumed trade-off between cases in which learners and demonstrators share an optimum versus cases in which they do not. In a second experiment that maximized the potential for social learners to follow their preferred strategies, a few social learners exhibited an inclination to follow the majority. Most, however, did not respond systematically to social information. Additionally, in the complete absence of information about their similarity to

  3. Treatment of fresh poultry carcases with emulsions of glycerol monocaprate (monocaprin) to reduce contamination with Campylobacter and psychrotrophic bacteria.

    Science.gov (United States)

    Thormar, H; Hilmarsson, H; Thrainsson, J H; Georgsson, F; Gunnarsson, E; Dadadottir, S

    2011-02-01

    1. A previous study has shown that emulsions of monocaprin in citrate lactate buffer at pH 4·1-4·3 are highly active in killing Campylobacter in water, where they reduce viable bacterial counts by more than 6 log(10) colony forming units (cfu) in 1 min at a concentration of 1·25 mM (0·03%). 2. The present study was carried out to evaluate whether monocaprin emulsions could be used to kill Campylobacter on raw poultry. 3. It was shown that immersion of naturally contaminated chicken legs in 20 mM (0·5%) monocaprin emulsion at pH 4·1 for 1 min at 20°C reduced the number of Campylobacter by 2·0 to 2·7 log(10) cfu. Pre-chill dipping of whole carcases into 20 mM monocaprin emulsion in the slaughterhouse also caused a significant reduction in Campylobacter contamination. 4. Immersion in monocaprin emulsions at pH 4·1 was also assessed as a means to reduce the number of psychrotrophic spoilage bacteria. There were lower psychrotrophic bacteria counts on treated chicken parts than on untreated controls after storage at 3°C for up to 14 d. 5. Immersion in emulsions of monocaprin, which is a natural lipid classified as GRAS, may be a feasible method to reduce the number of Campylobacter and spoilage bacteria on raw poultry. This method could reduce the risk of human exposure to Campylobacter, and at the same time increase the shelf-life of poultry products.

  4. Endozoicomonas Are Specific, Facultative Symbionts of Sea Squirts

    Science.gov (United States)

    Schreiber, Lars; Kjeldsen, Kasper U.; Funch, Peter; Jensen, Jeppe; Obst, Matthias; López-Legentil, Susanna; Schramm, Andreas

    2016-01-01

    Ascidians are marine filter feeders and harbor diverse microbiota that can exhibit a high degree of host-specificity. Pharyngeal samples of Scandinavian and Mediterranean ascidians were screened for consistently associated bacteria by culture-dependent and -independent approaches. Representatives of the Endozoicomonas (Gammaproteobacteria, Hahellaceae) clade were detected in the ascidian species Ascidiella aspersa, Ascidiella scabra, Botryllus schlosseri, Ciona intestinalis, Styela clava, and multiple Ascidia/Ascidiella spp. In total, Endozoicomonas was detected in more than half of all specimens screened, and in 25–100% of the specimens for each species. The retrieved Endozoicomonas 16S rRNA gene sequences formed an ascidian-specific subclade, whose members were detected by fluorescence in situ hybridization (FISH) as extracellular microcolonies in the pharynx. Two strains of the ascidian-specific Endozoicomonas subclade were isolated in pure culture and characterized. Both strains are chemoorganoheterotrophs and grow on mucin (a mucus glycoprotein). The strains tested negative for cytotoxic or antibacterial activity. Based on these observations, we propose ascidian-associated Endozoicomonas to be commensals, living off the mucus continuously secreted into the pharynx. Members of the ascidian-specific Endozoicomonas subclade were also detected in seawater from the Scandinavian sampling site, which suggests acquisition of the symbionts by horizontal transmission. The combined results indicate a host-specific, yet facultative symbiosis between ascidians and Endozoicomonas. PMID:27462299

  5. Cellulase production by pink pigmented facultative methylotrophic strains (PPFMs).

    Science.gov (United States)

    Jayashree, Shanmugam; Lalitha, Rajendran; Vadivukkarasi, Ponnusamy; Kato, Yuko; Seshadri, Sundaram

    2011-07-01

    Pink pigmented facultative methylotrophs (PPFM) isolated from water samples of Cooum and Adyar rivers in Chennai and soil samples of forests located in various districts of Tamil Nadu, India were screened for cellulase production using carboxymethylcellulose agar (CMC agar) medium. The strains showed wide variations in the production of clearing zones around the colonies on CMC agar medium flooded with Congo red. CMCase and filter paper assays were used to quantitatively measure the cellulase activity of 13 PPFM strains. Among the strains, Methylobacterium gregans, MNW 60, MHW 109, MSF 34, and MSF 40 showed cellulolytic activity ranging from 0.73 to 1.16 U mL(-1) with wide temperature (35-65°C) and pH (5 to 8) tolerance. SDS-PAGE analysis of the crude enzyme of PPFM strain MNW 60 exhibited several protein bands, and zymogram analysis revealed two dimeric cellulase bands with molecular mass of ~92 and 42 kDa. Scanning electron microscopic studies revealed significant morphological differences between the cells grown in normal and CMC amended medium. The strain MNW 60 was identified as Methylobacterium sp. based on biochemical, physiological, and morphological analyses, and the methylotrophic nature was authenticated by the presence of mxaF gene, encoding methanol dehydrogenase as a key indicator enzyme of methylotrophs, with 99% similarity to Methylobacterium lusitanum. With the 16S ribosomal RNA sequence showing 97% similarity to M. lusitanum strain MP2, this can be proposed as a novel taxon of the genus Methylobacterium. The study forms the first detailed report on the extracellular cellulase production by pink pigmented Methylobacterium sp., and it is expected that this might be the basis for further studies on cellulase production by PPFMs to explore the molecular mechanism, strain improvement, and large-scale cellulase production for its application.

  6. Effect of ADH II Deficiency on the Intracellular Redox Homeostasis in Zymomonas mobilis

    Directory of Open Access Journals (Sweden)

    Nina Galinina

    2012-01-01

    Full Text Available Mutant strain of the facultatively anaerobic, ethanol-producing bacterium Zymomonas mobilis, deficient in the Fe-containing alcohol dehydrogenase isoenzyme (ADH II, showed impaired homeostasis of the intracellular NAD(PH during transition from anaerobic to aerobic conditions, and also in steady-state continuous cultures at various oxygen supplies. At the same time, ADH II deficiency in aerobically grown cells was accompanied by a threefold increase of catalase activity and by about 50% increase of hydrogen peroxide excretion. It is concluded that ADH II under aerobic conditions functions to maintain intracellular redox homeostasis and to protect the cells from endogenous hydrogen peroxide.

  7. Effect of ADH II deficiency on the intracellular redox homeostasis in Zymomonas mobilis.

    Science.gov (United States)

    Galinina, Nina; Lasa, Zane; Strazdina, Inese; Rutkis, Reinis; Kalnenieks, Uldis

    2012-01-01

    Mutant strain of the facultatively anaerobic, ethanol-producing bacterium Zymomonas mobilis, deficient in the Fe-containing alcohol dehydrogenase isoenzyme (ADH II), showed impaired homeostasis of the intracellular NAD(P)H during transition from anaerobic to aerobic conditions, and also in steady-state continuous cultures at various oxygen supplies. At the same time, ADH II deficiency in aerobically grown cells was accompanied by a threefold increase of catalase activity and by about 50% increase of hydrogen peroxide excretion. It is concluded that ADH II under aerobic conditions functions to maintain intracellular redox homeostasis and to protect the cells from endogenous hydrogen peroxide.

  8. Psychrotrophic lactic acid bacteria associated with production batch recalls and sporadic cases of early spoilage in Belgium between 2010 and 2014.

    Science.gov (United States)

    Pothakos, Vasileios; Taminiau, Bernard; Huys, Geert; Nezer, Carine; Daube, Georges; Devlieghere, Frank

    2014-11-17

    Between 2010 and 2014 several spoilage cases in Belgium occurring in retail foodstuffs prior to the end of shelf-life have been reported to our laboratory. Overall, seven cases involved strictly psychrotrophic lactic acid bacteria (LAB) contamination in packaged and chilled-stored food products. The products derived either from recalls of entire production batches or as specimens of sporadic spoilage manifestations. Some of these samples were returned to the manufacturing companies by consumers who observed the alterations after purchasing the products. The products covered a wide range of foodstuffs (i.e. meat, dairy, vegetable, egg products and composite food) and denoted different spoilage defects. However, the microbiota determined by means of 16S rRNA gene high-throughput sequencing analysis underpin few LAB genera (i.e. Leuconostoc, Lactobacillus, Weissella and Lactococcus), which are frequently encountered nowadays as specific spoilage organisms (SSO) albeit overlooked by mesophilic enumeration methods due to their strictly psychrotrophic character. The present study confirms the spreading of psychrotrophic LAB in Belgian food processing environments leading to unexpected spoilage, corroborating their spoilage dynamics and prevalence in all kinds of packaged and refrigerated foodstuffs in Northern Europe. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Psychrotrophic members of Leuconostoc gasicomitatum, Leuconostoc gelidum and Lactococcus piscium dominate at the end of shelf-life in packaged and chilled-stored food products in Belgium.

    Science.gov (United States)

    Pothakos, Vasileios; Snauwaert, Cindy; De Vos, Paul; Huys, Geert; Devlieghere, Frank

    2014-05-01

    Previously, a considerable underestimation (+0.5-3.2 log CFU/g) on the contamination levels of psychrotrophic lactic acid bacteria (LAB) was observed for 33 retail, packaged food products stored at chilling temperature when the mesophilic enumeration technique was implemented as reference shelf-life parameter. In the present study, the microbial diversity of the dominant psychrotrophic LAB recovered after incubation of plates at 22 °C for 5 days was determined using a polyphasic taxonomic approach. A total of 212 LAB isolates were identified using a combination of rep-PCR fingerprinting, amplified fragment length polymorphism (AFLP) analysis and pheS gene sequencing. Leuconostoc gasicomitatum, Leuconostoc gelidum, Leuconostoc spp., Lactococcus piscium and Lactobacillus algidus proved to be the most competent and predominant species that may go undetected by the widely applied mesophilic enumeration protocols (ISO 4833:2003 and ISO 15214:1998). This study has assessed the interspecific variation among potential spoilage LAB, and highlights the significance of implementing a reference shelf-life parameter based on the enumeration of the total psychrotrophic bacterial load for industrial microbiological routine analyses.

  10. Intracellular Bacteria in Protozoa

    Science.gov (United States)

    Görtz, Hans-Dieter; Brigge, Theo

    Intracellular bacteria in humans are typically detrimental, and such infections are regarded by the patients as accidental and abnormal. In protozoa it seems obvious that many bacteria have coevolved with their hosts and are well adapted to the intracellular way of life. Manifold interactions between hosts and intracellular bacteria are found, and examples of antibacterial resistance of unknown mechanisms are observed. The wide diversity of intracellular bacteria in protozoa has become particularly obvious since they have begun to be classified by molecular techniques. Some of the bacteria are closely related to pathogens; others are responsible for the production of toxins.

  11. Properties of a subtilisin-like proteinase from a psychrotrophic Vibrio species comparison with proteinase K and aqualysin I.

    Science.gov (United States)

    Kristjánsson, M M; Magnússon, O T; Gudmundsson, H M; Alfredsson, G A; Matsuzawa, H

    1999-03-01

    An extracellular serine proteinase purified from cultures of a psychrotrophic Vibrio species (strain PA-44) belongs to the proteinase K family of the superfamily of subtilisin-like proteinases. The enzyme is secreted as a 47-kDa protein, but under mild heat treatment (30 min at 40 degrees C) undergoes autoproteolytic cleavage on the carboxyl-side of the molecule to give a proteinase with a molecular mass of about 36 kDa that apparently shares most of the enzymatic characteristics and the stability of the 47-kDa protein. In this study, selected enzymatic properties of the Vibrio proteinase were compared with those of the related proteinases, proteinase K and aqualysin I, as representative mesophilic and thermophilic enzymes, respectively. The catalytic efficiency (kcat/Km) for the amidase activity of the cold-adapted enzyme against succinyl-AAPF-p-nitroanilide was significantly higher than that of its mesophilic and thermophilic counterparts, especially when compared with aqualysin I. The stability of the Vibrio proteinase, both towards heat and denaturants, was found to be significantly lower than of either proteinase K or aqualysin I. One or more disulfide bonds in the psychrotrophic proteinase are important for the integrity of the active enzyme structure, as disulfide cleavage, either by reduction with dithiothreitol or by sulfitolysis, led to a loss in its activity. Under the same conditions, aqualysin I was also partially inactivated by dithiothreitol, but the activity of proteinase K was unaffected. The disulfides of either proteinase K or aqualysin I were not reactive towards sulfitolysis, except under denaturing conditions, while all disulfides of the Vibrio proteinase reacted in absence of a denaturant. The reactivity of the disulfides of the proteins as a function of denaturant concentration followed the order: Vibrio proteinase > proteinase K > aqualysin I. The same order of reactivity was also observed for the inactivation of the enzymes by H2O2

  12. The consequences of facultative sex in a prey adapting to predation.

    Science.gov (United States)

    Koch, H; Becks, L

    2017-01-01

    A species reproductive mode, along with its associated costs and benefits, can play a significant role in its evolution and survival. Facultative sexuality, being able to reproduce both sexually and asexually, has been deemed evolutionary favourable as the benefits of either mode may be fully realized. In fact, many studies have focused on identifying the benefits of sex and/or the forces selecting for increased rates of sex using facultative sexual species. The costs of either mode, however, can also have a profound impact on a population's evolutionary trajectory. Here, we used experimental evolution and fitness assays to investigate the consequences of facultative sexuality in prey adapting to predation. Specifically, we compared the adaptive response of algal prey populations exposed to constant rotifer predation and which had alternating cycles of asexual and sexual reproduction where sexual episodes were either facultative (sexual and asexual progeny simultaneously propagated) or obligate (only sexual progeny propagated). We found that prey populations with facultative sexual episodes reached a lower final relative fitness and suffered a greater trade-off in traits under selection, that is defence and competitive ability, as compared to prey populations with obligate sexual episodes. Our results suggest that costs associated with sexual reproduction (germination time) and asexual reproduction (selection interference) were amplified in the facultative sexual prey populations, leading to a reduction in the net advantage of sexuality. Additionally, we found evidence that the cost of sex was reduced in the obligate sexual prey populations because increased selection for sex was observed via the spontaneous production of sexual cells. These results show that certain costs associated with facultative sexuality can affect an organism's evolutionary trajectory.

  13. Oxygen regulated gene expression in facultatively anaerobic bacteria.

    Science.gov (United States)

    Unden, G; Becker, S; Bongaerts, J; Schirawski, J; Six, S

    1994-01-01

    In facultatively anaerobic bacteria such as Escherichia coli, oxygen and other electron acceptors fundamentally influence catabolic and anabolic pathways. E. coli is able to grow aerobically by respiration and in the absence of O2 by anaerobic respiration with nitrate, nitrite, fumarate, dimethylsulfoxide and trimethylamine N-oxide as acceptors or by fermentation. The expression of the various catabolic pathways occurs according to a hierarchy with 3 or 4 levels. Aerobic respiration at the highest level is followed by nitrate respiration (level 2), anaerobic respiration with the other acceptors (level 3) and fermentation. In other bacteria, different regulatory cascades with other underlying principles can be observed. Regulation of anabolism in response to O2 availability is important, too. It is caused by different requirements of cofactors or coenzymes in aerobic and anaerobic metabolism and by the requirement for different O2-independent biosynthetic routes under anoxia. The regulation mainly occurs at the transcriptional level. In E. coli, 4 global regulatory systems are known to be essential for the aerobic/anaerobic switch and the described hierarchy. A two-component sensor/regulator system comprising ArcB (sensor) and ArcA (transcriptional regulator) is responsible for regulation of aerobic metabolism. The FNR protein is a transcriptional sensor-regulator protein which regulates anaerobic respiratory genes in response to O2 availability. The gene activator FhlA regulates fermentative formate and hydrogen metabolism with formate as the inductor. ArcA/B and FNR directly respond to O2, FhlA indirectly by decreased levels of formate in the presence of O2. Regulation of nitrate/nitrite catabolism is effected by two 2-component sensor/regulator systems NarX(Q)/NarL(P) in response to nitrate/nitrite. Co-operation of the different regulatory systems at the target promoters which are in part under dual (or manifold) transcriptional control causes the expression

  14. DNA topology and adaptation of Salmonella typhimurium to an intracellular environment.

    Science.gov (United States)

    Marshall, D G; Bowe, F; Hale, C; Dougan, G; Dorman, C J

    2000-01-01

    The expression of genes coding for determinants of DNA topology in the facultative intracellular pathogen Salmonella typhimurium was studied during adaptation by the bacteria to the intracellular environment of J774A.1 macrophage-like cells. A reporter plasmid was used to monitor changes in DNA supercoiling during intracellular growth. Induction of the dps and spv genes, previously shown to be induced in the macrophage, was detected, as was expression of genes coding for DNA gyrase, integration host factor and the nucleoid-associated protein H-NS. The topA gene, coding for the DNA relaxing enzyme topoisomerase I, was not induced. Reporter plasmid data showed that bacterial DNA became relaxed following uptake of S. typhimurium cells by the macrophage. These data indicate that DNA topology in S. typhimurium undergoes significant changes during adaptation to the intracellular environment. A model describing how this process may operate is discussed. PMID:10874730

  15. Purification and characterization of an extracellular cold-adapted alkaline lipase produced by psychrotrophic bacterium Yersinia enterocolitica strain KM1.

    Science.gov (United States)

    Ji, Xiuling; Chen, Guiyuan; Zhang, Qi; Lin, Lianbing; Wei, Yunlin

    2015-06-01

    An extracellular cold-adapted alkaline lipase from the psychrotrophic Yersinia enterocolitica strain KM1 was purified 26-fold to homogeneity. The enzyme was active over a broad range spanning 0-60 °C with an optimum activity at 37 °C, and it was found to be alkaline-preferring with an optimum activity at pH 9.0. The molecular weight was estimated to be 34.3 KDa and monomeric. The lipase could be activated by Ca(2+) and low concentration (10%) of ethanol, dimethyl sulphoxide, methanol, and acetonitrile, whereas it was strongly inhibited by Zn(2+), Cu(2+), SDS, EDTA, and PMSF. Using p-nitrophenyl butyrate as a substrate at 37 °C, the Km and Vmax of the enzyme were found to be 16.58 mM and 5.24 × 10(5)  μM · min(-1), respectively. This extracellular cold-adapted alkaline lipase may be a good candidate for detergents and biocatalysts at low temperature.

  16. Extracellular cold active lipase from the psychrotrophic Halomonas sp. BRI 8 isolated from the Antarctic sea water

    Directory of Open Access Journals (Sweden)

    Vipra Vijay Jadhav

    2013-12-01

    Full Text Available An extracellular cold active lipase-producing psychrotrophic bacterium (BRI 8 was isolated from the Antarctic sea water sample. The 16s rRNA sequence study revealed that the isolate belongs to the genus Halomonas (929 bp. The present paper reports optimization of fermentation conditions for production of lipase (EC 3.1.1.3 from Halomonas sp. BRI 8. Highest lipase production was observed in the medium containing olive oil and peptone. The optimum pH and temperature for enzyme catalysis were 7.0 and 10°C respectively. The enzyme was relatively more stable in acidic pH range and retained 50% activity when incubated at 30°C for 1 h. The enzyme was stable in various organic solvents and showed more than 100% activity in presence of isoamyl alcohol. Significant enzyme activity was also observed in the presence of metal ions and detergents. The molecular mass of partially purified lipase was found to be around 66 kD.

  17. Lactococcus piscium: a psychrotrophic lactic acid bacterium with bioprotective or spoilage activity in food-a review.

    Science.gov (United States)

    Saraoui, T; Leroi, F; Björkroth, J; Pilet, M F

    2016-10-01

    The genus Lactococcus comprises 12 species, some known for decades and others more recently described. Lactococcus piscium, isolated in 1990 from rainbow trout, is a psychrotrophic lactic acid bacterium, probably disregarded because most of the strains are unable to grow at 30°C. During the last 10 years, this species has been isolated from a large variety of food: meat, seafood and vegetables, mostly packed under vacuum (VP) or modified atmosphere (MAP) and stored at chilled temperature. Recently, culture-independent techniques used for characterization of microbial ecosystems have highlighted the importance of Lc. piscium in food. Its role in food spoilage varies according to the strain and the food matrix. However, most studies have indicated that Lc. piscium spoils meat, whereas it does not degrade the sensory properties of seafood. Lactococcus piscium strains have a large antimicrobial spectrum, including Gram-positive and negative bacteria. In various seafoods, some strains have a protective effect against spoilage and can extend the sensory shelf-life of the products. They can also inhibit the growth of Listeria monocytogenes, by a cell-to-cell contact-dependent. This article reviews the physiological and genomic characteristics of Lc. piscium and discusses its spoilage or protective activities in food.

  18. Cold active hydrolytic enzymes production by psychrotrophic Bacilli isolated from three sub-glacial lakes of NW Indian Himalayas.

    Science.gov (United States)

    Yadav, Ajar Nath; Sachan, Shashwati Ghosh; Verma, Priyanka; Kaushik, Rajeev; Saxena, Anil Kumar

    2016-03-01

    The diversity of culturable, cold-active enzymes producing Bacilli was investigated from three sub-glacial lakes of north western Indian Himalayas. Amplified ribosomal DNA restriction analysis (ARDRA) using three restriction enzymes Alu I, Msp I, and Hae III led to the clustering of 136 Bacilli into 26, 23, and 22 clusters at 75% similarity index from Chandratal Lake, Dashair Lake, and Pangong Lake, respectively. Phylogenetic analysis based on 16S rRNA gene sequencing led to the identification of 35 Bacilli that could be grouped in seven families viz.: Bacillaceae (48%), Staphylococcaceae (14%), Bacillales incertae sedis (13%), Planococcaceae (12%), Paenibacillaceae (9%), Sporolactobacillaceae (3%), and Carnobacteriaceae (1%), which included twelve different genera Bacillus, Desemzia, Exiguobacterium, Jeotgalicoccus, Lysinibacillus, Paenibacillus, Planococcus, Pontibacillus, Sinobaca, Sporosarcina, Staphylococcus, and Virgibacillus. Based on their optimal temperature for growth, 35 Bacilli were grouped as psychrophilic (11 strains), psychrotrophic (17 strains), or psychrotolerant (7 strains), respectively. The representative isolates from each cluster were screened for cold-active enzyme activities. Amylase, β-glucosidase, pectinase, and protease activities at 4 °C were detected in more than 80% of the strains while approximately 40, 31, 23, 14, 11, and 9% of strains possessed cellulase, xylanase, β-galactosidase, laccase, chitinase, and lipase activity, respectively. Among 35 Bacilli, Bacillus amyloliquefaciens, Bacillus marisflavi, Exiguobacterium indicum, Paenibacillus terrae, Pontibacillus sp., Sporosarcina globispora, and Sporosarcina psychrophila were efficient producers of different cold-active enzymes. These cold-adapted Bacilli could play an important role in industrial and agricultural processes.

  19. Facultative cheating supports the coexistence of diverse quorum-sensing alleles.

    Science.gov (United States)

    Pollak, Shaul; Omer-Bendori, Shira; Even-Tov, Eran; Lipsman, Valeria; Bareia, Tasneem; Ben-Zion, Ishay; Eldar, Avigdor

    2016-02-23

    Bacterial quorum sensing enables bacteria to cooperate in a density-dependent manner via the group-wide secretion and detection of specific autoinducer molecules. Many bacterial species show high intraspecific diversity of autoinducer-receptor alleles, called pherotypes. The autoinducer produced by one pherotype activates its coencoded receptor, but not the receptor of another pherotype. It is unclear what selection forces drive the maintenance of pherotype diversity. Here, we use the ComQXPA system of Bacillus subtilis as a model system, to show that pherotype diversity can be maintained by facultative cheating--a minority pherotype exploits the majority, but resumes cooperation when its frequency increases. We find that the maintenance of multiple pherotypes by facultative cheating can persist under kin-selection conditions that select against "obligate cheaters" quorum-sensing response null mutants. Our results therefore support a role for facultative cheating and kin selection in the evolution of quorum-sensing diversity.

  20. Activity of 10 antimicrobial agents against intracellular Rhodococcus equi.

    Science.gov (United States)

    Giguère, Steeve; Berghaus, Londa J; Lee, Elise A

    2015-08-05

    Studies with facultative intracellular bacterial pathogens have shown that evaluation of the bactericidal activity of antimicrobial agents against intracellular bacteria is more closely associated with in vivo efficacy than traditional in vitro susceptibility testing. The objective of this study was to determine the relative activity of 10 antimicrobial agents against intracellular Rhodococcus equi. Equine monocyte-derived macrophages were infected with virulent R. equi and exposed to erythromycin, clarithromycin, azithromycin, rifampin, ceftiofur, gentamicin, enrofloxacin, vancomycin, imipenem, or doxycycline at concentrations achievable in plasma at clinically recommended dosages in foals. The number of intracellular R. equi was determined 48h after infection by counting colony forming units (CFUs). The number of R. equi CFUs in untreated control wells were significantly higher than those of monolayers treated with antimicrobial agents. Numbers of R. equi were significantly lower in monolayers treated with enrofloxacin followed by those treated with gentamicin, and vancomycin, when compared to monolayers treated with other antimicrobial agents. Numbers of R. equi in monolayers treated with doxycycline were significantly higher than those of monolayers treated with other antimicrobial agents. Differences in R. equi CFUs between monolayers treated with other antimicrobial agents were not statistically significant. Enrofloxacin, gentamicin, and vancomycin are the most active drugs in equine monocyte-derived macrophages infected with R. equi. Additional studies will be needed to determine if these findings correlate with in vivo efficacy. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. The outcome of Cryptococcus neoformans intracellular pathogenesis in human monocytes

    Directory of Open Access Journals (Sweden)

    Pirofski Liise-anne

    2009-03-01

    Full Text Available Abstract Background Cryptococcus neoformans is an encapsulated yeast that is a facultative intracellular pathogen. The interaction between macrophages and C. neoformans is critical for extrapulmonary dissemination of this pathogenic yeast. C. neoformans can either lyse macrophages or escape from within them through a process known as phagosomal extrusion. However, most studies of intracellular pathogenesis have been made with mouse cells and their relevance to human infection is uncertain. In this study we extended studies of C. neoformans-macrophage cellular interaction/s to human peripheral blood monocytes. Results This study demonstrated that C. neoformans can shed polysaccharide within human monocytes, spread from cell to cell, and be extruded from them. Furthermore, human monocytes responded to ingestion of C. neoformans with cell cycle progression from G1 to S. Conclusion Similarities between mouse and human cells support the suitability of mouse cells for the study of intracellular pathogenesis mechanisms. Given that these hosts diverged over 70 million years ago, the similar pathogenic strategies for C. neoformans in murine and human cells supports the hypothesis that the mechanism that underlies the mammalian intracellular pathogenesis of C. neoformans originated from interactions with a third host, possibly soil amoeboid predators, before the mammalian radiation.

  2. Hazard Analysis and Control of Psychrotrophic Bacterias and Heat Resistant Enzymes in Raw Milk and Dairy Products%乳和乳制品中嗜冷菌及其耐热酶的危害与控制研究

    Institute of Scientific and Technical Information of China (English)

    王娇; 张筠; 刘术明; 刘文丽

    2015-01-01

    嗜冷菌是乳低温贮藏中的主要腐败菌,嗜冷菌及其代谢酶的存在直接影响原料乳的质量,阐述了嗜冷菌及其耐热酶类对乳及乳制品的危害,并通过分析嗜冷菌的适冷机理和影响酶活的因素,探讨控制嗜冷菌及其代谢酶的方法,以保障原料乳质量。%Psychrotrophic bacteria is the primary putrefying bacteria in low-temperature storage raw milk , Psychrotrophic bacteria and the thermostable enzymes they made have enormous implications on raw milk ,in this article,the perniciousness Psychrotrophic bacteria and the thermostable enzymes they made to milk and dairy products are introduced,by analyze the mechanism how Psychrotrophic bacteria can live in low temperature and the factors that influence the enzymes' activities ,investigate the methods to control Psychrotrophic bacteria and the enzymes they produce to guarantee the quality of raw milk.

  3. Obligation of negotiation and facultative arbitration as part of constitutional law content on collective negotiation

    Directory of Open Access Journals (Sweden)

    Guillermo Boza Pró

    2013-12-01

    Full Text Available This work studies constitutional bases of two essential institutions of Collective Labor Law: employer’s obligation to negotiate collectively and facultative arbitration. In relation to the first one, using tools provided bythe Theory of Law, it is argued that the section 28º of Peruvian Constitution establishes a collective negotiation concept as claim (not as permission so, employers legal obligation of collective negotiation is constitutional. In theother hand, regarding facultative arbitration, it has constitutional support on state’s obligation to promote pacific ways to labor conflicts; even though, it is questioned that a «sub constitutional» rule has restrictively regulated this institution.

  4. Multilocus sequence typing of Leuconostoc gelidum subsp. gasicomitatum, a Psychrotrophic lactic acid bacterium causing spoilage of packaged perishable foods.

    Science.gov (United States)

    Rahkila, Riitta; Johansson, Per; Säde, Elina; Paulin, Lars; Auvinen, Petri; Björkroth, Johanna

    2015-04-01

    Leuconostoc gelidum subsp. gasicomitatum is a psychrotrophic lactic acid bacterium (LAB) that causes spoilage of a variety of modified-atmosphere-packaged (MAP) cold-stored food products. During the past 10 years, this spoilage organism has been increasingly reported in MAP meat and vegetable products in northern Europe. In the present study, the population structure within 252 L. gelidum subsp. gasicomitatum strains was determined based on a novel multilocus sequence-typing (MLST) scheme employing seven housekeeping genes. These strains had been isolated from meat and vegetable sources over a time span of 15 years, and all 68 previously detected pulsed-field gel electrophoresis (PFGE) genotypes were represented. A total of 46 sequence types (STs) were identified, with a majority of the strains (>60%) belonging to three major STs, which were grouped into three clonal complexes (CCs) and 17 singletons by Global Optimal eBURST (goeBURST). The results by Bayesian analysis of population structure (BAPS) mostly correlated with the grouping by goeBURST. Admixture analysis by BAPS indicated a very low level of exchange of genetic material between the subpopulations. Niche specificity was observed within the subpopulations: CC1 and BAPS cluster 1 consisted mostly of strains from a variety of MAP meats, whereas vegetable strains grouped together with strains from MAP poultry within CC2 and BAPS cluster 2. The MLST scheme presented in this study provides a shareable and continuously growing sequence database enabling global comparison of strains associated with spoilage cases. This will further advance our understanding of the microbial ecology of this industrially important LAB. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  5. Effects of sodium citrate plus sodium diacetate and buffered vinegar on Escherichia coli O157:H7 and psychrotrophic bacteria in brine-injected beef.

    Science.gov (United States)

    Ponrajan, Amudhan; Harrison, Mark A; Segers, Jacob R; Lowe, Bradley K; McKeith, Russell O; Pringle, T Dean; Martino, Karina G; Mulligan, Jake H; Stelzleni, Alexander M

    2011-03-01

    The objective of this research was to examine the effects of sodium citrate plus sodium diacetate or buffered vinegar on Escherichia coli O157:H7 and psychrotrophic bacteria when incorporated in brine solutions for injected beef. Two experiments were conducted in which 30 top rounds and 30 top sirloins were injected (110%) to contain (i) 0.5% sodium chloride and 0.4% sodium tripolyphosphate as the control (CNT); (ii) CNT with a 1% solution of 80% sodium citrate plus 20% sodium diacetate (SC + D); or (iii) CNT with 2% buffered vinegar (VIN) in the final product. For the E. coli challenge, muscles were surface inoculated to target 6 log CFU/cm(2). After injection and 10 days of storage in a vacuum package (4°C), one half of each muscle was sampled raw and the other half was cooked to an internal temperature of 60°C with a 12-min hold. For raw samples, a significant reduction of 0.6 and 1.0 log CFU/g of E. coli O157:H7 was observed in both SC + D- and VIN-injected top rounds and sirloins, respectively. All cooked samples were E. coli O157:H7 negative. For psychrotrophic analysis, subprimals were injected and vacuum packaged for 10 days at 0 ± 1°C. After 10 days of storage, steaks were fabricated and placed in aerobic display (4 ± 1°C) for 1, 7, 14, and 21 days. Psychrotrophic organism growth was restricted in SC + D and VIN samples when compared with CNT on all days except day 1. Sodium citrate plus sodium diacetate or buffered vinegar may improve the safety and shelf life of multineedle brine-injected beef.

  6. Global Attractivity of a Periodic Delayed N-Species Model of Facultative Mutualism

    Directory of Open Access Journals (Sweden)

    Ahmadjan Muhammadhaji

    2013-01-01

    Full Text Available Two classes of periodic N-species Lotka-Volterra facultative mutualism systems with distributed delays are discussed. Based on the continuation theorem of the coincidence degree theory developed by Gaines and Mawhin and the Lyapunov function method, some new sufficient conditions on the existence and global attractivity of positive periodic solutions are established.

  7. Facultative endosymbionts of aphid populations from coastal dunes of the North Sea

    NARCIS (Netherlands)

    Pena, de la E.; Vandomme, V.; Frago Clols, E.

    2014-01-01

    Aphids establish symbiotic associations with a diverse assemblage of mutualistic bacteria. Some of them are not required for the host’s survival but still have a crucial impact on the biology and ecology of their host. Facultative symbionts may modify important host-life-history traits and affect th

  8. Facultative endosymbionts of aphid populations from coastal dunes of the North Sea

    NARCIS (Netherlands)

    Pena, de la E.; Vandomme, V.; Frago Clols, E.

    2014-01-01

    Aphids establish symbiotic associations with a diverse assemblage of mutualistic bacteria. Some of them are not required for the host’s survival but still have a crucial impact on the biology and ecology of their host. Facultative symbionts may modify important host-life-history traits and affect th

  9. Faculté des sciences de Bordeaux, René-André Coulon architecte

    Directory of Open Access Journals (Sweden)

    Franck Delorme

    2012-01-01

    Full Text Available Au sortir de la seconde guerre mondiale, les facultés bordelaises étouffent dans leurs locaux du centre-ville édifiés sous la troisième République. Un ambitieux programme de transferts des établissements universitaires vers les banlieues de Pessac, Talence et Gradignan se met en place à partir des années 1950. L’architecte René-André Coulon (1908-1997 conçoit la faculté des sciences de Bordeaux, envisagée alors comme une véritable faculté « pour l’an 2000 ». Tout en disposant largement les différents bâtiments dans l’espace du campus, Coulon s’attache à leur donner un style original, tout empreint de rigueur et de rationalisme, mais qui montre également quelques écarts notables avec les normes régissant alors les constructions universitaires. Talence est la première réalisation universitaire de Coulon, qui travaillera par la suite sur des programmes fort différents, qu’il s’agisse du campus de HEC à Jouy-en-Josas ou de celui de la faculté des sciences de Paris-centre avec Édouard Albert.

  10. Facultative endosymbionts of aphid populations from coastal dunes of the North Sea

    NARCIS (Netherlands)

    Pena, de la E.; Vandomme, V.; Frago Clols, E.

    2014-01-01

    Aphids establish symbiotic associations with a diverse assemblage of mutualistic bacteria. Some of them are not required for the host’s survival but still have a crucial impact on the biology and ecology of their host. Facultative symbionts may modify important host-life-history traits and affect

  11. Non-oral gram-negative facultative rods in chronic periodontitis microbiota

    NARCIS (Netherlands)

    van Winkelhoff, Arie J; Rurenga, Patrick; Wekema-Mulder, Gepke J; Singadji, Zadnach; Rams, Thomas E

    2016-01-01

    OBJECTIVE: The subgingival prevalence of gram-negative facultative rods not usually inhabiting or indigenous to the oral cavity (non-oral GNFR), as well as selected periodontal bacterial pathogens, were evaluated by culture in untreated and treated chronic periodontitis patients. METHODS: Subgingiva

  12. Intracellular drug release nanosystems

    Directory of Open Access Journals (Sweden)

    Fenghua Meng

    2012-10-01

    Full Text Available In order to elicit therapeutic effects, many drugs including small molecule anticancer drugs, proteins, siRNA, and DNA have to be delivered and released into the specific cellular compartments typically the cytoplasm or nucleus of target cells. Intracellular environment-responsive nanosystems that exhibit good extracellular stability while rapidly releasing drugs inside cancer cells have been actively pursued for effective cancer therapy. Here, we highlight novel designs of smart nanosystems that release drugs in response to an intracellular biological signal of cancer cells such as acidic pH in endo/lysosomal compartments, enzymes in lysosomes, and redox potential in cytoplasm and the cell nucleus.

  13. Antibiotic multiresistance analysis of mesophilic and psychrotrophic Pseudomonas spp. isolated from goat and lamb slaughterhouse surfaces throughout the meat production process.

    Science.gov (United States)

    Lavilla Lerma, Leyre; Benomar, Nabil; Casado Muñoz, María del Carmen; Gálvez, Antonio; Abriouel, Hikmate

    2014-11-01

    The aim of this study was to investigate the phenotypic and genotypic antibiotic resistance profiles of pseudomonads isolated from surfaces of a goat and lamb slaughterhouse, which were representative of areas that are possible sources of meat contamination. Mesophilic (85 isolates) and psychrotrophic (37 isolates) pseudomonads identified at the species level generally were resistant to sulfamethoxazole, erythromycin, amoxicillin, ampicillin, chloramphenicol, trimethoprim, rifampin, and ceftazidime (especially mesophiles), as well as colistin and tetracycline (especially psychrotrophes). However, they generally were sensitive to ciprofloxacin, gentamicin, imipenem, and kanamycin regardless of species identity. Worryingly, in the present study, we found multidrug resistance (MDR) to up to 13 antibiotics, which was related to intrinsic and acquired resistance mechanisms. Furthermore, a link between various antimicrobial resistance genes was shown for beta-lactams and tetracycline, trimethoprim, and sulfonamides. The distribution and resistome-based analysis of MDR pseudomonads in different slaughterhouse zones indicated that the main sources of the identical or related pseudomonad strains were the animals (feet and wool) and the slaughterhouse environment, being disseminated from the beginning, or entrance environment, to the environment of the finished meat products. Those facts must be taken into consideration to avoid cross-contamination with the subsequent flow of mobile resistance determinants throughout all slaughterhouse zones and then to humans and the environment by the application of adequate practices of hygiene and disinfection measures, including those for animal wool and feet and also the entrance environment.

  14. Total mesophilic counts underestimate in many cases the contamination levels of psychrotrophic lactic acid bacteria (LAB) in chilled-stored food products at the end of their shelf-life.

    Science.gov (United States)

    Pothakos, Vasileios; Samapundo, Simbarashe; Devlieghere, Frank

    2012-12-01

    The major objective of this study was to determine the role of psychrotrophic lactic acid bacteria (LAB) in spoilage-associated phenomena at the end of the shelf-life of 86 various packaged (air, vacuum, modified-atmosphere) chilled-stored retail food products. The current microbiological standards, which are largely based on the total viable mesophilic counts lack discriminatory capacity to detect psychrotrophic LAB. A comparison between the total viable counts on plates incubated at 30 °C (representing the mesophiles) and at 22 °C (indicating the psychrotrophs) for 86 food samples covering a wide range - ready-to-eat vegetable salads, fresh raw meat, cooked meat products and composite food - showed that a consistent underestimation of the microbial load occurs when the total aerobic mesophilic counts are used as a shelf-life parameter. In 38% of the samples, the psychrotrophic counts had significantly higher values (+0.5-3 log CFU/g) than the corresponding total aerobic mesophilic counts. A total of 154 lactic acid bacteria, which were unable to proliferate at 30 °C were isolated. In addition, a further 43 with a poor recovery at this temperature were also isolated. This study highlights the potential fallacy of the total aerobic mesophilic count as a reference shelf-life parameter for chilled food products as it can often underestimate the contamination levels at the end of the shelf-life.

  15. Biodegradation of Acid Anthraquinone Dye in a Facultative-aerobic Process: Kinetics and Products

    Institute of Scientific and Technical Information of China (English)

    LI Yin; LI Zai-lei

    2009-01-01

    The transformation of an anthraquinone dye blue 324 in a facultative-aerobic (F-A) system was investigated. Kinetic parameter study showed that higher Vmax coupled with more recalcitrant chemical oxygen demand (COD) were found in the facuitative biofilm reactor (FBR) than in the aerobic reactor (AR). Results of the product analyses indicated that most of dye molecular could be facultatively broken down into simple intermediates, which would be further degraded under subsequent aerobic condition. The main metabolites in each reactor were detected by infrared (FT-IR) and high performance liquid chromatography and mass spectrometry (HPLC-MS). Comparison of the toxicities among the dye and its metabolites was conducted, surprisingly, the colorless intermediates from FBR possessed less inhibitory than original dye and the median effective luminescence concentration (EC50) in 15 min for aerobic effluent could not be detected, showing that hardly toxic products existed in the aerobic process effluent.

  16. Substrate preference, uptake kinetics and bioenergetics in a facultatively autotrophic, thermoacidophilic crenarchaeote.

    Science.gov (United States)

    Urschel, Matthew R; Hamilton, Trinity L; Roden, Eric E; Boyd, Eric S

    2016-05-01

    Facultative autotrophs are abundant components of communities inhabiting geothermal springs. However, the influence of uptake kinetics and energetics on preference for substrates is not well understood in this group of organisms. Here, we report the isolation of a facultatively autotrophic crenarchaeote, strain CP80, from Cinder Pool (CP, 88.7°C, pH 4.0), Yellowstone National Park. The 16S rRNA gene sequence from CP80 is 98.8% identical to that from Thermoproteus uzonensis and is identical to the most abundant sequence identified in CP sediments. Strain CP80 reduces elemental sulfur (S8°) and demonstrates hydrogen (H2)-dependent autotrophic growth. H2-dependent autotrophic activity is suppressed by amendment with formate at a concentration in the range of 20-40 μM, similar to the affinity constant determined for formate utilization. Synthesis of a cell during growth with low concentrations of formate required 0.5 μJ compared to 2.5 μJ during autotrophic growth with H2 These results, coupled to data indicating greater C assimilation efficiency when grown with formate as compared to carbon dioxide, are consistent with preferential use of formate for energetic reasons. Collectively, these results provide new insights into the kinetic and energetic factors that influence the physiology and ecology of facultative autotrophs in high-temperature acidic environments.

  17. Sex choice in plants: facultative adjustment of the sex ratio in the perennial herb Begonia gracilis.

    Science.gov (United States)

    López, S; Domínguez, C A

    2003-11-01

    Sex allocation theory predicts that reproducing individuals will increase their fitness by facultatively adjusting their relative investment towards the rarer sex in response to population shifts in operational sex ratio (OSR). The evolution of facultative manipulation of sex ratio depends on the ability of the parents to track the conditions favouring skewed sex allocation and on the mechanism controlling sex allocation. In animals, which have well-developed sensorial mechanisms, facultative adjustment of sex ratios has been demonstrated on many occasions. In this paper, we show that plants have mechanisms that allow them to evaluate the population OSR. We simulated three different conditions of population OSR by manipulating the amount of pollen received by the female flowers of a monoecious herb, and examined the effect of this treatment on the allocation to male vs. female flowers. A shortage of pollen on the stigmas resulted in a more male-skewed sex allocation, whereas plants that experienced a relatively pollen rich environment tended to produce a more female-skewed sex allocation pattern. Our results for Begonia gracilis demonstrate that the individuals of this species are able to respond to the levels of pollination intensity experienced by their female flowers and adjust their patterns of sex allocation in accordance to the expectations of sex allocation theory.

  18. Insights into the physiological responses of the facultative halophyte Aeluropus littoralis to the combined effects of salinity and phosphorus availability.

    Science.gov (United States)

    Talbi Zribi, Ons; Barhoumi, Zouhaier; Kouas, Saber; Ghandour, Mohamed; Slama, Ines; Abdelly, Chedly

    2015-09-15

    In this work, we investigate the physiological responses to P deficiency (5μM KH2PO4=D), salt stress (400mM NaCl=C+S), and their combination (D+S) on the facultative halophyte Aeluropus littoralis to understand how plants adapt to these combined stresses. When individually applied, both P deficiency and salinity significantly restricted whole plant growth, with a more marked effect of the latter stress. However, the effects of the two stresses were not additive in plant biomass production since the response of plants to combined salinity and P deficiency was similar to that of plants grown under salt stress alone. In addition the observed features under salinity alone are kept when plants are simultaneously subjected to the combined effects of salinity and P deficiency such as biomass partitioning; the synthesis of proline and the K(+)/Na(+) selectivity ratio. Thus, increasing P availability under saline conditions has no significant effect on salt tolerance in this species. Plants cultivated under the combined effects of salinity and P deficiency exhibited the lowest leaf water potential. This trend was associated with a high accumulation of Na(+), Cl(-) and proline in shoots of salt treated plants suggesting the involvement of these solutes in osmotic adjustment. Proline could be involved in other physiological processes such as free radical scavenging. Furthermore, salinity has no significant effect on phosphorus acquisition when combined with a low P supply and it significantly decreased this parameter when combined with a sufficient P supply. This fact was probably due to salt's effect on P transporters. In addition, shoot soluble sugars accumulation under both P deficiency treatments with and without salt likely play an important role in the adaptation of A. littoralis plants to P shortage applied alone or combined with salinity. Moreover, there was a strong correlation between shoot and root intracellular acid phosphatase activity and phosphorus use

  19. Environmental and intracellular regulation of Francisella tularensis ripA

    Directory of Open Access Journals (Sweden)

    Taft-Benz Sharon

    2009-10-01

    Full Text Available Abstract Background Francisella tularensis is a highly virulent, facultative intracellular pathogen and the etiologic agent of the zoonotic disease Tularemia. RipA is a cytoplasmic membrane protein that is conserved among Francisella species and is required for intracellular growth. F. tularensis ripA deletion mutants escape the phagosome of infected cells, but unlike wild type organisms fail to replicate in the host cell cytoplasm. Results Further analysis of ripA with respect to environmental effects on the growth of mutant strains and expression levels revealed that RipA is required for optimal growth at pH 7.5 but not pH 6.5. Using a combination of RT-PCR, ripA-lacZ transcriptional and translational fusions, and a RipA-tetracysteine tag fusion protein we found that both ripA transcription and RipA protein levels were elevated in organisms grown at pH 7.5 as compared to organisms grown at pH 5.5. A number of genes, including iglA, that are required for intracellular growth are regulated by the transcriptional regulators MglA and SspA, and are induced upon infection of host cells. We quantified ripA and iglA expression at different stages of intracellular growth and found that the expression of each increased between 1 and 6 hours post infection. Given the similar intracellular expression patterns of ripA and iglA and that MglA and SspA are positive regulators of iglA we tested the impact of mglA and sspA deletions on ripA and iglA expression. In the deletion mutant strains iglA expression was reduced dramatically as expected, however ripA expression was increased over 2-fold. Conclusion Expression of ripA is required for growth at neutral pH, is pH sensitive, and is responsive to the intracellular environment. The intracellular expression pattern of ripA coincided with iglA, which is positively regulated by MglA and SspA. However, in contrast to their positive impact on iglA expression, MglA and SspA negatively impacted ripA expression in

  20. Effect of antioxidants on thiobarbituric acid reactive substances, psychrotrophic bacteria and functional properties of mechanically deboned chicken meat irradiated with Cobalto-60 and electron beam sources

    Energy Technology Data Exchange (ETDEWEB)

    Brito, Poliana de Paula; Azevedo, Heliana de; Roque, Claudio Vitor; Pomarico Neto, Walter, E-mail: hgomes@cnen.gov.br, E-mail: pbrito@cnen.gov.br, E-mail: cvroque@cnen.gov.br, E-mail: abrusqui@cnen.gov.br [Comissao Nacional de Energia Nuclear (LAPOC/CNEN-MG), Pocos de Caldas, MG (Brazil). Laboratorio de Pocos de Caldas; Mourao, Gerson Barreto; Orlando, Eduardo Adilson; Miyagusku, Luciana, E-mail: marciamh@ital.sp.gov.br, E-mail: eduardo.orlando@ital.sp.gov.br [Instituto de Tecnologia dos Alimentos (ITAL), Campinas, SP (Brazil)

    2013-07-01

    Samples of MDCM with skin were divided into three groups: control (without antioxidants), Antioxidant 1 (Sodium Polyphosphate and Sodium Ascorbate and Antioxidant 2 (Rosemary Extract and α-Tocopherol. The three batches of samples were divided into nine groups: no antioxidant and non-irradiated (C), with antioxidant A1 and non-irradiated (A1), with antioxidant A2 and non-irradiated (A2) without antioxidant and irradiated in Cobalt-60 source (Co), with antioxidant A1 irradiated in Cobalt 60 source (A1Co) with antioxidant A2 irradiated in Cobalt-60 source (A2Co) with antioxidant A1 irradiated in Electron beam (A1Eb) and with antioxidant A2 irradiated in Electron beam (A2Eb). The samples was conditioned in a transparent, low density frozen overnight at a temperature of -18 ± 1 deg C in a chamber, and irradiated in this state with a dose of 3.0 kGy, used two sources of radiation: Cobalt-60 (3.1 kGy/h) and electron beam (7.86 kGy/s). After this process, the samples were evaluated during the refrigeration period (2 ± 1 deg C) for 11 days for the following analysis: total psychrotrophic bacteria count and thiobarbituric acid reactive substances (TBARS) and the analysis of functional properties were performed after the irradiation process. The use of the combination of rosemary antioxidant and α-tocopherol were able to significantly decrease TBARS values caused by the irradiation of samples in MDCM cobalt-60 sources and electron beam, and show a synergetic effect to processing with ionizing radiation to reduce of psychrotrophic bacteria count. The use of irradiation processing of MDCM did not negatively affect the functional properties studied. (author)

  1. Evolution of intracellular compartmentalization.

    Science.gov (United States)

    Diekmann, Yoan; Pereira-Leal, José B

    2013-01-15

    Cells compartmentalize their biochemical functions in a variety of ways, notably by creating physical barriers that separate a compartment via membranes or proteins. Eukaryotes have a wide diversity of membrane-based compartments, many that are lineage- or tissue-specific. In recent years, it has become increasingly evident that membrane-based compartmentalization of the cytosolic space is observed in multiple prokaryotic lineages, giving rise to several types of distinct prokaryotic organelles. Endosymbionts, previously believed to be a hallmark of eukaryotes, have been described in several bacteria. Protein-based compartments, frequent in bacteria, are also found in eukaryotes. In the present review, we focus on selected intracellular compartments from each of these three categories, membrane-based, endosymbiotic and protein-based, in both prokaryotes and eukaryotes. We review their diversity and the current theories and controversies regarding the evolutionary origins. Furthermore, we discuss the evolutionary processes acting on the genetic basis of intracellular compartments and how those differ across the domains of life. We conclude that the distinction between eukaryotes and prokaryotes no longer lies in the existence of a compartmentalized cell plan, but rather in its complexity.

  2. An intracellular replication niche for Vibrio cholerae in the amoeba Acanthamoeba castellanii.

    Science.gov (United States)

    Van der Henst, Charles; Scrignari, Tiziana; Maclachlan, Catherine; Blokesch, Melanie

    2016-04-01

    Vibrio cholerae is a human pathogen and the causative agent of cholera. The persistence of this bacterium in aquatic environments is a key epidemiological concern, as cholera is transmitted through contaminated water. Predatory protists, such as amoebae, are major regulators of bacterial populations in such environments. Therefore, we investigated the interaction between V. cholerae and the amoeba Acanthamoeba castellanii at the single-cell level. We observed that V. cholerae can resist intracellular killing. The non-digested bacteria were either released or, alternatively, established a replication niche within the contractile vacuole of A. castellanii. V. cholerae was maintained within this compartment even upon encystment. The pathogen ultimately returned to its aquatic habitat through lysis of A. castellanii, a process that was dependent on the production of extracellular polysaccharide by the pathogen. This study reinforces the concept that V. cholerae is a facultative intracellular bacterium and describes a new host-pathogen interaction.

  3. Colony stage and not facultative policing explains pattern of worker reproduction in the Saxon wasp.

    Science.gov (United States)

    Bonckaert, W; van Zweden, J S; d'Ettorre, P; Billen, J; Wenseleers, T

    2011-08-01

    Inclusive fitness theory predicts that in colonies of social Hymenoptera headed by a multiple-mated queen, workers should benefit from policing eggs laid by other workers. Foster & Ratnieks provided evidence that in the vespine wasp Dolichovespula saxonica, workers police other workers' eggs only in colonies headed by a multiple-mated queen, but not in those headed by a single-mated one. This conclusion, however, was based on a relatively small sample size, and the original study did not control for possible confounding variables such as the seasonal colony progression of the nests. Our aim, therefore, was to reinvestigate whether or not facultative worker policing occurs in D. saxonica. Remarkably, our data show that in the studied Danish population, there was no correlation between worker-worker relatedness and the percentage of worker-derived males. In addition, we show that variability in cuticular hydrocarbon profiles among the workers did not significantly correlate with relatedness and that workers therefore probably did not have sufficient information on queen mating frequency from the workers' cuticular hydrocarbon profiles. Hence, there was no evidence that workers facultatively policed other workers' eggs in response to queen mating frequency. Nevertheless, our data do show that the seasonal progression of the nest and the location in which the males were reared both explain the patterns of worker reproduction found. Overall, our results suggest that the earlier evidence for facultative worker policing in D. saxonica may have been caused by accidental correlations with certain confounding variables, or, alternatively, that there are large interpopulation differences in the expression of worker policing. © 2011 Blackwell Publishing Ltd.

  4. Anaerobic facultative bacteria isolated from the gut of rabbits fed different diets.

    Science.gov (United States)

    Canganella, F; Zirletta, G; Gualterio, L; Massa, S; Trovatelli, L D

    1992-11-01

    Anaerobic facultative bacteria colonizing the intestinal tract of conventional rabbits fed three different diets (standard pellet, hay and pellet/hay mixture) were enumerated in brain heart infusion agar. Colony counts recovered from homogenized samples of small intestine, caecum and rectum differed with reference to the diet given. Among anaerobic groups, identified from rabbit fed pellet/hay mixture, Enterococci (E. faecalis, E. avium, E. faecium and E. durans) represented the predominant flora. Enterobacters (E. cloacae and E. aerogenes) accounted for about 10 to 25% of the bacteria in the rectum and colon respectively, whereas Staphylococci (S. intermedius, S. epidermidis and S. lentus) represented 11% of the bacteria isolated from colon.

  5. Conditional Reduction of Predation Risk Associated with a Facultative Symbiont in an Insect.

    Directory of Open Access Journals (Sweden)

    Sarah Polin

    Full Text Available Symbionts are widespread among eukaryotes and their impacts on the ecology and evolution of their hosts are meaningful. Most insects harbour obligate and facultative symbiotic bacteria that can influence their phenotype. In the pea aphid Acyrthosiphon pisum, an astounding symbiotic-mediated phenotype has been recently observed: when infected with the symbiotic bacteria Rickettsiella viridis, young red aphid larvae become greener at adulthood and even darker green when co-infected with Rickettsiella viridis and Hamiltonella defensa. As body colour affects the susceptibility towards natural enemies in aphids, the influence of the colour change due to these facultative symbionts on the host survival in presence of predators was tested. Our results suggested that the Rickettsiella viridis infection may impact positively host survival by reducing predation risk. Due to results from uninfected aphids (i.e., more green ones attacked, the main assumption is that this symbiotic infection would deter the predatory ladybird feeding by reducing the profitability of their hosts rather than decreasing host detection through body colour change. Aphids co-infected with Rickettsiella viridis and Hamiltonella defensa were, however, more exposed to predation suggesting an ecological cost associated with multiple infections. The underlying mechanisms and ecological consequences of these symbiotic effects are discussed.

  6. No facultative worker policing in the honey bee (Apis mellifera L.).

    Science.gov (United States)

    Loope, Kevin J; Seeley, Thomas D; Mattila, Heather R

    2013-05-01

    Kin selection theory predicts that in colonies of social Hymenoptera with multiply mated queens, workers should mutually inhibit ("police") worker reproduction, but that in colonies with singly mated queens, workers should favor rearing workers' sons instead of queens' sons. In line with these predictions, Mattila et al. (Curr Biol 22:2027-2031, 2012) documented increased ovary development among workers in colonies of honey bees with singly mated queens, suggesting that workers can detect and respond adaptively to queen mating frequency and raising the possibility that they facultative police. In a follow-up experiment, we test and reject the hypothesis that workers in single-patriline colonies prefer worker-derived males and are able to reproduce directly; we show that their eggs are policed as strongly as those of workers in colonies with multiply mated queens. Evidently, workers do not respond facultatively to a kin structure that favors relaxed policing and increased direct reproduction. These workers may instead be responding to a poor queen or preparing for possible queen loss.

  7. Growth of the facultative anaerobes from Antarctica, Alaska, and Patagonia at low temperatures

    Science.gov (United States)

    Pikuta, Elena V.; Hoover, Richard B.

    2004-11-01

    Psychrotolerance, as an adaptation for surviving in extreme environments, is widespread among mesophilic microorganisms. Physico-chemical factors such as pressure, red-ox potential, pH and salinity could significantly alter the features of ecosystems by providing liquid water at subzero temperatures. Furthermore, organisms can respond to temperature changes by several known mechanisms, including changing the conformation capacities of constitutional proteins or by the synthesis of mucopolysaccharides around the cell wall and membrane. Such protective mechanisms make it possible for cells to not only passively survive low temperatures in a state of anabiosis, but also to be capable of actively metabolizing substrates and reproducing normally. The physiological and biochemical characteristics of the species, as well as genetics, could be remarkably changed due to adaptation and surviving in extreme environments. The cold shock genes of some of the studied strains of psychrotolerant facultative anaerobes were reported previously. In this paper we present experimental data for psychrotolerant, non spore-forming, facultative anaerobes isolated from geographically different cold regions of our planet. We show the growth response on changing from anaerobic conditions to aerobic with cultivation at low temperatures.

  8. Temperature dependent virulence of obligate and facultative fungal pathogens of honeybee brood.

    Science.gov (United States)

    Vojvodic, S; Jensen, A B; James, R R; Boomsma, J J; Eilenberg, J

    2011-04-21

    Chalkbrood (Ascosphaera apis) and stonebrood (Aspergillus flavus) are well known fungal brood diseases of honeybees (Apis mellifera), but they have hardly been systematically studied because the difficulty of rearing larvae in vitro has precluded controlled experimentation. Chalkbrood is a chronic honeybee-specific disease that can persist in colonies for years, reducing both brood and honey production, whereas stonebrood is a rare facultative pathogen that also affects hosts other than honeybees and can likely survive outside insect hosts. Hive infection trials have indicated that accidental drops in comb temperature increase the prevalence of chalkbrood, but it has remained unclear whether virulence is directly temperature-dependent. We used a newly established in vitro rearing technique for honeybee larvae to test whether there are systematic temperature effects on mortality induced by controlled infections, and whether such effects differed between the two fungal pathogens. We found that increasing spore dosage at infection had a more dramatic effect on mortality from stonebrood compared to chalkbrood. In addition, a 24h cooling period after inoculation increased larval mortality from chalkbrood infection, whereas such a cooling period decreased mortality after stonebrood infection. These results raise interesting questions about honeybee defenses against obligate and facultative pathogens and about the extent to which stress factors in the host (dis)favor pathogens with lesser degrees of specialization. Copyright © 2010 Elsevier B.V. All rights reserved.

  9. Serratia symbiotica from the aphid Cinara cedri: a missing link from facultative to obligate insect endosymbiont.

    Directory of Open Access Journals (Sweden)

    Araceli Lamelas

    2011-11-01

    Full Text Available The genome sequencing of Buchnera aphidicola BCc from the aphid Cinara cedri, which is the smallest known Buchnera genome, revealed that this bacterium had lost its symbiotic role, as it was not able to synthesize tryptophan and riboflavin. Moreover, the biosynthesis of tryptophan is shared with the endosymbiont Serratia symbiotica SCc, which coexists with B. aphidicola in this aphid. The whole-genome sequencing of S. symbiotica SCc reveals an endosymbiont in a stage of genome reduction that is closer to an obligate endosymbiont, such as B. aphidicola from Acyrthosiphon pisum, than to another S. symbiotica, which is a facultative endosymbiont in this aphid, and presents much less gene decay. The comparison between both S. symbiotica enables us to propose an evolutionary scenario of the transition from facultative to obligate endosymbiont. Metabolic inferences of B. aphidicola BCc and S. symbiotica SCc reveal that most of the functions carried out by B. aphidicola in A. pisum are now either conserved in B. aphidicola BCc or taken over by S. symbiotica. In addition, there are several cases of metabolic complementation giving functional stability to the whole consortium and evolutionary preservation of the actors involved.

  10. Genome characteristics of facultatively symbiotic Frankia sp. strains reflect host range and host plant biogeography

    Science.gov (United States)

    Normand, Philippe; Lapierre, Pascal; Tisa, Louis S.; Gogarten, Johann Peter; Alloisio, Nicole; Bagnarol, Emilie; Bassi, Carla A.; Berry, Alison M.; Bickhart, Derek M.; Choisne, Nathalie; Couloux, Arnaud; Cournoyer, Benoit; Cruveiller, Stephane; Daubin, Vincent; Demange, Nadia; Francino, Maria Pilar; Goltsman, Eugene; Huang, Ying; Kopp, Olga R.; Labarre, Laurent; Lapidus, Alla; Lavire, Celine; Marechal, Joelle; Martinez, Michele; Mastronunzio, Juliana E.; Mullin, Beth C.; Niemann, James; Pujic, Pierre; Rawnsley, Tania; Rouy, Zoe; Schenowitz, Chantal; Sellstedt, Anita; Tavares, Fernando; Tomkins, Jeffrey P.; Vallenet, David; Valverde, Claudio; Wall, Luis G.; Wang, Ying; Medigue, Claudine; Benson, David R.

    2007-01-01

    Soil bacteria that also form mutualistic symbioses in plants encounter two major levels of selection. One occurs during adaptation to and survival in soil, and the other occurs in concert with host plant speciation and adaptation. Actinobacteria from the genus Frankia are facultative symbionts that form N2-fixing root nodules on diverse and globally distributed angiosperms in the “actinorhizal” symbioses. Three closely related clades of Frankia sp. strains are recognized; members of each clade infect a subset of plants from among eight angiosperm families. We sequenced the genomes from three strains; their sizes varied from 5.43 Mbp for a narrow host range strain (Frankia sp. strain HFPCcI3) to 7.50 Mbp for a medium host range strain (Frankia alni strain ACN14a) to 9.04 Mbp for a broad host range strain (Frankia sp. strain EAN1pec.) This size divergence is the largest yet reported for such closely related soil bacteria (97.8%–98.9% identity of 16S rRNA genes). The extent of gene deletion, duplication, and acquisition is in concert with the biogeographic history of the symbioses and host plant speciation. Host plant isolation favored genome contraction, whereas host plant diversification favored genome expansion. The results support the idea that major genome expansions as well as reductions can occur in facultative symbiotic soil bacteria as they respond to new environments in the context of their symbioses. PMID:17151343

  11. Glucose transporter 1-positive endothelial cells in infantile hemangioma exhibit features of facultative stem cells

    Science.gov (United States)

    Huang, Lan; Nakayama, Hironao; Klagsbrun, Michael; Mulliken, John B.; Bischoff, Joyce

    2014-01-01

    Endothelial glucose transporter 1 (GLUT1) is a definitive and diagnostic marker for infantile hemangioma (IH), a vascular tumor of infancy. To date, GLUT1-positive endothelial cells in IH have not been quantified nor directly isolated and studied. We isolated GLUT1-positive and GLUT1-negative endothelial cells from IH specimens and characterized their proliferation, differentiation and response to propranolol, a first-line therapy for IH, and to rapamycin, an mTOR pathway inhibitor used to treat an increasingly wide array of proliferative disorders. Although freshly isolated GLUT1-positive cells, selected using anti-GLUT1 magnetic beads, expressed endothelial markers CD31, VE-Cadherin and VEGFR2, they converted to a mesenchymal phenotype after three weeks in culture. In contrast, GLUT1-negative endothelial cells exhibited a stable endothelial phenotype in vitro. GLUT1-selected cells were clonogenic when plated as single cells and could be induced to re-differentiate into endothelial cells, or into pericyte/smooth muscle cells or into adipocytes, indicating a stem cell-like phenotype. These data demonstrate that, although they appear and function in the tumor as bona fide endothelial cells, the GLUT1-positive endothelial cells display properties of facultative stem cells. Pretreatment with rapamycin for 4 days significantly slowed proliferation of GLUT1-selected cells, whereas propranolol pretreatment had no effect. These results reveal for the first time the facultative nature of GLUT1-positive endothelial cells in infantile hemangioma. PMID:25187207

  12. The Facultative Symbiont Rickettsia Protects an Invasive Whitefly against Entomopathogenic Pseudomonas syringae Strains.

    Science.gov (United States)

    Hendry, Tory A; Hunter, Martha S; Baltrus, David A

    2014-12-01

    Facultative endosymbionts can benefit insect hosts in a variety of ways, including context-dependent roles, such as providing defense against pathogens. The role of some symbionts in defense may be overlooked, however, when pathogen infection is transient, sporadic, or asymptomatic. The facultative endosymbiont Rickettsia increases the fitness of the sweet potato whitefly (Bemisia tabaci) in some populations through mechanisms that are not yet understood. In this study, we investigated the role of Rickettsia in mediating the interaction between the sweet potato whitefly and Pseudomonas syringae, a common environmental bacterium, some strains of which are pathogenic to aphids. Our results show that P. syringae multiplies within whiteflies, leading to host death, and that whiteflies infected with Rickettsia show a decreased rate of death due to P. syringae. Experiments using plants coated with P. syringae confirmed that whiteflies can acquire the bacteria at a low rate while feeding, leading to increased mortality, particularly when the whiteflies are not infected with Rickettsia. These results suggest that P. syringae may affect whitefly populations in nature and that Rickettsia can ameliorate this effect. This study highlights the possible importance of interactions among opportunistic environmental pathogens and endosymbionts of insects.

  13. No facultative worker policing in the honey bee ( Apis mellifera L.)

    Science.gov (United States)

    Loope, Kevin J.; Seeley, Thomas D.; Mattila, Heather R.

    2013-05-01

    Kin selection theory predicts that in colonies of social Hymenoptera with multiply mated queens, workers should mutually inhibit ("police") worker reproduction, but that in colonies with singly mated queens, workers should favor rearing workers' sons instead of queens' sons. In line with these predictions, Mattila et al. (Curr Biol 22:2027-2031, 2012) documented increased ovary development among workers in colonies of honey bees with singly mated queens, suggesting that workers can detect and respond adaptively to queen mating frequency and raising the possibility that they facultative police. In a follow-up experiment, we test and reject the hypothesis that workers in single-patriline colonies prefer worker-derived males and are able to reproduce directly; we show that their eggs are policed as strongly as those of workers in colonies with multiply mated queens. Evidently, workers do not respond facultatively to a kin structure that favors relaxed policing and increased direct reproduction. These workers may instead be responding to a poor queen or preparing for possible queen loss.

  14. A new dye uptake assay to test the activity of antibioticsagainst intracellular Francisella tularensis

    Directory of Open Access Journals (Sweden)

    Vivien eSutera

    2014-03-01

    Full Text Available Francisella tularensis, a facultative intracellular bacterium, is the aetiological agent of tularaemia. Antibiotic treatment of this zoonosis is based on the administration of a fluoroquinolone or a tetracycline for cases with mild to moderate severity, whereas an aminoglycoside (streptomycin or gentamicin is advocated for severe cases. However, treatment failures and relapses remain frequent, especially in patients suffering from chronic lymph node suppuration. Therefore, new treatment alternatives are needed. We have developed a dye uptake assay for determination of minimal inhibitory extracellular concentrations (MIECs of antibiotics against intracellular F. tularensis, and validated the method by comparing the results obtained using a CFU-enumerating method. We also compared MIECs with MICs of the same compounds determined using a CLSI broth microdilution method. We tested the activity of 11 antibiotics against two clinical strains of F. tularensis subsp. holarctica isolated in France. Both strains displayed low MICs (= 8 µg/mL were found for carbapenems (imipenem and meropenem, daptomycin and linezolid. Erythromycin MICs were 4.0 and 16.0 µg/mL, respectively, for the two clinical strains. MIECs were almost the same with the two methods used. They were concordant with MICs, except for erythromycin and linezolid (respectively, four and eight times more active against intracellular F. tularensis and gentamicin (four to eight times less active against intracellular F. tularensis. This study validated the dye uptake assay as a new tool for determination of the activity of a large panel of antibiotics against intracellular F. tularensis. This test confirmed the intracellular activity of first-line antibiotics used for tularaemia treatment, but also revealed significant activity of linezolid against intracellular F. tularensis.

  15. Spoilage potential of psychrotrophic lactic acid bacteria (LAB) species: Leuconostoc gelidum subsp. gasicomitatum and Lactococcus piscium, on sweet bell pepper (SBP) simulation medium under different gas compositions.

    Science.gov (United States)

    Pothakos, Vasileios; Nyambi, Clarice; Zhang, Bao-Yu; Papastergiadis, Antonios; De Meulenaer, Bruno; Devlieghere, Frank

    2014-05-16

    Sweet bell peppers are a significant constituent of retail, chilled-stored and packaged food products like fresh salads, marinades and ready-to-eat (RTE) meals. Previously, through general screening of the Belgian market and by means of source tracking analysis in a plant manufacturing minimally processed, vegetable salads the susceptibility of fresh-cut sweet bell peppers to lactic acid bacterium (LAB) contamination was substantiated. The determination of the metabolic profiles of Leuconostoc gelidum subsp. gasicomitatum and Lactococcus piscium, two major psychrotrophic, spoilage-related LAB species, on sweet bell pepper (SBP) simulation medium under different packaging conditions - 1.) vacuum: 100% N2, 2.) air: 21% O2, 79% N2, 3.) MAP1: 30% CO2, 70% N2 and 4.) MAP2: 50% O2, 50% CO2 - facilitated a better understanding of the spoilage potential of these microbes as well as the presumptive contribution of O2 in the spectrum of produced volatile organic compounds (VOCs) associated with poor organoleptic properties of food products. Generally, none of the applied gas compositions inhibited the growth of the 4 L. gelidum subsp. gasicomitatum isolates, however the presence of O2 resulted in buttery off-odors by inducing primarily the accumulation of diacetyl and pungent "vinegar" smell due to acetic acid. The 3 tested isolates of L. piscium varied greatly among their growth dynamics and inhibition at MAP2. They exhibited either weak spoilage profile or very offensive metabolism confirming significant intraspecies diversity.

  16. capA, a cspA-like gene that encodes a cold acclimation protein in the psychrotrophic bacterium Arthrobacter globiformis SI55.

    Science.gov (United States)

    Berger, F; Normand, P; Potier, P

    1997-09-01

    By use of Arthrobacter globiformis SI55, a psychrotrophic bacterium capable of growth between -5 and +32 degrees C, we cloned and sequenced capA, a gene homologous to cspA encoding the major cold shock protein in Escherichia coli. The deduced protein sequence has a high level of identity with the sequences of other CspA-related proteins from various sources, and no particular residue or domain that could be specific to cold-adapted microorganisms emerged. We show that CapA was produced very rapidly following cold shock, but unlike its mesophilic counterparts, it was still expressed during prolonged growth at low temperature. Its synthesis is regulated at the translational level, and we showed that growth resumption following a temperature downshift correlated with CapA expression. Transient inhibitions in protein synthesis during the first stages of the cold shock response severely impaired the subsequent acclimation of A. globiformis SI55 to low temperature and delayed CapA expression. The cold shock response in A. globiformis SI55 is an adaptative process in which CapA may play a crucial role. We suggest that low-temperature acclimation is conditioned mainly by the ability of cells to restore an active translational machinery after cold shock in a process that may be different from that present in mesophiles.

  17. Isolation and characteristics of one marine psychrotrophic cellulase-generating bacterium Ar/w/b/75°/10/5 from Chuckchi Sea, Arctic

    Institute of Scientific and Technical Information of China (English)

    曾胤新; 陈波

    2002-01-01

    Microorganisms living in polar zones play an important part as the potential source of organic activity materials with low temperature characteristics in the biotechnological applications. A psychrotrophic bacterium (strain Ar/w/b/75°/10/5), producing cellulase at low temperatures during late-exponential and early-stationary phases of cell growth, was isolated from sea ice-covered surface water in Chuckchi Sea, Arctic. This bacterium, with rod cells, was Gram-negative, slightly halophilic. Colony growing on agar plate was in black. Optimum growth temperature was 15℃. No cell growth was observed at 35℃ or above. Optimum salt concentration for cell growth was between 2 and 3 % of sodium chloride in media. Maximal cellulase activity was detected at a temperature of 35℃ and pH8. Cellulase was irreversibly inactivated when incubated at 55℃ within 30 min. Enzyme can be kept stable at the temperature no higher than 25℃. Of special interest was that this bacterium produced various extracellular enzymes including cellulase, amylase, agar hydrolase and protease, at low or moderate temperature conditions, which is certainly of it potential value for applications.

  18. First report of interspecific facultative social parasitism in the paper wasp genus Mischocyttarus Saussure (Hymenoptera, Vespidae

    Directory of Open Access Journals (Sweden)

    Thiago S. Montagna

    2012-06-01

    Full Text Available First report of interspecific facultative social parasitism in the paper wasp genus Mischocyttarus Saussure (Hymenoptera, Vespidae. Parasitism of colonies of the social wasp Mischocyttarus cerberus Ducke, 1918 by females of Mischocyttarus consimilis Zikán, 1949 was observed in a rural area of Dourados, state of Mato Grosso do Sul, Brazil. In all monitored cases, the invasion occurred in the pre-emergence colony stage, generally by a single female of M. consimilis. The period of establishment of the foreign female in the host colony was marked by antagonistic behaviors between the host female and the invasive. In general, the architecture of the parasitized nest was modified from the typical architecture of the host species nest.

  19. Enhanced start-up of anaerobic facultatively autotrophic biocathodes in bioelectrochemical systems.

    Science.gov (United States)

    Zaybak, Zehra; Pisciotta, John M; Tokash, Justin C; Logan, Bruce E

    2013-12-01

    Biocathodes in bioelectrochemical systems (BESs) can be used to convert CO2 into diverse organic compounds through a process called microbial electrosynthesis. Unfortunately, start-up of anaerobic biocathodes in BESs is a difficult and time consuming process. Here, a pre-enrichment method was developed to improve start-up of anaerobic facultatively autotrophic biocathodes capable of using cathodes as the electron donor (electrotrophs) and CO2 as the electron acceptor. Anaerobic enrichment of bacteria from freshwater bog sediment samples was first performed in batch cultures fed with glucose and then used to inoculate BES cathode chambers set at -0.4V (versus a standard hydrogen electrode; SHE). After two weeks of heterotrophic operation of BESs, CO2 was provided as the sole electron acceptor and carbon source. Consumption of electrons from cathodes increased gradually and was sustained for about two months in concert with a significant decrease in cathode chamber headspace CO2. The maximum current density consumed was -34 ± 4 mA/m(2). Biosynthesis resulted in organic compounds that included butanol, ethanol, acetate, propionate, butyrate, and hydrogen gas. Bacterial community analyses based on 16S rRNA gene clone libraries revealed Trichococcus palustris DSM 9172 (99% sequence identity) as the prevailing species in biocathode communities, followed by Oscillibacter sp. and Clostridium sp. Isolates from autotrophic cultivation were most closely related to Clostridium propionicum (99% sequence identity; ZZ16), Clostridium celerecrescens (98-99%; ZZ22, ZZ23), Desulfotomaculum sp. (97%; ZZ21), and Tissierella sp. (98%; ZZ25). This pre-enrichment procedure enables simplified start-up of anaerobic biocathodes for applications such as electrofuel production by facultatively autotrophic electrotrophs.

  20. Geographic variation in a facultative mutualism: consequences for local arthropod composition and diversity.

    Science.gov (United States)

    Rudgers, Jennifer A; Savage, Amy M; Rúa, Megan A

    2010-08-01

    Geographic variation in the outcome of interspecific interactions may influence not only the evolutionary trajectories of species but also the structure of local communities. We investigated this community consequence of geographic variation for a facultative mutualism between ants and wild cotton (Gossypium thurberi). Ants consume wild cotton extrafloral nectar and can protect plants from herbivores. We chose three sites that differed in interaction outcome, including a mutualism (ants provided the greatest benefits to plant fitness and responded to manipulations of extrafloral nectar), a potential commensalism (ants increased plant fitness but were unresponsive to extrafloral nectar), and a neutral interaction (ants neither affected plant fitness nor responded to extrafloral nectar). At all sites, we manipulated ants and extrafloral nectar in a factorial design and monitored the abundance, diversity, and composition of other arthropods occurring on wild cotton plants. We predicted that the effects of ants and extrafloral nectar on arthropods would be largest in the location with the mutualism and weakest where the interaction was neutral. A non-metric multidimensional scaling analysis revealed that the presence of ants altered arthropod composition, but only at the two sites in which ants increased plant fitness. At the site with the mutualism, ants also suppressed detritivore/scavenger abundance and increased aphids. The presence of extrafloral nectar increased arthropod abundance where mutual benefits were the strongest, whereas both arthropod abundance and morphospecies richness declined with extrafloral nectar availability at the site with the weakest ant-plant interaction. Some responses were geographically invariable: total arthropod richness and evenness declined by approximately 20% on plants with ants, and extrafloral nectar reduced carnivore abundance when ants were excluded from plants. These results demonstrate that a facultative ant-plant mutualism

  1. Effects of Recent Climate Change on Facultative and Spontaneous Torpor in Apline Habitats

    Science.gov (United States)

    Frank, C. L.; Hood, W. R.; Stevens, M.; Gary, G.

    2007-12-01

    Mean annual air temperatures have increased in North America by 1.0°C during the past 100 years, and are predicted to increase by 4-8 °C further within the next 70 years. Hibernating mammals may be particularly sensitive to climate change since body temperatures during torpor are strongly influence by ambient temperature. We conducted 3-7 year studies on the relationship between ambient (air/soil) temperature and the torpor patterns of free-ranging facultative and spontaneous hibernators. The facultative hibernation of eastern chipmunks (Tamias striatus) in New York State and the spontaneous hibernation by golden-mantled ground squirrels (Spermophilus lateralis) in the mountains of California were continuously monitored using temperature sensitive radiocollars during winters of 2000-01 through 2006-07. Mean air/soil temperatures during the winter 2001-02 were much greater than those observed during the same periods of 2000-1 and 2002-3 winters at both sites, and the winter of 2001-2 was one of the warmest measured in both New York State and California since 1895. Consequently T. striatus during the winter 2001-2 had: a) fewer individuals using torpor, b) reduced the time spent in torpor by 96%, and, c) increased energy expenditure by 100% when compared to the torpor patterns of the same population during the colder winters of 2000-1/2002-3. Likewise, S. lateralis during the winter of 2001-2 had: a) delayed the entrance into hibernation by a mean of 12.2 days, b) increased mean body temperatures by an average of 5.4 C during torpor, and, c) increased metabolic rate during torpor by 75% when compared to the torpor patterns of the same population during the winters of 2000-1/2002-3.

  2. The content of arbitration agreement: Facultative elements as an instrument for exercising contractual freedom

    Directory of Open Access Journals (Sweden)

    Janićijević Dejan

    2014-01-01

    Full Text Available The content of an arbitration agreement is of great practical importance, given the effect its provisions may have on the rights and interests of the parties in the arbitration proceedings. Beside the mandatory elements, an arbitration agreement may include numerous facultative provisions, where parties express their contractual freedom, limited only by the imperative law and institutional arbitration rules. Even in cases when contractual provisions are contrary to the imperative law, in many states, the policy of maintaining arbitration agreement would result in ignoring the problematic provisions. The list of facultative elements of an arbitration agreement is practically limitless; among other elements, it may include: time limitations on the validity of an arbitration agreement; determinations referring to the composition of the arbitration tribunal; detailed rules pertaining to the procedure, decision-making processes and content of the arbitration award as well as its annulment; provisions on the choice of law; the right of third parties to intervene; joinder, consolidation and the authority of arbitration tribunal to decide as amiable compositeur or ex aequo et bono. The development of international commerce as well as arbitration legislature and practice, featuring transactions of ever-increasing complexity, have brought about the need to establish dispute resolution mechanisms modulated according to their specific characteristics. The customary practice of concluding arbitration agreements in the form of standardized and simple clauses is being challenged by the growing complexity of international trade. Therefore, not only should a modern arbitration agreement clearly demonstrate the parties' intent to submit a dispute to arbitration but it should also conform the arbitration procedure (as much as possible] to the content of the parties' relationship and the dispute that has emerged or may arise thereof.

  3. O2-sensing and O2-dependent gene regulation in facultatively anaerobic bacteria.

    Science.gov (United States)

    Unden, G; Becker, S; Bongaerts, J; Holighaus, G; Schirawski, J; Six, S

    1995-08-01

    Availability of O2 is one of the most important regulatory signals in facultatively anaerobic bacteria. Various two- or one-component sensor/regulator systems control the expression of aerobic and anaerobic metabolism in response to O2. Most of the sensor proteins contain heme or Fe as cofactors that interact with O2 either by binding or by a redox reaction. The ArcA/ArcB regulator of aerobic metabolism in Escherichia coli may use a different sensory mechanism. In two-component regulators, the sensor is located in the cytoplasmic membrane, whereas one-component regulators are located in the cytoplasm. Under most conditions, O2 can readily reach the cytoplasm and could provide the signal in the cytoplasm. The transcriptional regulator FNR of E. Coli controls the expression of many genes required for anaerobic metabolism in response to O2. Functional homologs of FNR are present in facultatively anaerobic Proteobacteria and presumably also in gram-positive bacteria. The target genes of FNR are mostly under multiple regulation by FNR and other regulators that respond to O2, nitrate, or glucose. FNR represents a 'one-component' sensor/regulator and contains Fe for signal perception. In response to O2 availability, FNR is converted reversibly from the aerobic (inactive) state to the anaerobic (active) state. Experiments suggest that the Fe cofactor is bound by four essential cysteine residues. The O2-triggered transformation between active and inactive FNR presumably is due to a redox reaction at the Fe cofactor, but other modes of interaction cannot be excluded. O2 seems to affect the site-specific DNA binding of FNR at target genes or the formation of an active transcriptional complex with RNA polymerase.

  4. Photoperiod extension enhances sexual megaspore formation and triggers metabolic reprogramming in facultative apomictic Ranunculus auricomus

    Directory of Open Access Journals (Sweden)

    Simone eKlatt

    2016-03-01

    Full Text Available Abstract••Meiosis, the key step of sexual reproduction, persists in facultative apomictic plants functional to some extent. However, it still remains unclear how and why proportions of reproductive pathways vary under different environmental stress conditions. We hypothesized that oxidative stress mediates alterations of developmental pathways. In apomictic plants we expected that megasporogenesis, the stage directly after meiosis, would be more affected than later stages of seed development.•To simulate moderate stress conditions we subjected clone-mates of facultative apomictic Ranunculus auricomus to 10 h photoperiods, reflecting natural conditions, and extended ones (16.5 h. Reproduction mode was screened directly after megasporogenesis (microscope and at seed stage (flow cytometric seed screening. Targeted metabolite profiles were performed with HPLC−DAD to explore if and which metabolic reprogramming was caused by the extended photoperiod.•Prolonged photoperiods resulted in increased frequencies of sexual vs. aposporous initials directly after meiosis, but did not affect frequencies of sexual vs. asexual seed formation. Changes in secondary metabolite profiles under extended photoperiods affected all classes of compounds, and c. 20% of these changes separated the two treatments. Unexpectedly, the renowned antioxidant phenylpropanoids and flavonoids added more to clone-mate variation than to treatment differentiation. Among others, chlorophyll degradation products, non-assigned phenolic compounds and more lipophilic metabolites also contributed to the dissimilarity of the metabolic profiles of plants that had been exposed to the two different photoperiods.•The hypothesis of moderate light stress effects was supported by increased proportions of sexual megaspore development at the expense of aposporous initial formation. The lack of effects at the seed stage confirms the basic assumption that only meiosis and sporogenesis would

  5. Enhanced start-up of anaerobic facultatively autotrophic biocathodes in bioelectrochemical systems

    KAUST Repository

    Zaybak, Zehra

    2013-12-01

    Biocathodes in bioelectrochemical systems (BESs) can be used to convert CO2 into diverse organic compounds through a process called microbial electrosynthesis. Unfortunately, start-up of anaerobic biocathodes in BESs is a difficult and time consuming process. Here, a pre-enrichment method was developed to improve start-up of anaerobic facultatively autotrophic biocathodes capable of using cathodes as the electron donor (electrotrophs) and CO2 as the electron acceptor. Anaerobic enrichment of bacteria from freshwater bog sediment samples was first performed in batch cultures fed with glucose and then used to inoculate BES cathode chambers set at -0.4V (versus a standard hydrogen electrode; SHE). After two weeks of heterotrophic operation of BESs, CO2 was provided as the sole electron acceptor and carbon source. Consumption of electrons from cathodes increased gradually and was sustained for about two months in concert with a significant decrease in cathode chamber headspace CO2. The maximum current density consumed was -34±4mA/m2. Biosynthesis resulted in organic compounds that included butanol, ethanol, acetate, propionate, butyrate, and hydrogen gas. Bacterial community analyses based on 16S rRNA gene clone libraries revealed Trichococcus palustris DSM 9172 (99% sequence identity) as the prevailing species in biocathode communities, followed by Oscillibacter sp. and Clostridium sp. Isolates from autotrophic cultivation were most closely related to Clostridium propionicum (99% sequence identity; ZZ16), Clostridium celerecrescens (98-99%; ZZ22, ZZ23), Desulfotomaculum sp. (97%; ZZ21), and Tissierella sp. (98%; ZZ25). This pre-enrichment procedure enables simplified start-up of anaerobic biocathodes for applications such as electrofuel production by facultatively autotrophic electrotrophs. © 2013 Elsevier B.V.

  6. Coalescent Times and Patterns of Genetic Diversity in Species with Facultative Sex: Effects of Gene Conversion, Population Structure, and Heterogeneity.

    Science.gov (United States)

    Hartfield, Matthew; Wright, Stephen I; Agrawal, Aneil F

    2016-01-01

    Many diploid organisms undergo facultative sexual reproduction. However, little is currently known concerning the distribution of neutral genetic variation among facultative sexual organisms except in very simple cases. Understanding this distribution is important when making inferences about rates of sexual reproduction, effective population size, and demographic history. Here we extend coalescent theory in diploids with facultative sex to consider gene conversion, selfing, population subdivision, and temporal and spatial heterogeneity in rates of sex. In addition to analytical results for two-sample coalescent times, we outline a coalescent algorithm that accommodates the complexities arising from partial sex; this algorithm can be used to generate multisample coalescent distributions. A key result is that when sex is rare, gene conversion becomes a significant force in reducing diversity within individuals. This can reduce genomic signatures of infrequent sex (i.e., elevated within-individual allelic sequence divergence) or entirely reverse the predicted patterns. These models offer improved methods for assessing null patterns of molecular variation in facultative sexual organisms.

  7. Patho-epigenetics of Infectious Diseases Caused by Intracellular Bacteria.

    Science.gov (United States)

    Niller, Hans Helmut; Minarovits, Janos

    2016-01-01

    In multicellular eukaryotes including plants, animals and humans, epigenetic reprogramming may play a role in the pathogenesis of a wide variety of diseases. Recent studies revealed that in addition to viruses, pathogenic bacteria are also capable to dysregulate the epigenetic machinery of their target cells. In this chapter we focus on epigenetic alterations induced by bacteria infecting humans. Most of them are obligate or facultative intracellular bacteria that produce either bacterial toxins and surface proteins targeting the host cell membrane, or synthesise effector proteins entering the host cell nucleus. These bacterial products typically elicit histone modifications, i.e. alter the "histone code". Bacterial pathogens are capable to induce alterations of host cell DNA methylation patterns, too. Such changes in the host cell epigenotype and gene expression pattern may hinder the antibacterial immune response and create favourable conditions for bacterial colonization, growth, or spread. Epigenetic dysregulation mediated by bacterial products may also facilitate the production of inflammatory cytokines and other inflammatory mediators affecting the epigenotype of their target cells. Such indirect epigenetic changes as well as direct interference with the epigenetic machinery of the host cells may contribute to the initiation and progression of malignant tumors associated with distinct bacterial infections.

  8. Functional genomics of intracellular bacteria.

    Science.gov (United States)

    de Barsy, Marie; Greub, Gilbert

    2013-07-01

    During the genomic era, a large amount of whole-genome sequences accumulated, which identified many hypothetical proteins of unknown function. Rapidly, functional genomics, which is the research domain that assign a function to a given gene product, has thus been developed. Functional genomics of intracellular pathogenic bacteria exhibit specific peculiarities due to the fastidious growth of most of these intracellular micro-organisms, due to the close interaction with the host cell, due to the risk of contamination of experiments with host cell proteins and, for some strict intracellular bacteria such as Chlamydia, due to the absence of simple genetic system to manipulate the bacterial genome. To identify virulence factors of intracellular pathogenic bacteria, functional genomics often rely on bioinformatic analyses compared with model organisms such as Escherichia coli and Bacillus subtilis. The use of heterologous expression is another common approach. Given the intracellular lifestyle and the many effectors that are used by the intracellular bacteria to corrupt host cell functions, functional genomics is also often targeting the identification of new effectors such as those of the T4SS of Brucella and Legionella.

  9. Analysis of raw milk quality at reception and during cold storage: combined effects of somatic cell counts and psychrotrophic bacteria on lipolysis.

    Science.gov (United States)

    Gargouri, Ahmed; Hamed, Houda; Elfeki, Abdelfettah

    2013-09-01

    The objective of the article was to analyze the influence of psychrotrophic bacteria counts (PBCs) and somatic cell counts (SCCs) on the extent of lipolysis in bulk samples of cow's milk at reception and during cold storage. Samples of milk were analyzed on the day of sampling and subsequently during cold storage. The acidity, fat, density, chloride content, electrical conductivity (EC), bulk milk SCCs (BMSCC), and PBC values were analyzed on the day of sampling and the levels of acidity, EC, SCC, and PBC were analyzed during cold storage at 4 °C for 72 h. The SCC value 869 × 10(3) mL(-1) was higher than the recommended threshold. Lipolysis level at sampling day was related more closely with SCC than with PBC. There was no significant correlation between milk acidity and PBC among others parameters, while the milk mean density was only significant (P < 0.01) correlated with the fat content. The EC and chloride content were consistently correlated (P < 0.001) with BMSCC that allowed them to be used as indicators of mammary gland infection. The milk acidity, EC, PBC, and lipolysis levels increased in relation to the storage time at 4 °C. The lipolysis level during storage was in closer relation to the SCC, but not relation to the PBC. Effects of SCC and PBC on lipolysis decreased throughout the chilling period. It was concluded that initial lipolysis level and intrinsic milk lipoprotein lipase appear more effective than SCC and PBC on the development of lipolysis during storage.

  10. Metallochaperones regulate intracellular copper levels.

    Directory of Open Access Journals (Sweden)

    W Lee Pang

    Full Text Available Copper (Cu is an important enzyme co-factor that is also extremely toxic at high intracellular concentrations, making active efflux mechanisms essential for preventing Cu accumulation. Here, we have investigated the mechanistic role of metallochaperones in regulating Cu efflux. We have constructed a computational model of Cu trafficking and efflux based on systems analysis of the Cu stress response of Halobacterium salinarum. We have validated several model predictions via assays of transcriptional dynamics and intracellular Cu levels, discovering a completely novel function for metallochaperones. We demonstrate that in addition to trafficking Cu ions, metallochaperones also function as buffers to modulate the transcriptional responsiveness and efficacy of Cu efflux. This buffering function of metallochaperones ultimately sets the upper limit for intracellular Cu levels and provides a mechanistic explanation for previously observed Cu metallochaperone mutation phenotypes.

  11. Mechanisms of Intracellular Chlamydiae Survival

    Directory of Open Access Journals (Sweden)

    Lukic Ruzica

    2016-06-01

    Full Text Available Chlamydiae are Gram-negative, non-motile, obligate intracellular, and spherically shaped bacteria with a diameter of 0.2-1.5 μm. Chlamydiae are present in several different morphological forms: the elementary body, the reticular body, and in the last several years, there has been the observation of a third form known as the persistent or atypical form. The intracellular localization of Chlamydia provides a unique replication cycle that occurs inside a membrane-surrounded vacuole in the host cell cytoplasm and is significantly different from the method of multiplication of other microorganisms. Chlamydiae are capable of manipulating different signalling pathways inside the infected cell, thus avoiding the host immune response. This ensures intracellular multiplication, survival, and long-term persistence of Chlamydiae. There are two basic means of achieving this persistence: inhibition of apoptosis and manipulation of NF-κB (nuclear factor kappa B-mediated signals in the host.

  12. Sexual imprinting misguides species recognition in a facultative interspecific brood parasite.

    Science.gov (United States)

    Sorenson, Michael D; Hauber, Mark E; Derrickson, Scott R

    2010-10-22

    Sexual reproduction relies on the recognition of conspecifics for breeding. Most experiments in birds have implicated a critical role for early social learning in directing subsequent courtship behaviours and mating decisions. This classical view of avian sexual imprinting is challenged, however, by studies of megapodes and obligate brood parasites, species in which reliable recognition is achieved despite the lack of early experience with conspecifics. By rearing males with either conspecific or heterospecific brood mates, we experimentally tested the effect of early social experience on the association preferences and courtship behaviours of two sympatrically breeding ducks. We predicted that redheads (Aythya americana), which are facultative interspecific brood parasites, would show a diminished effect of early social environment on subsequent courtship preferences when compared with their host and congener, the canvasback (Aythya valisineria). Contrary to expectations, cross-fostered males of both species courted heterospecific females and preferred them in spatial association tests, whereas control males courted and associated with conspecific females. These results imply that ontogenetic constraints on species recognition may be a general impediment to the initial evolution of interspecific brood parasitism in birds. Under more natural conditions, a variety of mechanisms may mitigate or counteract the effects of early imprinting for redheads reared in canvasback broods.

  13. Introduire l'ELAO dans une Faculté de Lettres

    Directory of Open Access Journals (Sweden)

    Paul Bogaards

    2000-01-01

    Full Text Available Dans cet article je me propose de décrire les stratégies qui ont été utilisées pour introduire l'enseignement des langues assisté par ordinateur (ELAO dans la Faculté de Lettres de l'Université de Leyde (Pays-Bas. Je crois que l'approche choisie, si elle ne peut aucunement prétendre à servir de modèle, est tout de même suffisamment exemplaire pour pouvoir donner des idées, voire de l'espoir, à ceux et celles qui sont appelés à faire bénéficier l'enseignement des langues des acquis des technologies nouvelles dans des situations qui, à première vue, ne s'y prêtent guère. Très souvent les structures universitaires sont telles que toute modernisation doit se faire au prix de luttes qui demandent des sacrifices personnels. Mais une fois que les oppositions les plus féroces ont été vaincues, il s'agit de procéder de façon méthodique pour introduire les moyens informatiques dans le curriculum et pour faire en sorte que le plus grand nombre d'enseignants et d'étudiants puissent en profiter.

  14. Seroepidemiologic Survey of Potential Pathogens in Obligate and Facultative Scavenging Avian Species in California.

    Directory of Open Access Journals (Sweden)

    Mary H Straub

    Full Text Available Throughout the world, populations of scavenger birds are declining rapidly with some populations already on the brink of extinction. Much of the current research into the factors contributing to these declines has focused on exposure to drug residues, lead, and other toxins. Despite increased monitoring of these declining populations, little is known about infectious diseases affecting scavenger bird species. To assess potential infectious disease risks to both obligate and facultative scavenger bird species, we performed a serosurvey for eleven potential pathogens in three species of scavenging birds in California: the California condor (Gymnogyps californianus, turkey vulture (Cathartes aura and golden eagle (Aquila chrysaetos. California condors were seropositive for avian adenovirus, infectious bronchitis virus, Mycoplasma gallisepticum, avian paramyxovirus-2, West Nile virus (WNV and Toxoplasma gondii. Golden eagles were seropositive for avian adenovirus, Chlamydophila psittaci and Toxoplasma gondii, and turkey vultures were seropositive for avian adenovirus, Chlamydophila psittaci, avian paramyxovirus-1, Toxoplasma gondii and WNV. Risk factor analyses indicated that rearing site and original release location were significantly associated with a positive serologic titer to WNV among free-flying condors. This study provides preliminary baseline data on infectious disease exposure in these populations for aiding in early disease detection and provides potentially critical information for conservation of the endangered California condor as it continues to expand its range and encounter new infectious disease threats.

  15. Genomes of three facultatively symbiotic Frankia sp. strainsreflect host plant biogeography

    Energy Technology Data Exchange (ETDEWEB)

    Normand, Philippe; Lapierre, Pascal; Tisa, Louis S.; Gogarten, J.Peter; Alloisio, Nicole; Bagnarol, Emilie; Bassi, Carla A.; Berry,Alison; Bickhart, Derek M.; Choisne, Nathalie; Couloux, Arnaud; Cournoyer, Benoit; Cruveiller, Stephane; Daubin, Vincent; Demange, Nadia; Francino, M. Pilar; Ggoltsman, Eugene; Huang, Ying; Kopp, Olga; Labarre,Laurent; Lapidus, Alla; Lavire, Celine; Marechal, Joelle; Martinez,Michele; Mastronunzio, Juliana E.; Mullin, Beth; Niemann, James; Pujic,Pierre; Rawnsley, Tania; Rouy, Zoe; Schenowitz, Chantal; Sellstedt,Anita; Tavares, Fernando; Tomkins, Jeffrey P.; Vallenet, David; Valverde,Claudio; Wall, Luis; Wang, Ying; Medigue, Claudine; Benson, David R.

    2006-02-01

    Filamentous actinobacteria from the genus Frankia anddiverse woody trees and shrubs together form N2-fixing actinorhizal rootnodule symbioses that are a major source of new soil nitrogen in widelydiverse biomes 1. Three major clades of Frankia sp. strains are defined;each clade is associated with a defined subset of plants from among theeight actinorhizal plant families 2,3. The evolution arytrajectoriesfollowed by the ancestors of both symbionts leading to current patternsof symbiont compatibility are unknown. Here we show that the competingprocesses of genome expansion and contraction have operated in differentgroups of Frankia strains in a manner that can be related to thespeciation of the plant hosts and their geographic distribution. Wesequenced and compared the genomes from three Frankia sp. strains havingdifferent host plant specificities. The sizes of their genomes variedfrom 5.38 Mbp for a narrow host range strain (HFPCcI3) to 7.50Mbp for amedium host range strain (ACN14a) to 9.08 Mbp for a broad host rangestrain (EAN1pec.) This size divergence is the largest yet reported forsuch closely related bacteria. Since the order of divergence of thestrains is known, the extent of gene deletion, duplication andacquisition could be estimated and was found to be inconcert with thebiogeographic history of the symbioses. Host plant isolation favoredgenome contraction, whereas host plant diversification favored genomeexpansion. The results support the idea that major genome reductions aswell as expansions can occur in facultatively symbiotic soil bacteria asthey respond to new environments in the context of theirsymbioses.

  16. Distribution of pink-pigmented facultative methylotrophs on leaves of vegetables.

    Science.gov (United States)

    Mizuno, Masayuki; Yurimoto, Hiroya; Yoshida, Naoko; Iguchi, Hiroyuki; Sakai, Yasuyoshi

    2012-01-01

    The distribution of pink-pigmented facultative methylotrophs (PPFMs) on the leaves of various vegetables was studied. All kinds of vegetable leaves tested gave pink-pigmented colonies on agar plates containing methanol as sole carbon source. The numbers of PPFMs on the leaves, colony-forming units (CFU)/g of fresh leaves, differed among the plants, although they were planted and grown at the same farm. Commercial green perilla, Perilla frutescens viridis (Makino) Makino, gave the highest counts of PPFMs (2.0-4.1×10(7) CFU/g) of all the commercial vegetable leaves tested, amounting to 15% of total microbes on the leaves. The PPFMs isolated from seeds of two varieties of perilla, the red and green varieties, exhibited high sequence similarity as to the 16S rRNA gene to two different Methylobacterium species, M. fujisawaense DSM5686(T) and M. radiotolerans JCM2831(T) respectively, suggesting that there is specific interaction between perilla and the PPFMs.

  17. Mutualism exploitation: predatory drosophilid larvae sugar-trap ants and jeopardize facultative ant-plant mutualism.

    Science.gov (United States)

    Vidal, Mayra C; Sendoya, Sebastian F; Oliveira, Paulo S

    2016-07-01

    An open question in the evolutionary ecology of ant-plant facultative mutualism is how other members of the associated community can affect the interaction to a point where reciprocal benefits are disrupted. While visiting Qualea grandiflora shrubs to collect sugary rewards at extrafloral nectaries, tropical savanna ants deter herbivores and reduce leaf damage. Here we show that larvae of the fly Rhinoleucophenga myrmecophaga, which develop on extrafloral nectaries, lure potentially mutualistic, nectar-feeding ants and prey on them. Foraging ants spend less time on fly-infested foliage. Field experiments showed that predation (or the threat of predation) on ants by fly larvae produces cascading effects through three trophic levels, resulting in fewer protective ants on leaves, increased numbers of chewing herbivores, and greater leaf damage. These results reveal an undocumented mode of mutualism exploitation by an opportunistic predator at a plant-provided food source, jeopardizing ant-derived protection services to the plant. Our study documents a rather unusual case of predation of adult ants by a dipteran species and demonstrates a top-down trophic cascade within a generalized ant-plant mutualism. © 2016 by the Ecological Society of America.

  18. Involvement of facultative apomixis in inheritance of EPSPS gene amplification in glyphosate-resistant Amaranthus palmeri.

    Science.gov (United States)

    Ribeiro, Daniela N; Pan, Zhiqiang; Duke, Stephen O; Nandula, Vijay K; Baldwin, Brian S; Shaw, David R; Dayan, Franck E

    2014-01-01

    The inheritance of glyphosate resistance in two Amaranthus palmeri populations (R1 and R2) was examined in reciprocal crosses (RC) and second reciprocal crosses (2RC) between glyphosate-resistant (R) and -susceptible (S) parents of this dioecious species. R populations and Female-R × Male-S crosses contain higher 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene copy numbers than the S population. EPSPS expression, EPSPS enzyme activity, EPSPS protein quantity, and level of resistance to glyphosate correlated positively with genomic EPSPS relative copy number. Transfer of resistance was more influenced by the female than the male parent in spite of the fact that the multiple copies of EPSPS are amplified in the nuclear genome. This led us to hypothesize that this perplexing pattern of inheritance may result from apomictic seed production in A. palmeri. We confirmed that reproductively isolated R and S female plants produced seeds, indicating that A. palmeri can produce seeds both sexually and apomictically (facultative apomixis). This apomictic trait accounts for the low copy number inheritance in the Female-S × Male-R offsprings. Apomixis may also enhance the stability of the glyphosate resistance trait in the R populations in the absence of reproductive partners.

  19. FACULTATIVE PARASITISM BY THE BIVALVE KURTIELLA PEDROANA IN THE SAND CRAB EMERITA ANALOGA.

    Science.gov (United States)

    Bhaduri, Ritin; Valentich-Scott, Paul; Hilgers, Mark; Singh, Rajvir; Hickman, Mikaila; Lafferty, Kevin

    2017-08-31

    It is rare that an organism capable of independent or commensalistic existence can also become endoparasitic on a host. In this study, we documented a potential step toward parasitism in the commensal clam Kurtiella pedroana (Bivalvia: Galeommatoidea). Galeommatoideans are known commensals of various invertebrates, including crustaceans. Emerita analoga (Decapoda: Hippidae) is an abundant intertidal decapod inhabiting sandy beaches of the Pacific coast of North and South America. Crabs collected from Monterey Bay, California, were measured and examined externally and internally for associated molluscs. Out of the 520 crabs, 37 large female individuals harbored 49 bivalves (prevalence of 7.11% and mean intensity of 1.3). Forty-one ectocommensal clams were either inside the crab's branchial chambers or on their lateroventral surfaces, and were attached by byssal threads. Our key finding was eight clams that lacked byssal threads and were living in the hemocoel. These internal clams were significantly smaller than the ectocommensals. Because these internal clams lacked access to their normal food, we hypothesize they might have fed on their host's hemolymph as would a parasite. This clam species likely can't reproduce inside its host, implying that endoparasitism is a dead-end state for K. pedroana. Facultative parasitism in a free-living or an ectocommensal is uncommon and suggests a pathway to parasitism.

  20. Validation Hydrodynamic Models of Three Topological Models of Secondary Facultative Ponds

    Directory of Open Access Journals (Sweden)

    Aponte-Reyes Alxander

    2014-10-01

    Full Text Available A methodology was developed to analyze boundary conditions, the size of the mesh and the turbulence of a mathematical model of CFD, which could explain hydrodynamic behavior on facultative stabilization ponds, FSP, built to pilot scale: conventional pond, CP, baffled pond, BP, and baffled-mesh pond, BMP. Models dispersion studies were performed in field for validation, taking samples into and out of the FSP, the information was used to carry out CFD model simulations of the three topologies. Evaluated mesh sizes ranged from 500,000 to 2,000,000 elements. The boundary condition in Pared surface-free slip showed good qualitative behavior and the turbulence model κ–ε Low Reynolds yielded good results. The biomass contained in LFS generates interference on dispersion studies and should be taken into account in assessing the CFD modeling, the tracer injection times, its concentration at the entrance, the effect of wind on CFD, and the flow models adopted as a basis for modeling are parameters to be taken into account for the CFD model validation and calibration.

  1. Isolation and characterization of facultative mixotrophic ammonia-oxidizing bacteria from constructed wetlands

    Institute of Scientific and Technical Information of China (English)

    Soulwène Kouki; Neila Saidi; Fadhel M'hiri; Houda Nasr; Hanène Cherif; Hadda Ouzari; Abdermaceur Hassen

    2011-01-01

    Autotrophic ammonia-oxidizing bacteria (AOB) have been widely studied in constructed wetlands systems,while mixotrophic AOB have been less thoroughly examined.Heterotrophic bacteria were isolated from wastewater and rhizospheres of macrophytes of constructed wetlands,and then cultivated in a mixotrophic medium containing ammonium and acetic acid.A molecular characterization was accomplished using ITS-PCR amplification,and phylogenetic analysis based on 16S rRNA gene sequences.Results showed the presence of 35 bacteria,among 400 initially heterotrophic isolates,that were able to remove ammonia.These 35 isolates were classified into 10 genetically different groups based on ITS pattern.Then,a collection of 10 isolates were selected because of their relatively high ammonia removal efficiencies (ARE ≥ 80%) and their phylogenetic diversity.In conditions of mixotrophy,these strains were shown to be able to grow (increase of optical density OD660 during incubation with assimilation of nitrogen into cellular biomass) and to oxidize ammonia (important ammonia oxidation efficiencies,AOE between 79% and 87%).Among these facultative mixotrophic AOB,four isolates were genetically related to Firmicutes (Bacillus and Exiguobacterium),three isolates were affiliated to Actinobacteria (Arthrobacter) and three other isolates were associated with Proteobacteria (Pseudomonas,Ochrobactrum and Bordetella).

  2. Facultative control of matrix production optimizes competitive fitness in Pseudomonas aeruginosa PA14 biofilm models.

    Science.gov (United States)

    Madsen, Jonas S; Lin, Yu-Cheng; Squyres, Georgia R; Price-Whelan, Alexa; de Santiago Torio, Ana; Song, Angela; Cornell, William C; Sørensen, Søren J; Xavier, Joao B; Dietrich, Lars E P

    2015-12-01

    As biofilms grow, resident cells inevitably face the challenge of resource limitation. In the opportunistic pathogen Pseudomonas aeruginosa PA14, electron acceptor availability affects matrix production and, as a result, biofilm morphogenesis. The secreted matrix polysaccharide Pel is required for pellicle formation and for colony wrinkling, two activities that promote access to O2. We examined the exploitability and evolvability of Pel production at the air-liquid interface (during pellicle formation) and on solid surfaces (during colony formation). Although Pel contributes to the developmental response to electron acceptor limitation in both biofilm formation regimes, we found variation in the exploitability of its production and necessity for competitive fitness between the two systems. The wild type showed a competitive advantage against a non-Pel-producing mutant in pellicles but no advantage in colonies. Adaptation to the pellicle environment selected for mutants with a competitive advantage against the wild type in pellicles but also caused a severe disadvantage in colonies, even in wrinkled colony centers. Evolution in the colony center produced divergent phenotypes, while adaptation to the colony edge produced mutants with clear competitive advantages against the wild type in this O2-replete niche. In general, the structurally heterogeneous colony environment promoted more diversification than the more homogeneous pellicle. These results suggest that the role of Pel in community structure formation in response to electron acceptor limitation is unique to specific biofilm models and that the facultative control of Pel production is required for PA14 to maintain optimum benefit in different types of communities.

  3. Cultivation of anaerobic and facultatively anaerobic bacteria from spacecraft-associated clean rooms.

    Science.gov (United States)

    Stieglmeier, Michaela; Wirth, Reinhard; Kminek, Gerhard; Moissl-Eichinger, Christine

    2009-06-01

    In the course of this biodiversity study, the cultivable microbial community of European spacecraft-associated clean rooms and the Herschel Space Observatory located therein were analyzed during routine assembly operations. Here, we focused on microorganisms capable of growing without oxygen. Anaerobes play a significant role in planetary protection considerations since extraterrestrial environments like Mars probably do not provide enough oxygen for fully aerobic microbial growth. A broad assortment of anaerobic media was used in our cultivation strategies, which focused on microorganisms with special metabolic skills. The majority of the isolated strains grew on anaerobic, complex, nutrient-rich media. Autotrophic microorganisms or microbes capable of fixing nitrogen were also cultivated. A broad range of facultatively anaerobic bacteria was detected during this study and also, for the first time, some strictly anaerobic bacteria (Clostridium and Propionibacterium) were isolated from spacecraft-associated clean rooms. The multiassay cultivation approach was the basis for the detection of several bacteria that had not been cultivated from these special environments before and also led to the discovery of two novel microbial species of Pseudomonas and Paenibacillus.

  4. Direct fitness correlates and thermal consequences of facultative aggregation in a desert lizard.

    Science.gov (United States)

    Rabosky, Alison R Davis; Corl, Ammon; Liwanag, Heather E M; Surget-Groba, Yann; Sinervo, Barry

    2012-01-01

    Social aggregation is a common behavioral phenomenon thought to evolve through adaptive benefits to group living. Comparing fitness differences between aggregated and solitary individuals in nature--necessary to infer an evolutionary benefit to living in groups--has proven difficult because communally-living species tend to be obligately social and behaviorally complex. However, these differences and the mechanisms driving them are critical to understanding how solitary individuals transition to group living, as well as how and why nascent social systems change over time. Here we demonstrate that facultative aggregation in a reptile (the Desert Night Lizard, Xantusia vigilis) confers direct reproductive success and survival advantages and that thermal benefits of winter huddling disproportionately benefit small juveniles, which can favor delayed dispersal of offspring and the formation of kin groups. Using climate projection models, however, we estimate that future aggregation in night lizards could decline more than 50% due to warmer temperatures. Our results support the theory that transitions to group living arise from direct benefits to social individuals and offer a clear mechanism for the origin of kin groups through juvenile philopatry. The temperature dependence of aggregation in this and other taxa suggests that environmental variation may be a powerful but underappreciated force in the rapid transition between social and solitary behavior.

  5. Direct fitness correlates and thermal consequences of facultative aggregation in a desert lizard.

    Directory of Open Access Journals (Sweden)

    Alison R Davis Rabosky

    Full Text Available Social aggregation is a common behavioral phenomenon thought to evolve through adaptive benefits to group living. Comparing fitness differences between aggregated and solitary individuals in nature--necessary to infer an evolutionary benefit to living in groups--has proven difficult because communally-living species tend to be obligately social and behaviorally complex. However, these differences and the mechanisms driving them are critical to understanding how solitary individuals transition to group living, as well as how and why nascent social systems change over time. Here we demonstrate that facultative aggregation in a reptile (the Desert Night Lizard, Xantusia vigilis confers direct reproductive success and survival advantages and that thermal benefits of winter huddling disproportionately benefit small juveniles, which can favor delayed dispersal of offspring and the formation of kin groups. Using climate projection models, however, we estimate that future aggregation in night lizards could decline more than 50% due to warmer temperatures. Our results support the theory that transitions to group living arise from direct benefits to social individuals and offer a clear mechanism for the origin of kin groups through juvenile philopatry. The temperature dependence of aggregation in this and other taxa suggests that environmental variation may be a powerful but underappreciated force in the rapid transition between social and solitary behavior.

  6. Comparative Genomics of Facultative Bacterial Symbionts Isolated from European Orius Species Reveals an Ancestral Symbiotic Association

    Directory of Open Access Journals (Sweden)

    Xiaorui Chen

    2017-10-01

    Full Text Available Pest control in agriculture employs diverse strategies, among which the use of predatory insects has steadily increased. The use of several species within the genus Orius in pest control is widely spread, particularly in Mediterranean Europe. Commercial mass rearing of predatory insects is costly, and research efforts have concentrated on diet manipulation and selective breeding to reduce costs and improve efficacy. The characterisation and contribution of microbial symbionts to Orius sp. fitness, behaviour, and potential impact on human health has been neglected. This paper provides the first genome sequence level description of the predominant culturable facultative bacterial symbionts associated with five Orius species (O. laevigatus, O. niger, O. pallidicornis, O. majusculus, and O. albidipennis from several geographical locations. Two types of symbionts were broadly classified as members of the genera Serratia and Leucobacter, while a third constitutes a new genus within the Erwiniaceae. These symbionts were found to colonise all the insect specimens tested, which evidenced an ancestral symbiotic association between these bacteria and the genus Orius. Pangenome analyses of the Serratia sp. isolates offered clues linking Type VI secretion system effector–immunity proteins from the Tai4 sub-family to the symbiotic lifestyle.

  7. Stable isotopes reveal habitat-related diet shifts in facultative deposit-feeders

    Science.gov (United States)

    Rossi, Francesca; Baeta, Alexandra; Marques, João C.

    2015-01-01

    Seagrass patches interspersed in a sediment matrix may vary environmental conditions and affect feeding habits of consumers and food-web structure. This paper investigates diet shifts between bare sediments and a Zostera noltei (Hornemann, 1832) meadow for three facultative deposit-feeding macrofaunal consumers, notably the bivalve Scrobicularia plana (da Costa, 1778), the polychaete Hediste diversicolor (O.T. Müller, 1776), and the gastropod Hydrobia ulvae (Pennant, 1778). In July 2008, one eelgrass meadow and two bare sediment locations were chosen in the Mondego estuary (40° 08″ N, 8° 50‧ W, Portugal) and sampled for stable isotope signatures (δ13C and δ15N) of macrofauna consumers and some of their potential basal food sources, such as sedimentary organic matter (SOM), microphytobenthos (MPB), seagrass shoots, leaves and seaweeds laying on the surface sediment. The δ15N of H. diversicolor was 3‰ higher in the eelgrass meadow than in bare sediment, indicating a change of trophic position, whereas the Bayesian stable-isotope mixing model showed that S. plana assimilated more macroalgal detritus than microphytobenthos in the eelgrass bed. Such habitat-related diet shifts have the potential to change structure and spatial dynamics of benthic food webs.

  8. Three stage cultivation process of facultative strain of Chlorella sorokiniana for treating dairy farm effluent and lipid enhancement.

    Science.gov (United States)

    Hena, S; Fatihah, N; Tabassum, S; Ismail, N

    2015-09-01

    Reserve lipids of microalgae are promising for biodiesel production. However, economically feasible and sustainable energy production from microalgae requires optimization of cultivation conditions for both biomass yield and lipid production of microalgae. Biomass yield and lipid production in microalgae are a contradictory problem because required conditions for both targets are different. Simultaneously, the mass cultivation of microalgae for biofuel production also depends extremely on the performance of the microalgae strains used. In this study a green unicellular microalgae Chlorella sorokiniana (DS6) isolated from the holding tanks of farm wastewater treatment plant using multi-step screening and acclimation procedures was found high-lipid producing facultative heterotrophic microalgae strain capable of growing on dairy farm effluent (DFE) for biodiesel feedstock and wastewater treatment. Morphological features and the phylogenetic analysis for the 18S rRNA identified the isolated strains. A novel three stage cultivation process of facultative strain of C. sorokiniana was examined for lipid production.

  9. Identifying potential evolutionary relationships within a facultative lycaenid-ant system: Ant association,oviposition, and butterfly-ant conflict

    Institute of Scientific and Technical Information of China (English)

    NEIL COLLIER

    2007-01-01

    Facultative associations are commonly encountered between ants and lycaenids,although the nature and patterns of associations are typically unclear. This study investigated a facultative symbiosis involving the lycaenid Theclinesthes albocincta (Lycaenidae), its host plant Adriana quadripartita and Australian native ants. Ants in the genera Ochetellus and Iridomyrmex were most frequently found in association with T. albocincta larvae,although Iridomyrmex ants were found in much lower abundance than were ants in Ochetellus. The abundances of Ochetellus and Iridomyrmex were highly correlated with larval abundance, but not egg abundance. Observations and experiments recorded oviposition on male inflorescences on more than 95% of occasions, but oviposition was not greater on inflorescences with ants present. Behavioral assays showed that Iridomyrmex ants were aggressive towards female butterflies on significantly more occasions than were Ochetellus ants. These findings indicate potential evolutionary relationships between T. albocincta and two genera of ants that were abundant within the habitat.

  10. Differences of metal concentrations and morpho-anatomical adaptations between obligate and facultative serpentinophytes from Western Serbia

    Directory of Open Access Journals (Sweden)

    Dudić B.

    2007-01-01

    Full Text Available In the present study, some important macro- and micronutrients (Ca, Mg, Fe, Ni and the trace element Cr are analyzed in the obligate serpentinophytes Fumana bonapartei and Stachys recta var. chrysophaea, Balkan endemic species, as well as in Seseli rigidum, a widespread facultative serpentinophyte. Differences of adaptive structural features between these plants relative to serpentine tolerance strategy are also presented. Our investigations were carried out on plant samples deriving from the serpentine area of Mt. Tara in Western Serbia.

  11. Gene flow between sexual and facultatively asexual lineages of an aphid species and the maintenance of reproductive mode variation.

    Science.gov (United States)

    Halkett, F; Plantegenest, M; Bonhomme, J; Simon, J-C

    2008-06-01

    Many organisms considered as strictly clonal may in fact experience some rare events of sexual reproduction with their sexual relatives. However, the rate of sexual-asexual gene flow has rarely been assessed mainly because its evaluation is difficult to achieve in the field. In the cyclically parthenogenetic aphid Rhopalosiphum padi, two main sets of lineages, differing in their investment in sexual reproduction and in their genetic attributes, co-exist even at a very fine scale: the 'sexual' lineages which have a full commitment to the sexual reproduction, and the 'facultatively asexual' lineages, which allocate investment in the sexual and parthenogenetic reproduction. This system offers a unique opportunity to tackle the genetic interactions between two contrasting reproductive modes. Here, we provide evidence that gene flow occurred between sexual and facultatively asexual lineages of R. padi. We carefully examined the shuffling in phenotypic and genotypic variation following a sexual reproduction event that took place in the field. Combining genotypic data and phenotypic measurements showed that this gene mixing led to the production of a wide array of reproductive modes, including strictly asexual lineages. Finally, we discuss the central role played by facultatively asexual lineages on the maintenance of reproductive mode variation.

  12. Arabidopsis ABCB21 is a facultative auxin importer/exporter regulated by cytoplasmic auxin concentration.

    Science.gov (United States)

    Kamimoto, Yoshihisa; Terasaka, Kazuyoshi; Hamamoto, Masafumi; Takanashi, Kojiro; Fukuda, Shoju; Shitan, Nobukazu; Sugiyama, Akifumi; Suzuki, Hideyuki; Shibata, Daisuke; Wang, Bangjun; Pollmann, Stephan; Geisler, Markus; Yazaki, Kazufumi

    2012-12-01

    The phytohormone auxin is critical for plant growth and many developmental processes. Members of the P-glycoprotein (PGP/ABCB) subfamily of ATP-binding cassette (ABC) transporters have been shown to function in the polar movement of auxin by transporting auxin over the plasma membrane in both monocots and dicots. Here, we characterize a new Arabidopsis member of the ABCB subfamily, ABCB21/PGP21, a close homolog of ABCB4, for which conflicting transport directionalities have been reported. ABCB21 is strongly expressed in the abaxial side of cotyledons and in junctions of lateral organs in the aerial part, whereas in roots it is specifically expressed in pericycle cells. Membrane fractionation by sucrose density gradient centrifugation followed by Western blot showed that ABCB21 is a plasma membrane-localized ABC transporter. A transport assay with Arabidopsis protoplasts suggested that ABCB21 was involved in IAA transport in an outward direction, while naphthalene acetic acid (NAA) was a less preferable substrate for ABCB21. Further functional analysis of ABCB21 using yeast import and export assays showed that ABCB21 mediates the 1-N-naphthylphthalamic acid (NPA)-sensitive translocation of auxin in an inward direction when the cytoplasmic IAA concentration is low, whereas this transporter mediates outward transport under high internal IAA. An increase in the cytoplasmic IAA concentration by pre-loading of IAA into yeast cells abolished the IAA uptake activity by ABCB21 as well as ABCB4. These findings suggest that ABCB21 functions as a facultative importer/exporter controlling auxin concentrations in plant cells.

  13. Facultative Apomixis in Garcinia atroviridis (Clusiaceae) and Effects of Different Pollination Regimes on Reproductive Success.

    Science.gov (United States)

    Pangsuban, Sasithorn; Bamroongrugsa, Noparat; Kanchanapoom, Kamnoon; Nualsri, Charassri

    2009-12-01

    Various aspects of the reproductive success of Garcinia atroviridis Griff. were studied. Controlled pollination experiments were carried out in an orchard located in Songkhla province, southern Thailand, from February to July 2003. Floral longevity, stigma receptivity, and pollen viability were examined before carrying out the experiments. Three pollination treatments were compared: open pollination, manual pollination with bags, and bags without pollination (apogamy). Although there was no significant difference in the initial fruit set, bagged and manual pollination produced a significantly greater fruit drop rate than apogamy or natural pollination at one week after the flowers had been pollinated. On the other hand, the apogamy treatment had a greater fruit drop rate than natural and manual pollination treatments before fruit maturation. In addition, unpollinated bagged flowers bore fewer and smaller fruit than naturally and manually cross-pollinated flowers. Although the fruits from unpollinated flowers were capable of asexual seed formation, they produced fewer seeds and had poorer seed quality (defined as average fresh weight and germination rate) than those from the other treatments. The occurrence of asexual and sexual reproduction was also studied using Random Amplification of Polymorphic DNA (RAPD) analysis and by comparing the patterns of bands produced from DNA extracted from the offspring of the naturally cross-pollinated fruits. On average, 58% of the offspring had a genetic constitution identical to that of the maternal parent (ranging from 36% to 87%), indicating that some offspring were produced without prior fertilisation. However, the remainder showed polymorphism, demonstrating the occurrence of sexual reproduction. These findings indicate that facultative apomixis occurred in the study population. However, a residual sexuality was important for fruit production, fruit size, normal seed set and seed quality.

  14. Growth of the facultative anaerobe Shewanella putrefaciens by elemental sulfur reduction

    Science.gov (United States)

    Moser, D. P.; Nealson, K. H.

    1996-01-01

    The growth of bacteria by dissimilatory elemental sulfur reduction is generally associated with obligate anaerobes and thermophiles in particular. Here we describe the sulfur-dependent growth of the facultatively anaerobic mesophile Shewanella putrefaciens. Six of nine representative S. putrefaciens isolates from a variety of environments proved able to grow by sulfur reduction, and strain MR-1 was chosen for further study. Growth was monitored in a minimal medium (usually with 0.05% Casamino Acids added as a growth stimulant) containing 30 mM lactate and limiting concentrations of elemental sulfur. When mechanisms were provided for the removal of the metabolic end product, H2S, measurable growth was obtained at sulfur concentrations of from 2 to 30 mM. Initial doubling times were ca. 1.5 h and substrate independent over the range of sulfur concentrations tested. In the cultures with the highest sulfur concentrations, cell numbers increased by greater than 400-fold after 48 h, reaching a maximum density of 6.8 x 10(8) cells ml-1. Yields were determined as total cell carbon and ranged from 1.7 to 5.9 g of C mol of S(0) consumed-1 in the presence of the amino acid supplement and from 0.9 to 3.4 g of C mol of S(0-1) in its absence. Several lines of evidence indicate that cell-to-sulfur contact is not required for growth. Approaches for the culture of sulfur-metabolizing bacteria and potential ecological implications of sulfur reduction in Shewanella-like heterotrophs are discussed.

  15. Shewanella amazonensis sp. nov., a novel metal-reducing facultative anaerobe from Amazonian shelf muds

    Science.gov (United States)

    Venkateswaran, K.; Dollhopf, M. E.; Aller, R.; Stackebrandt, E.; Nealson, K. H.

    1998-01-01

    A new bacterial species belonging to the genus Shewanella is described on the basis of phenotypic characterization and sequence analysis of its 16S rRNA-encoding and gyrase B (gyrB) genes. This organism, isolated from shallow-water marine sediments derived from the Amazon River delta, is a Gram-negative, motile, polarly flagellated, facultatively anaerobic, rod-shaped eubacterium and has a G&C content of 51.7 mol%. Strain SB2BT is exceptionally active in the anaerobic reduction of iron, manganese and sulfur compounds. SB2BT grows optimally at 35 degrees C, with 1-3% NaCl and over a pH range of 7-8. Analysis of the 16S rDNA sequence revealed a clear affiliation between strain SB2BT and members of the gamma subclass of the class Proteobacteria. High similarity values were found with certain members of the genus Shewanella, especially with Shewanella putrefaciens, and this was supported by cellular fatty acid profiles and phenotypic characterization. DNA-DNA hybridization between strain SB2BT and its phylogenetically closest relatives revealed low similarity values (24.6-42.7%) which indicated species status for strain SB2BT. That SB2BT represents a distinct bacterial species within the genus Shewanella is also supported by gyrB sequence analysis. Considering the source of the isolate, the name Shewanella amazonensis sp. nov. is proposed and strain SB2BT (= ATCC 700329T) is designated as the type strain.

  16. Draft genome of an Aerophobetes bacterium reveals a facultative lifestyle in deep-sea anaerobic sediments

    Institute of Scientific and Technical Information of China (English)

    Yong Wang; Zhao-Ming Gao; Jiang-Tao Li; Salim Bougouffa; Ren Mao Tian; Vladimir B.Bajic; Pei-Yuan Qian

    2016-01-01

    Aerophobetes (or CD12) is a recently defined bacterial phylum,of which the metabolic processes and ecological importance remain unclear.In the present study,we obtained the draft genome of an Aerophobetes bacterium TCS1 from saline sediment near the Thuwal cold seep in the Red Sea using a genome binning method.Analysis of 16S rRNA genes of TCS1 and close relatives revealed wide distribution of Aerophobetes in deep-sea sediments.Phylogenetic relationships showed affinity between Aerophobetes TCS1 and some thermophilic bacterial phyla.The genome of TCS1 (at least 1.27 Mbp)contains a full set of genes encoding core metabolic pathways,including glycolysis and pyruvate fermentation to produce acetyl-CoA and acetate.The identification of cross-membrane sugar transporter genes further indicates its potential ability to consume carbohydrates preserved in the sediment under the microbial mat.Aerophobetes bacterium TCS1 therefore probably carried out saccharolytic and fermentative metabolism.The genes responsible for autotrophic synthesis of acetyl-CoA via the Wood-Ljungdahl pathway were also found in the genome.Phylogenetic study of the essential genes for the Wood-Ljungdahl pathway implied relative independence of Aerophobetes bacterium from the known acetogens and methanogens.Compared with genomes of acetogenic bacteria,Aerophobetes bacterium TCS 1 genome lacks the genes involved in nitrogen metabolism,sulfur metabolism,signal transduction and cell motility.The metabolic activities of TCS1 might depend on geochemical conditions such as supplies of CO2,hydrogen and sugars,and therefore the TCS1 might be a facultative bacterium in anaerobic saline sediments near cold seeps.

  17. Population genomics of the facultatively mutualistic bacteria Sinorhizobium meliloti and S. medicae.

    Science.gov (United States)

    Epstein, Brendan; Branca, Antoine; Mudge, Joann; Bharti, Arvind K; Briskine, Roman; Farmer, Andrew D; Sugawara, Masayuki; Young, Nevin D; Sadowsky, Michael J; Tiffin, Peter

    2012-01-01

    The symbiosis between rhizobial bacteria and legume plants has served as a model for investigating the genetics of nitrogen fixation and the evolution of facultative mutualism. We used deep sequence coverage (>100×) to characterize genomic diversity at the nucleotide level among 12 Sinorhizobium medicae and 32 S. meliloti strains. Although these species are closely related and share host plants, based on the ratio of shared polymorphisms to fixed differences we found that horizontal gene transfer (HGT) between these species was confined almost exclusively to plasmid genes. Three multi-genic regions that show the strongest evidence of HGT harbor genes directly involved in establishing or maintaining the mutualism with host plants. In both species, nucleotide diversity is 1.5-2.5 times greater on the plasmids than chromosomes. Interestingly, nucleotide diversity in S. meliloti but not S. medicae is highly structured along the chromosome - with mean diversity (θ(π)) on one half of the chromosome five times greater than mean diversity on the other half. Based on the ratio of plasmid to chromosome diversity, this appears to be due to severely reduced diversity on the chromosome half with less diversity, which is consistent with extensive hitchhiking along with a selective sweep. Frequency-spectrum based tests identified 82 genes with a signature of adaptive evolution in one species or another but none of the genes were identified in both species. Based upon available functional information, several genes identified as targets of selection are likely to alter the symbiosis with the host plant, making them attractive targets for further functional characterization.

  18. Draft genome of an Aerophobetes bacterium reveals a facultative lifestyle in deep-sea anaerobic sediments

    KAUST Repository

    Wang, Yong

    2016-07-01

    Aerophobetes (or CD12) is a recently defined bacterial phylum, of which the metabolic processes and ecological importance remain unclear. In the present study, we obtained the draft genome of an Aerophobetes bacterium TCS1 from saline sediment near the Thuwal cold seep in the Red Sea using a genome binning method. Analysis of 16S rRNA genes of TCS1 and close relatives revealed wide distribution of Aerophobetes in deep-sea sediments. Phylogenetic relationships showed affinity between Aerophobetes TCS1 and some thermophilic bacterial phyla. The genome of TCS1 (at least 1.27 Mbp) contains a full set of genes encoding core metabolic pathways, including glycolysis and pyruvate fermentation to produce acetyl-CoA and acetate. The identification of cross-membrane sugar transporter genes further indicates its potential ability to consume carbohydrates preserved in the sediment under the microbial mat. Aerophobetes bacterium TCS1 therefore probably carried out saccharolytic and fermentative metabolism. The genes responsible for autotrophic synthesis of acetyl-CoA via the Wood–Ljungdahl pathway were also found in the genome. Phylogenetic study of the essential genes for the Wood–Ljungdahl pathway implied relative independence of Aerophobetes bacterium from the known acetogens and methanogens. Compared with genomes of acetogenic bacteria, Aerophobetes bacterium TCS1 genome lacks the genes involved in nitrogen metabolism, sulfur metabolism, signal transduction and cell motility. The metabolic activities of TCS1 might depend on geochemical conditions such as supplies of CO2, hydrogen and sugars, and therefore the TCS1 might be a facultative bacterium in anaerobic saline sediments near cold seeps. © 2016, Science China Press and Springer-Verlag Berlin Heidelberg.

  19. Are hummingbirds facultatively ammonotelic? Nitrogen excretion and requirements as a function of body size.

    Science.gov (United States)

    McWhorter, Todd J; Powers, Donald R; Martínez Del Rio, Carlos

    2003-01-01

    Most birds are uricotelic. An exception to this rule may be nectar-feeding birds, which excrete significant amounts of ammonia under certain conditions. Although ammonia is toxic, because it is highly water soluble its excretion may be facilitated in animals that ingest and excrete large amounts of water. Bird-pollinated plants secrete carbohydrate- and water-rich floral nectars that contain exceedingly little protein. Thus, nectar-feeding birds are faced with the dual challenge of meeting nitrogen requirements while disposing of large amounts of water. The peculiar diet of nectar-feeding birds suggests two hypotheses: (1) these birds must have low protein requirements, and (2) when they ingest large quantities of water their primary nitrogen excretion product may be ammonia. To test these hypotheses, we measured maintenance nitrogen requirements (MNR) and total endogenous nitrogen losses (TENL) in three hummingbird species (Archilochus alexandri, Eugenes fulgens, and Lampornis clemenciae) fed on diets with varying sugar, protein, and water content. We also quantified the form in which the by-products of nitrogen metabolism were excreted. The MNR and TENL of the hummingbirds examined were exceptionally low. However, no birds excreted more than 50% of nitrogen as ammonia or more nitrogen as ammonia than urates. Furthermore, ammonia excretion was not influenced by either water or protein intake. The smallest species (A. alexandri) excreted a significantly greater proportion (>25%) of their nitrogenous wastes as ammonia than the larger hummingbirds ( approximately 4%). Our results support the hypothesis that nectar-feeding birds have low protein requirements but cast doubt on the notion that they are facultatively ammonotelic. Our data also hint at a possible size-dependent dichotomy in hummingbirds, with higher ammonia excretion in smaller species. Differences in proportionate water loads and/or postrenal modification of urine may explain this dichotomy.

  20. 低温污水中耐冷微生物的筛选及多样性分析%Diversity of Psychrotrophs from Low-temperature Sewage

    Institute of Scientific and Technical Information of China (English)

    顾美英; 谢玉清; 唐琦勇; 张志东; 房世杰; 包慧芳; 茆军

    2009-01-01

    采用4种分离培养基对新疆乌鲁木齐市污水处理厂低温污水中的耐冷微生物进行分离筛选,共获得各种耐冷微生物154株,其中丝状真菌12株,酵母46株,放线菌6株,细菌90株.并对部分菌株的耐受性和产酶特性进行了研究,获得了耐4%NaCl的菌株44株,耐0.2%苯酚的16株,耐0.5%SDS的33株,具有淀粉酶活性的菌株40株,蛋白酶活性的30株,脂酶活性的27株.对其中60株细菌进行16S rRNA基因测序,序列比对分析表明,其分属13个属,其中菌株39与不动杆菌属中的标准菌株Acinetobacter lwoffii(DSM2403)同源性最近,为96.6%,极有可能为新种.证明低温污水中存在着丰富的耐冷微生物及种属多样性,且具有较好的耐受性及产酶特性,预示着低温污水可能是重要潜在的微生物资源库.%Psychrotrophs were isolated by using four media from low-temperature sewage of sewage treatment plant in Urumqi, Xinjiang. Totally, 154 strains were obtained including 12 filamentous fungi, 46 yeasts, 6 actinomycetes and 90 bacteria. The results of tolerance tests of the isolates to salt, phenol and SDS, and enzyme producing characters of amylase, proteinase and esterase were shown. Then 60 bacterial strains were chosen for 16S rRNA gene sequencing and analysis. The blasting results showed that the strains were assigned to 13 recognized genera , and the Strain 39 exhibited 96.6% similarity to Acinetobacter Iwof-fii(DSM2403), indicating that it might be a novel species. These results suggested that there were a lot of psychrotrophs and rich bacterial diversity in low-temperature sewage. In addition, which maybe an important and potential library of microbial resources.

  1. Stochastic models of intracellular transport

    KAUST Repository

    Bressloff, Paul C.

    2013-01-09

    The interior of a living cell is a crowded, heterogenuous, fluctuating environment. Hence, a major challenge in modeling intracellular transport is to analyze stochastic processes within complex environments. Broadly speaking, there are two basic mechanisms for intracellular transport: passive diffusion and motor-driven active transport. Diffusive transport can be formulated in terms of the motion of an overdamped Brownian particle. On the other hand, active transport requires chemical energy, usually in the form of adenosine triphosphate hydrolysis, and can be direction specific, allowing biomolecules to be transported long distances; this is particularly important in neurons due to their complex geometry. In this review a wide range of analytical methods and models of intracellular transport is presented. In the case of diffusive transport, narrow escape problems, diffusion to a small target, confined and single-file diffusion, homogenization theory, and fractional diffusion are considered. In the case of active transport, Brownian ratchets, random walk models, exclusion processes, random intermittent search processes, quasi-steady-state reduction methods, and mean-field approximations are considered. Applications include receptor trafficking, axonal transport, membrane diffusion, nuclear transport, protein-DNA interactions, virus trafficking, and the self-organization of subcellular structures. © 2013 American Physical Society.

  2. Enoyl-Acyl Carrier Protein Reductase I (FabI) Is Essential for the Intracellular Growth of Listeria monocytogenes

    Science.gov (United States)

    Ericson, Megan E.; Frank, Matthew W.

    2016-01-01

    Enoyl-acyl carrier protein reductase catalyzes the last step in each elongation cycle of type II bacterial fatty acid synthesis and is a key regulatory protein in bacterial fatty acid synthesis. Genes of the facultative intracellular pathogen Listeria monocytogenes encode two functional enoyl-acyl carrier protein isoforms based on their ability to complement the temperature-sensitive growth phenotype of Escherichia coli strain JP1111 [fabI(Ts)]. The FabI isoform was inactivated by the FabI selective inhibitor AFN-1252, but the FabK isoform was not affected by the drug, as expected. Inhibition of FabI by AFN-1252 decreased endogenous fatty acid synthesis by 80% and lowered the growth rate of L. monocytogenes in laboratory medium. Robust exogenous fatty acid incorporation was not detected in L. monocytogenes unless the pathway was partially inactivated by AFN-1252 treatment. However, supplementation with exogenous fatty acids did not restore normal growth in the presence of AFN-1252. FabI inactivation prevented the intracellular growth of L. monocytogenes, showing that neither FabK nor the incorporation of host cellular fatty acids was sufficient to support the intracellular growth of L. monocytogenes. Our results show that FabI is the primary enoyl-acyl carrier protein reductase of type II bacterial fatty acid synthesis and is essential for the intracellular growth of L. monocytogenes. PMID:27736774

  3. Enoyl-Acyl Carrier Protein Reductase I (FabI) Is Essential for the Intracellular Growth of Listeria monocytogenes.

    Science.gov (United States)

    Yao, Jiangwei; Ericson, Megan E; Frank, Matthew W; Rock, Charles O

    2016-12-01

    Enoyl-acyl carrier protein reductase catalyzes the last step in each elongation cycle of type II bacterial fatty acid synthesis and is a key regulatory protein in bacterial fatty acid synthesis. Genes of the facultative intracellular pathogen Listeria monocytogenes encode two functional enoyl-acyl carrier protein isoforms based on their ability to complement the temperature-sensitive growth phenotype of Escherichia coli strain JP1111 [fabI(Ts)]. The FabI isoform was inactivated by the FabI selective inhibitor AFN-1252, but the FabK isoform was not affected by the drug, as expected. Inhibition of FabI by AFN-1252 decreased endogenous fatty acid synthesis by 80% and lowered the growth rate of L. monocytogenes in laboratory medium. Robust exogenous fatty acid incorporation was not detected in L. monocytogenes unless the pathway was partially inactivated by AFN-1252 treatment. However, supplementation with exogenous fatty acids did not restore normal growth in the presence of AFN-1252. FabI inactivation prevented the intracellular growth of L. monocytogenes, showing that neither FabK nor the incorporation of host cellular fatty acids was sufficient to support the intracellular growth of L. monocytogenes Our results show that FabI is the primary enoyl-acyl carrier protein reductase of type II bacterial fatty acid synthesis and is essential for the intracellular growth of L. monocytogenes. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  4. Characterization of Leptin Intracellular Trafficking

    Directory of Open Access Journals (Sweden)

    E Walum

    2009-12-01

    Full Text Available Leptin is produced by adipose tissue, and its concentration in plasma is related to the amount of fat in the body. The leptin receptor (OBR is a member of the class I cytokine receptor family and several different isoforms, produced by alternative mRNA splicing are found in many tissues, including the hypothalamus. The two predominant isoforms includes a long form (OBRl with an intracellular domain of 303 amino acids and a shorter form (OBRs with an intracellular domain of 34 amino acids. Since OBRl is mainly expressed in the hypotalamus, it has been suggested to be the main signalling form. The peripheral production of leptin by adipocyte tissue and its effects as a signal of satiety in the central nervous system imply that leptin gains access to regions of the brain regulating in energy balance by crossing the blood-brain barrier. In an attempt to characterize the intracellular transport of leptin, we have followed binding internalization and degradation of leptin in HEK293 cells. We have also monitored the intracellular transport pathway of fluorescent conjugated leptin in HEK293 cells. Phenylarsine oxide, a general inhibitor of endocytosis, as well as incubation at mild hypertonic conditions, prevented the uptake of leptin, confirming a receptor-mediated internalization process. When internalized, 125I-leptin was rapidly accumulated inside the cells and reached a maximum after 10 min. After 70 minutes about 40-50% of total counts in each time point were found in the medium as TCA-soluble material. Leptin sorting, at the level of early endosomes, did not seem to involve recycling endosomes, since FITC-leptin was sorted from Cy3- transferrin containing compartments at 37°C. At 45 minutes of continuos internalization, FITC-leptin appeared mainly accumulated in late endocytic structures colocalizing with internalized rhodamine coupled epidermial growth factor (EGF and the lysosomal marker protein lamp-1. The transport of leptin was also shown

  5. Intracardiac intracellular angiotensin system in diabetes.

    NARCIS (Netherlands)

    Kumar, R.; Yong, Q.C.; Thomas, C.M.G.; Baker, K.M.

    2012-01-01

    The renin-angiotensin system (RAS) has mainly been categorized as a circulating and a local tissue RAS. A new component of the local system, known as the intracellular RAS, has recently been described. The intracellular RAS is defined as synthesis and action of ANG II intracellularly. This RAS appea

  6. Control of wilt and rot pathogens of tomato by antagonistic pink pigmented facultative methylotrophic Delftia lacustris and Bacillus spp.

    Directory of Open Access Journals (Sweden)

    Veeranan Janahiraman

    2016-11-01

    Full Text Available The studies on the biocontrol potential of pink pigmented facultative methylotrophic (PPFM bacteria other than the genus Methylobacterium are scarce. In the present study, we report three facultative methylotrophic isolates; PPO-1, PPT-1 and PPB-1, respectively identified as Delftia lacustris, Bacillus subtilis and Bacillus cereus by 16S rRNA gene sequence analysis. Hemolytic activity was tested to investigate the potential pathogenicity of isolates to plants and humans, the results indicates that the isolates PPO-1, PPT-1 and PPB-1 are not pathogenic strains. Under in vitro conditions, D. lacustris PPO-1, B. subtilis PPT-1 and B. cereus PPB-1 showed direct antagonistic effect by inhibiting the mycelial growth of fungal pathogens; Fusarium oxysporum f. sp. lycopersici (2.15, 2.05 and 1.95 cm, Sclerotium rolfsii (2.14, 2.04 and1.94 cm, Pythium ultimum (2.12, 2.02 and 1.92cm, and Rhizoctonia solani (2.18, 2.08 and 1.98 cm and also produced volatile inhibitory compounds. Under plant growth chamber condition methylotrophic bacterial isolates; D. lacustris PPO-1, B. subtilis PPT-1 and B. cereus PPB-1 significantly reduced the disease incidence of tomato. Under greenhouse condition, D. lacustris PPO-1, B. subtilis PPT-1 and B. cereus PPB-1 inoculated tomato plants, when challenged with F. oxysporum f. sp. lycopersici, S. rolfsii, P. ultimum and R. solani, increased the pathogenesis related proteins (β-1, 3-glucanase and chitinase and defense enzymes (phenylalanine ammonia lyase, peroxidase, polyphenol oxidase, and catalase on day 5 after inoculation. In the current study, we first report the facultative methylotrophy in pink pigmented Delftia lacustris, B. subtilis, and B. cereus and their antagonistic potential against fungal pathogens. Direct antagonistic and ISR effects of these isolates against fungal pathogens of tomato evidenced their possible use as a biocontrol agent.

  7. Control of Wilt and Rot Pathogens of Tomato by Antagonistic Pink Pigmented Facultative Methylotrophic Delftia lacustris and Bacillus spp.

    Science.gov (United States)

    Janahiraman, Veeranan; Anandham, Rangasamy; Kwon, Soon W.; Sundaram, Subbiah; Karthik Pandi, Veeranan; Krishnamoorthy, Ramasamy; Kim, Kiyoon; Samaddar, Sandipan; Sa, Tongmin

    2016-01-01

    The studies on the biocontrol potential of pink pigmented facultative methylotrophic (PPFM) bacteria other than the genus Methylobacterium are scarce. In the present study, we report three facultative methylotrophic isolates; PPO-1, PPT-1, and PPB-1, respectively, identified as Delftia lacustris, Bacillus subtilis, and Bacillus cereus by 16S rRNA gene sequence analysis. Hemolytic activity was tested to investigate the potential pathogenicity of isolates to plants and humans, the results indicates that the isolates PPO-1, PPT-1, and PPB-1 are not pathogenic strains. Under in vitro conditions, D. lacustris PPO-1, B. subtilis PPT-1, and B. cereus PPB-1 showed direct antagonistic effect by inhibiting the mycelial growth of fungal pathogens; Fusarium oxysporum f. sp. lycopersici (2.15, 2.05, and 1.95 cm), Sclerotium rolfsii (2.14, 2.04, and 1.94 cm), Pythium ultimum (2.12, 2.02, and 1.92 cm), and Rhizoctonia solani (2.18, 2.08, and 1.98 cm) and also produced volatile inhibitory compounds. Under plant growth chamber condition methylotrophic bacterial isolates; D. lacustris PPO-1, B. subtilis PPT-1, and B. cereus PPB-1 significantly reduced the disease incidence of tomato. Under greenhouse condition, D. lacustris PPO-1, B. subtilis PPT-1, and B. cereus PPB-1 inoculated tomato plants, when challenged with F. oxysporum f. sp. lycopersici, S. rolfsii, P. ultimum, and R. solani, increased the pathogenesis related proteins (β-1,3-glucanase and chitinase) and defense enzymes (phenylalanine ammonia lyase, peroxidase, polyphenol oxidase, and catalase) on day 5 after inoculation. In the current study, we first report the facultative methylotrophy in pink pigmented D. lacustris, B. subtilis, and B. cereus and their antagonistic potential against fungal pathogens. Direct antagonistic and ISR effects of these isolates against fungal pathogens of tomato evidenced their possible use as a biocontrol agent. PMID:27872630

  8. HiPIP oxido-reductase activity in membranes from aerobically grown cells of the facultative phototroph Rhodoferax fermentans

    DEFF Research Database (Denmark)

    Hochkoeppler, Alejandro; Kofod, Pauli; Zannoni, Davide

    1995-01-01

    The role of the periplasmically located, water-soluble, HiPIP (high-potential iron-sulfur protein) in the respiratory chain of the facultative phototroph Rhodoferax fermentans has been examined. The oxidized HiPIP is reduced by succinate-dependent respiration via the bc 1 complex, this reaction...... being inhibited by myxothiazol and/or stigmatellin. The reduced HiPIP can be oxidized by the membrane-bound cytochrome oxidase, this reaction being inhibited by 0.1 mM cyanide. We conclude that aerobically grown Rf. fermentans contains a redox chain in which HiPIP mediates electron transfer between...

  9. Comparative antimicrobial susceptibility of aerobic and facultative bacteria from community-acquired bacteremia to ertapenem in Taiwan

    Directory of Open Access Journals (Sweden)

    Fung Chang-Phone

    2007-07-01

    Full Text Available Abstract Background Ertapenem is a once-a-day carbapenem and has excellent activity against many gram-positive and gram-negative aerobic, facultative, and anaerobic bacteria. The susceptibility of isolates of community-acquired bacteremia to ertapenem has not been reported yet. The present study assesses the in vitro activity of ertapenem against aerobic and facultative bacterial pathogens isolated from patients with community-acquired bacteremia by determining and comparing the MICs of cefepime, cefoxitin, ceftazidime, ceftriaxone, ertapenem, piperacillin, piperacillin-tazobactam, ciprofloxacin, amikacin and gentamicin. The prevalence of extended broad spectrum β-lactamases (ESBL producing strains of community-acquired bacteremia and their susceptibility to these antibiotics are investigated. Methods Aerobic and facultative bacteria isolated from blood obtained from hospitalized patients with community-acquired bacteremia within 48 hours of admission between August 1, 2004 and September 30, 2004 in Chang Gung Memorial Hospital at Keelung, Taiwan, were identified using standard procedures. Antimicrobial susceptibility was evaluated by Etest according to the standard guidelines provided by the manufacturer and document M100-S16 Performance Standards of the Clinical Laboratory of Standard Institute. Antimicrobial agents including cefepime, cefoxitin, ceftazidime, ceftriaxone, ertapenem, piperacillin, piperacillin-tazobactam, ciprofloxacin, amikacin and gentamicin were used against the bacterial isolates to test their MICs as determined by Etest. For Staphylococcus aureus isolates, MICs of oxacillin were also tested by Etest to differentiate oxacillin-sensitive and oxacillin-resistant S. aureus. Results Ertapenem was highly active in vitro against many aerobic and facultative bacterial pathogens commonly recovered from patients with community-acquired bacteremia (128/159, 80.5 %. Ertapenem had more potent activity than ceftriaxone, piperacillin

  10. Swedish isolates of Vibrio cholerae enhance their survival when interacted intracellularly with Acanthamoeba castellanii

    Directory of Open Access Journals (Sweden)

    Salah Shanan

    2016-04-01

    Full Text Available Vibrio cholerae is a Gram-negative bacterium that occurs naturally in aquatic environment. Only V. cholerae O1 and V. cholerae O139 produce cholera toxin and cause cholera, other serogroups can cause gastroenteritis, open wounds infection, and septicaemia. V. cholerae O1 and V. cholerae O139 grow and survive inside Acanthamoeba castellanii. The aim of this study is to investigate the interactions of the Swedish clinical isolates V. cholerae O3, V. cholerae O4, V. cholerae O5, V. cholerae O11, and V. cholerae O160 with A. castellanii. The interaction between A. castellanii and V. cholerae strains was studied by means of amoeba cell counts, viable counts of the bacteria in the absence or presence of amoebae, and of the intracellularly growing bacteria, visualised by electron microscopy. These results show that all V. cholerae can grow and survive outside and inside the amoebae, disclosing that V. cholerae O3, V. cholerae O4, V. cholerae O5, V. cholerae O11, and V. cholerae O160 all can be considered as facultative intracellular bacteria.

  11. Swedish isolates of Vibrio cholerae enhance their survival when interacted intracellularly with Acanthamoeba castellanii.

    Science.gov (United States)

    Shanan, Salah; Bayoumi, Magdi; Saeed, Amir; Sandström, Gunnar; Abd, Hadi

    2016-01-01

    Vibrio cholerae is a Gram-negative bacterium that occurs naturally in aquatic environment. Only V. cholerae O1 and V. cholerae O139 produce cholera toxin and cause cholera, other serogroups can cause gastroenteritis, open wounds infection, and septicaemia. V. cholerae O1 and V. cholerae O139 grow and survive inside Acanthamoeba castellanii. The aim of this study is to investigate the interactions of the Swedish clinical isolates V. cholerae O3, V. cholerae O4, V. cholerae O5, V. cholerae O11, and V. cholerae O160 with A. castellanii. The interaction between A. castellanii and V. cholerae strains was studied by means of amoeba cell counts, viable counts of the bacteria in the absence or presence of amoebae, and of the intracellularly growing bacteria, visualised by electron microscopy. These results show that all V. cholerae can grow and survive outside and inside the amoebae, disclosing that V. cholerae O3, V. cholerae O4, V. cholerae O5, V. cholerae O11, and V. cholerae O160 all can be considered as facultative intracellular bacteria.

  12. Conjugal transfer of a virulence plasmid in the opportunistic intracellular actinomycete Rhodococcus equi.

    Science.gov (United States)

    Tripathi, V N; Harding, W C; Willingham-Lane, J M; Hondalus, M K

    2012-12-01

    Rhodococcus equi is a facultative intracellular, Gram-positive, soilborne actinomycete which can cause severe pyogranulomatous pneumonia with abscessation in young horses (foals) and in immunocompromised people, such as persons with AIDS. All strains of R. equi isolated from foals and approximately a third isolated from humans contain a large, ~81-kb plasmid which is essential for the intramacrophage growth of the organism and for virulence in foals and murine in vivo model systems. We found that the entire virulence plasmid could be transferred from plasmid-containing strains of R. equi (donor) to plasmid-free R. equi strains (recipient) at a high frequency and that plasmid transmission reestablished the capacity for intracellular growth in macrophages. Plasmid transfer required living cells and cell-to-cell contact and was unaffected by the presence of DNase, factors pointing to conjugation as the major means of genetic transfer. Deletion of a putative relaxase-encoding gene, traA, located in the proposed conjugative region of the plasmid, abolished plasmid transfer. Reversion of the traA mutation restored plasmid transmissibility. Finally, plasmid transmission to other Rhodococcus species and some additional related organisms was demonstrated. This is the first study showing a virulence plasmid transfer in R. equi, and it establishes a mechanism by which the virulence plasmid can move among bacteria in the soil.

  13. Modulation of Macrophage Inflammatory Nuclear Factor κB (NF-κB) Signaling by Intracellular Cryptococcus neoformans.

    Science.gov (United States)

    Hayes, James B; Sircy, Linda M; Heusinkveld, Lauren E; Ding, Wandi; Leander, Rachel N; McClelland, Erin E; Nelson, David E

    2016-07-22

    Cryptococcus neoformans (Cn) is a common facultative intracellular pathogen that can cause life-threatening fungal meningitis in immunocompromised individuals. Shortly after infection, Cn is detectable as both extra- and intracellular yeast particles, with Cn being capable of establishing long-lasting latent infections within host macrophages. Although recent studies have shown that shed capsular polysaccharides and intact extracellular Cn can compromise macrophage function through modulation of NF-κB signaling, it is currently unclear whether intracellular Cn also affects NF-κB signaling. Utilizing live cell imaging and computational modeling, we find that extra- and intracellular Cn support distinct modes of NF-κB signaling in cultured murine macrophages. Specifically, in RAW 264.7 murine macrophages treated with extracellular glucuronoxylomannan (GXM), the major Cn capsular polysaccharide, LPS-induced nuclear translocation of p65 is inhibited, whereas in cells with intracellular Cn, LPS-induced nuclear translocation of p65 is both amplified and sustained. Mathematical simulations and quantification of nascent protein expression indicate that this is a possible consequence of Cn-induced "translational interference," impeding IκBα resynthesis. We also show that long term Cn infection induces stable nuclear localization of p65 and IκBα proteins in the absence of additional pro-inflammatory stimuli. In this case, nuclear localization of p65 is not accompanied by TNFα or inducible NOS (iNOS) expression. These results demonstrate that capsular polysaccharides and intact intracellular yeast manipulate NF-κB via multiple distinct mechanisms and provide new insights into how Cn might modulate cellular signaling at different stages of an infection.

  14. Contagem, isolamento e caracterização de bactérias psicrotróficas contaminantes de leite cru refrigerado Counting, isolation and characterization of psychrotrophic bacteria from refrigerated raw milk

    Directory of Open Access Journals (Sweden)

    Edna Froeder Arcuri

    2008-11-01

    Full Text Available Com os objetivos de quantificar, isolar e caracterizar bactérias psicrotróficas contaminantes de leite cru refrigerado, produzido na região da Zona da Mata de Minas Gerais e Sudeste do Rio de Janeiro, foram analisadas amostras de leite coletadas de 20 tanques coletivos e 23 tanques individuais. As contagens de bactérias psicrotróficas nas amostras de leite para os dois tipos de tanques de refrigeração variaram entre 10² e 10(7 Unidades Formadoras de Colônias (UFC ml-1, porém, um maior número de tanques coletivos apresentou contagens acima de 1 x 10(5 UFC ml-1. Foi verificada a predominância de bactérias psicrotróficas gram-negativas (81,2%, que foram identificadas pelos sistemas API 20E e API 20NE nos gêneros: Aeromonas, Alcaligenes, Acinetobacter, Burkholderia,Chryseomonas, Enterobacter, Ewingella, Klebsiella, Hafnia, Methylobacterium, Moraxella, Pantoea, Pseudomonas, Serratia, Sphingomonas e Yersinia. As bactérias gram-positivas (18,8% foram identificadas com API 50 CH, API Coryne e API Staph, nos gêneros: Bacillus, Brevibacterium, Cellum/Microbacterium, Kurthia e Staphylococcus. Os sistemas API utilizados não identificaram todos os isolados bacterianos. Pseudomonas foi o gênero mais isolado e P. fluorescens foi a espécie predominante. A maioria dos isolados bacterianos apresentou atividade proteolítica e/ou lipolítica a temperaturas de refrigeração de 4°C, 7°C e 10°C, evidenciando seu alto potencial de deterioração do leite e dos produtos lácteos. Os resultados ressaltam que maior atenção deve ser dada aos procedimentos que impeçam a contaminação do leite por esses microrganismos.This study aimed to quantify, isolate and characterize psychrotrophic bacteria from refrigerated raw milk produced at the ‘Mata’ Region of Minas Gerais State and Southeast of Rio de Janeiro State, Brazil. Raw milk samples, were collected at the farms, from 20 collective refrigerated tanks and 23 individual refrigerated tanks

  15. The 'male escape hypothesis': sex-biased metamorphosis in response to climatic drivers in a facultatively paedomorphic amphibian.

    Science.gov (United States)

    Mathiron, Anthony G E; Lena, Jean-Paul; Baouch, Sarah; Denoël, Mathieu

    2017-04-26

    Paedomorphosis is a major evolutionary process that bypasses metamorphosis and allows reproduction in larvae. In newts and salamanders, it can be facultative with paedomorphs retaining gills and metamorphs dispersing. The evolution of these developmental processes is thought to have been driven by the costs and benefits of inhabiting aquatic versus terrestrial habitats. In this context, we aimed at testing the hypothesis that climatic drivers affect phenotypic transition and the difference across sexes because sex-ratio is biased in natural populations. Through a replicated laboratory experiment, we showed that paedomorphic palmate newts (Lissotriton helveticus) metamorphosed at a higher frequency when water availability decreased and metamorphosed earlier when temperature increased in these conditions. All responses were sex-biased, and males were more prone to change phenotype than females. Our work shows how climatic variables can affect facultative paedomorphosis and support theoretical models predicting life on land instead of in water. Moreover, because males metamorphose and leave water more often and earlier than females, these results, for the first time, give an experimental explanation for the rarity of male paedomorphosis (the 'male escape hypothesis') and suggest the importance of sex in the evolution of paedomorphosis versus metamorphosis. © 2017 The Author(s).

  16. Effects of gamma ray and electron-beam irradiations on survival of anaerobic and facultatively anaerobic bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Miyahara, Michiko; Miyahara, Makoto [National Inst. of Health Sciences, Tokyo (Japan)

    2002-10-01

    An extension of the approval for food irradiation is desired due to the increase in the incidence of food poisoning in the world. One anaerobic (Clostridium perfringens) and four facultatively anaerobic (Bacillus cereus, Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella Enteritidis) bacteria irradiated with gamma ray or electron beam (E-beam) were tested in terms of survival on agar under packaging atmosphere. Using pouch pack, effects of two irradiations on survival of anaerobic and facultatively anaerobic bacteria were evaluated comparatively. E-beam irradiation was more effective than gamma ray irradiation in decreasing the lethal dose 10% (D{sub 10}) value of B. cereus at 4 deg C, slightly more effective in that of E. coli O157, and similarly effective in that of the other three bacteria at 4 deg C. The gamma irradiation of the bacteria without incubation at 4 deg C before irradiation was more effective than that of the bacteria with incubation overnight at 4 deg C before irradiation in decreasing the D10 values of these bacteria (B. cereus, E. coli O157, and L. monocytogenes). Furthermore, ground beef patties inoculated with bacteria were irradiated with 1 kGy by E-beam (5 MeV) at 4 deg C. The inoculated bacteria in the 1-9 mm beef patties were killed by 1 kGy E-beam irradiation and some bacteria in more than 9 mm beef patties were not killed by the irradiation. (author)

  17. Familiarity is more important than phenotypic similarity in shaping social relationships in a facultative female dispersed primate, Colobus vellerosus.

    Science.gov (United States)

    Wikberg, Eva C; Ting, Nelson; Sicotte, Pascale

    2014-07-01

    Animals often bias affiliative behaviors toward kin, but it is unclear what mechanism most species use to discriminate kin. We investigated if facultative dispersed female primates use phenotype matching and/or familiarity to discriminate female kin. We studied 38 adult female Colobus vellerosus at Boabeng-Fiema, Ghana. We determined dyadic co-residency status and age proximity using long-term demographic data, R-values from 17 short tandem repeat loci, and interaction rates using focal samples collected during one year. Approach rates were not strongly affected by how long females had resided together, which contrasts to the familiarity hypothesis. Females approached and groomed maternal kin more than other females, which supports the mother-mediated familiarity hypothesis. Females did not discriminate paternal half siblings from non-kin, and they did not prefer to interact with females of similar age. Short-term co-resident kin did not bias affiliation toward each other, indicating that female colobus cannot consistently recognize less familiar kin via phenotype matching or that biasing behaviors toward less familiar kin is not beneficial. Despite showing facultative dispersal that may reduce the accuracy of using familiarity as a kin recognition mechanism, female choice of social partners was based on familiarity, which conforms to the pattern observed in many female philopatric primates.

  18. Mathematical Analysis for the Optimization of a Design in a Facultative Pond: Indicator Organism and Organic Matter

    Directory of Open Access Journals (Sweden)

    Facundo Cortés Martínez

    2014-01-01

    Full Text Available Stabilization ponds are easy to operate and their maintenance is simple. Treatment is carried out naturally and they are recommended in developing countries. The main disadvantage of these systems is the large land area they occupy. The aim of this study was to perform an optimization in the design and cost of a facultative pond, considering a mathematical analysis of the traditional methodology to determine the model constraints (fecal coliforms and organic matter. Matlab optimization toolbox was used for nonlinear programming. A facultative pond with the traditional method was designed and then the optimization system was applied. Both analyses meet the treated water quality requirements for the discharge to the receiving bodies. The results show a reduction of hydraulic retention time by 4.82 days, and a decrease in the area of 17.9 percent over the traditional method. A sensitivity analysis of the mathematical model is included. It is recommended to realize a full-scale study in order to verify the results of the optimization.

  19. Distribution and dietary regulation of an associated facultative Rhizobiales-related bacterium in the omnivorous giant tropical ant, Paraponera clavata

    Science.gov (United States)

    Larson, Hannah K.; Goffredi, Shana K.; Parra, Erica L.; Vargas, Orlando; Pinto-Tomas, Adrián A.; McGlynn, Terrence P.

    2014-05-01

    We document a facultative Bartonella-like Rhizobiales bacterium in the giant tropical ant, Paraponera clavata. In a lowland tropical rainforest in Costa Rica, 59 colonies were assayed for the prevalence of the Bartonella-like bacterium (BLB), 14 of which were positive. We addressed three questions: First, how does the prevalence of BLB within colonies vary with environmental conditions? Second, how does diet affect the prevalence of BLB in P. clavata? Third, how does the distribution of BLB among colonies reflect ambient differences in food resources and foraging habits? A variety of environmental variables that may be predictive of the presence of BLB were measured, and diet manipulations were conducted to test whether the prevalence of BLB responded to supplemental carbohydrate or prey. The ambient frequency of BLB is much higher in young secondary forests, but is nearly absent from older secondary forests. The prevalence of BLB inside field colonies increased over the duration of a 2-week carbohydrate supplementation; however, water and prey supplementation did not alter the prevalence of BLB. The diets of the colonies located in young secondary forest, compared to other habitats, have a diet richer in carbohydrates and lower in prey. The abundance of carbohydrate, or the relative lack of N, in a colony's diet influences the occurrence of the BLB microbe in P. clavata. As experimental diet manipulations can affect the facultative presence of an N-cycling microbe, a consistent diet shift in diet may facilitate the emergence of tighter symbioses.

  20. Discordant coral-symbiont structuring: factors shaping geographical variation of Symbiodinium communities in a facultative zooxanthellate coral genus, Oculina

    Science.gov (United States)

    Leydet, Karine Posbic; Hellberg, Michael E.

    2016-06-01

    Understanding the factors that help shape the association between corals and their algal symbionts, zooxanthellae ( Symbiodinium), is necessary to better understand the functional diversity and acclimatization potential of the coral host. However, most studies focus on tropical zooxanthellate corals and their obligate algal symbionts, thus limiting our full comprehension of coral-algal symbiont associations. Here, we examine algal associations in a facultative zooxanthellate coral. We survey the Symbiodinium communities associated with Oculina corals in the western North Atlantic and the Mediterranean using one clade-level marker ( psbA coding region) and three fine-scale markers ( cp23S- rDNA, b7sym15 flanking region, and b2sym17). We ask whether Oculina spp. harbor geographically different Symbiodinium communities across their geographic range and, if so, whether the host's genetics or habitat differences are correlated with this geographical variation. We found that Oculina corals harbor different Symbiodinium communities across their geographical range. Of the habitat differences (including chlorophyll a concentration and depth), sea surface temperature is better correlated with this geographical variation than the host's genetics, a pattern most evident in the Mediterranean. Our results suggest that although facultative zooxanthellate corals may be less dependent on their algal partners compared to obligate zooxanthellate corals, the Symbiodinium communities that they harbor may nevertheless reflect acclimatization to environmental variation among habitats.

  1. Enhanced biohydrogen production from beverage industrial wastewater using external nitrogen sources and bioaugmentation with facultative anaerobic strains.

    Science.gov (United States)

    Kumar, Gopalakrishnan; Bakonyi, Péter; Sivagurunathan, Periyasamy; Kim, Sang-Hyoun; Nemestóthy, Nándor; Bélafi-Bakó, Katalin; Lin, Chiu-Yue

    2015-08-01

    In this work biohydrogen generation and its improvement possibilities from beverage industrial wastewater were sought. Firstly, mesophilic hydrogen fermentations were conducted in batch vials by applying heat-treated (80°C, 30 min) sludge and liquid (LB-grown) cultures of Escherichia coli XL1-Blue/Enterobacter cloacae DSM 16657 strains for bioaugmentation purposes. The results showed that there was a remarkable increase in hydrogen production capacities when facultative anaerobes were added in the form of inoculum. Furthermore, experiments were carried out in order to reveal whether the increment occurred either due to the efficient contribution of the facultative anaerobic microorganisms or the culture ingredients (in particular yeast extract and tryptone) supplied when the bacterial suspensions (LB media-based inocula) were mixed with the sludge. The outcome of these tests was that both the applied nitrogen sources and the bacteria (E. coli) could individually enhance hydrogen formation. Nevertheless, the highest increase took place when they were used together. Finally, the optimal initial wastewater concentration was determined as 5 g/L.

  2. 长春南湖水耐冷菌的筛选及其产酶特性%Screening and Properties of Enzyme Production of Psychrotrophs from Nanhu Lake of Changchun

    Institute of Scientific and Technical Information of China (English)

    黄继国; 杨柳; 王玉; 朱婧; 丛伟

    2011-01-01

    The samples from the South Lake at a low temperature were taken and diluted onto coated tablet for concentration inoculation. A total of 35 cold-adapted-strains were isolated, which contained 14 bacteria and 21 fungi. The colony morphology and physiological and biochemical results show that the psychrotrophic bacteria belong to Lactobacilus, Saccharococcus, Staphyloccus, Actinomyces and the fungi belong to molds and yeast.Some of them can produce hydrolases including oxidase, catalase, amylase and gelatinase, and some of psychrotrophs can make nitrate reduction, nitrite reduction and so on.%采用低温环境下的南湖湖水为样品,稀释后涂布平板进行富集培养,并进行细菌菌落形态及生理生化性质测定.实验共培养出耐冷菌35株,包括细菌14株,真菌21株.其中细菌分属于乳杆菌属、葡萄球菌属、糖球菌属、放线菌属;真菌分属于霉菌和酵母菌.分离的菌种能产生多种大分子物质水解酶类,包括氧化酶、接触酶、淀粉水解酶、明胶酶等,并且大多数菌种具有硝酸盐还原作用阳性,还有一些耐冷菌具有亚硝酸盐还原作用阳性、反硝化作用阳性等.

  3. 原料乳嗜冷菌分泌热稳定性酶类检测方法的研究进展%Advancement on Rapid Detection Method for Psychrotrophic Heat Stable Extracellular Enzymes from Raw Milk

    Institute of Scientific and Technical Information of China (English)

    黄丽; 赵玉娟; 李盛钰

    2015-01-01

    嗜冷菌是导致原料乳以及乳制品腐败的主要微生物类群。巴氏杀菌或超高温瞬时(UHT)灭菌后,几乎除去了全部的嗜冷菌,但细菌分泌的热稳定的蛋白酶和脂肪酶却并未完全钝化,进一步影响原料乳风味以及质地。测定原料奶中热稳定蛋白酶和脂肪酶的活性是控制嗜冷菌污染原料奶的前提。本文综述了原料乳中嗜冷菌分泌的热稳定性酶类的快速检测技术,比较了这些方法的优缺点,并对此领域的技术发展方向进行了展望。%Psychrotrophic microorganisms play a leading role in spoilage of raw milk and milk products. Pasteurized and ultra high temperature sterilization is usually used to kill psychrophile, but they are unable to remove the heat stable protease and lipase produced by the psychrophile, ifnally to affect the lfavor and texture of dairy products. Detecting the activities of heat stable protease and lipase is the base of controlling contamination of psychrophile. The present study summarizes and discusses some rapid detection methods of psychrotrophic heat stable extracellular enzymes in raw milk. The advantages and disadvantages in industrial application are also compared, and the development rapid, sensitive, and meaningful directions of those techniques in this ifeld are discussed.

  4. Palladium-mediated intracellular chemistry

    Science.gov (United States)

    Yusop, Rahimi M.; Unciti-Broceta, Asier; Johansson, Emma M. V.; Sánchez-Martín, Rosario M.; Bradley, Mark

    2011-03-01

    Many important intracellular biochemical reactions are modulated by transition metals, typically in the form of metalloproteins. The ability to carry out selective transformations inside a cell would allow researchers to manipulate or interrogate innumerable biological processes. Here, we show that palladium nanoparticles trapped within polystyrene microspheres can enter cells and mediate a variety of Pd0-catalysed reactions, such as allylcarbamate cleavage and Suzuki-Miyaura cross-coupling. The work provides the basis for the customization of heterogeneous unnatural catalysts as tools to carry out artificial chemistries within cells. Such in cellulo synthesis has potential for a plethora of applications ranging from cellular labelling to synthesis of modulators or inhibitors of cell function.

  5. Misregulation of the broad-range phospholipase C activity increases the susceptibility of Listeria monocytogenes to intracellular killing by neutrophils.

    Science.gov (United States)

    Blank, Bryant S; Abi Abdallah, Delbert S; Park, Justin J; Nazarova, Evgeniya V; Pavinski Bitar, Alan; Maurer, Kirk J; Marquis, Hélène

    2014-02-01

    Listeria monocytogenes is a facultative intracellular bacterial pathogen that tightly regulates the activities of various virulence factors during infection. A mutant strain (the plcBDpro mutant) that has lost the ability to control the activity of a phospholipase C (PC-PLC) is attenuated a hundred fold in mice. This attenuation is not due to a lack of bacterial fitness, but appears to result from a modified host response to infection. The transcriptomic pattern of immune-related genes indicated that PC-PLC did not enhance the innate immune response in infected macrophages. However, it partially protected the cells from bacteria-mediated mitochondrial fragmentation. In mice, the plcBDpro mutant transiently caused an increase in liver pathology, as judged by the size of neutrophil-filled micro-abscesses. Moreover, the plcBDpro mutant was more susceptible to intracellular killing by neutrophils than wild-type L. monocytogenes. Together, these data indicate that in vivo attenuation of the plcBDpro mutant results from its reduced ability to disrupt mitochondrial homeostasis and to resist intracellular killing by neutrophils.

  6. Intracellular peptides: From discovery to function

    Directory of Open Access Journals (Sweden)

    Emer S. Ferro

    2014-06-01

    Full Text Available Peptidomics techniques have identified hundreds of peptides that are derived from proteins present mainly in the cytosol, mitochondria, and/or nucleus; these are termed intracellular peptides to distinguish them from secretory pathway peptides that function primarily outside of the cell. The proteasome and thimet oligopeptidase participate in the production and metabolism of intracellular peptides. Many of the intracellular peptides are common among mouse tissues and human cell lines analyzed and likely to perform a variety of functions within cells. Demonstrated functions include the modulation of signal transduction, mitochondrial stress, and development; additional functions will likely be found for intracellular peptides.

  7. EFFECT OF TIME AND TEMPERATURE OF REFRIGERATION IN THE DEVELOPMENT OF PSYCHROTROPHIC MICRORGANISMS IN COOLED RAW MILK COLLECTED IN THE MACROREGION OF GOIÂNIA – GO EFEITO DO TEMPO E DA TEMPERATURA DE REFRIGERAÇÃO NO DESENVOLVIMENTO DE MICRORGANISMOS PSICROTRÓFICOS EM LEITE CRU REFRIGERADO COLETADO NA MACRORREGIÃO DE GOIÂNIA – GO

    Directory of Open Access Journals (Sweden)

    Antônio Nonato de Oliveira

    2009-12-01

    Full Text Available

    The objective of the research were evaluated the effect of storage time and of refrigeration temperature under the counting of psychrotrophics microrganisms and proteolytics psychrotrophics in refrigerated raw milk samples. In the period of August the September of 2005 34 refrigerated raw milk samples were collected of refrigeration tanks, being 29 individuals and five collectives. Milk was proceeding from bovine flocks. For the collection of the samples followed a route determined for the industry located in the macroregion of Goiânia – GO, Brazil. For the microbiological evaluation was carried the standard plate count method of psychrotrophics microrganisms and proteolytics psychrotrophics. For the determination and quantification of the glycomacropeptide (GMP were adopted methodology proposal in Brazil (1991. The counting of psychrotrophics microrganisms and proteolytics psychrotrophics increased with the storage time of refrigerated bovine milk. Were observed the inadequate tanks were observed e presence of GMP for the High Performance Liquid Chromatography (HPLC was not evidenced.

    KEY WORDS: Proteolysis, Pseudomonas spp, raw milk.
    O objetivo da pesquisa foi avaliar o efeito do tempo de estocagem e da temperatura de refrigeração sob a contagem de microrganismos psicrotróficos e psicrotróficos proteolíticos em amostras de leite cru refrigerado. No período de agosto a setembro de 2005 coletaram-se 34 amostras de leite cru refrigerado de tanques de expansão, sendo 29 individuais e cinco coletivos. Para a coleta das amostras seguiu-se uma rota determinada pela indústria localizada na macrorregião de Goiânia, GO. Para a avaliação microbiológica, foi realizada a contagem-padrão em placas de microrganismos psicrotróficos e psicrotróficos proteolíticos. Para a determinação e quantificação do caseinomacropeptídeo (CMP, adotou-se a metodologia proposta em BRASIL (1991. A contagem de

  8. The complete genome of Teredinibacter turnerae T7901: an intracellular endosymbiont of marine wood-boring bivalves (shipworms).

    Science.gov (United States)

    Yang, Joyce C; Madupu, Ramana; Durkin, A Scott; Ekborg, Nathan A; Pedamallu, Chandra S; Hostetler, Jessica B; Radune, Diana; Toms, Bradley S; Henrissat, Bernard; Coutinho, Pedro M; Schwarz, Sandra; Field, Lauren; Trindade-Silva, Amaro E; Soares, Carlos A G; Elshahawi, Sherif; Hanora, Amro; Schmidt, Eric W; Haygood, Margo G; Posfai, Janos; Benner, Jack; Madinger, Catherine; Nove, John; Anton, Brian; Chaudhary, Kshitiz; Foster, Jeremy; Holman, Alex; Kumar, Sanjay; Lessard, Philip A; Luyten, Yvette A; Slatko, Barton; Wood, Nicole; Wu, Bo; Teplitski, Max; Mougous, Joseph D; Ward, Naomi; Eisen, Jonathan A; Badger, Jonathan H; Distel, Daniel L

    2009-07-01

    Here we report the complete genome sequence of Teredinibacter turnerae T7901. T. turnerae is a marine gamma proteobacterium that occurs as an intracellular endosymbiont in the gills of wood-boring marine bivalves of the family Teredinidae (shipworms). This species is the sole cultivated member of an endosymbiotic consortium thought to provide the host with enzymes, including cellulases and nitrogenase, critical for digestion of wood and supplementation of the host's nitrogen-deficient diet. T. turnerae is closely related to the free-living marine polysaccharide degrading bacterium Saccharophagus degradans str. 2-40 and to as yet uncultivated endosymbionts with which it coexists in shipworm cells. Like S. degradans, the T. turnerae genome encodes a large number of enzymes predicted to be involved in complex polysaccharide degradation (>100). However, unlike S. degradans, which degrades a broad spectrum (>10 classes) of complex plant, fungal and algal polysaccharides, T. turnerae primarily encodes enzymes associated with deconstruction of terrestrial woody plant material. Also unlike S. degradans and many other eubacteria, T. turnerae dedicates a large proportion of its genome to genes predicted to function in secondary metabolism. Despite its intracellular niche, the T. turnerae genome lacks many features associated with obligate intracellular existence (e.g. reduced genome size, reduced %G+C, loss of genes of core metabolism) and displays evidence of adaptations common to free-living bacteria (e.g. defense against bacteriophage infection). These results suggest that T. turnerae is likely a facultative intracellular ensosymbiont whose niche presently includes, or recently included, free-living existence. As such, the T. turnerae genome provides insights into the range of genomic adaptations associated with intracellular endosymbiosis as well as enzymatic mechanisms relevant to the recycling of plant materials in marine environments and the production of cellulose

  9. The complete genome of Teredinibacter turnerae T7901: an intracellular endosymbiont of marine wood-boring bivalves (shipworms.

    Directory of Open Access Journals (Sweden)

    Joyce C Yang

    Full Text Available Here we report the complete genome sequence of Teredinibacter turnerae T7901. T. turnerae is a marine gamma proteobacterium that occurs as an intracellular endosymbiont in the gills of wood-boring marine bivalves of the family Teredinidae (shipworms. This species is the sole cultivated member of an endosymbiotic consortium thought to provide the host with enzymes, including cellulases and nitrogenase, critical for digestion of wood and supplementation of the host's nitrogen-deficient diet. T. turnerae is closely related to the free-living marine polysaccharide degrading bacterium Saccharophagus degradans str. 2-40 and to as yet uncultivated endosymbionts with which it coexists in shipworm cells. Like S. degradans, the T. turnerae genome encodes a large number of enzymes predicted to be involved in complex polysaccharide degradation (>100. However, unlike S. degradans, which degrades a broad spectrum (>10 classes of complex plant, fungal and algal polysaccharides, T. turnerae primarily encodes enzymes associated with deconstruction of terrestrial woody plant material. Also unlike S. degradans and many other eubacteria, T. turnerae dedicates a large proportion of its genome to genes predicted to function in secondary metabolism. Despite its intracellular niche, the T. turnerae genome lacks many features associated with obligate intracellular existence (e.g. reduced genome size, reduced %G+C, loss of genes of core metabolism and displays evidence of adaptations common to free-living bacteria (e.g. defense against bacteriophage infection. These results suggest that T. turnerae is likely a facultative intracellular ensosymbiont whose niche presently includes, or recently included, free-living existence. As such, the T. turnerae genome provides insights into the range of genomic adaptations associated with intracellular endosymbiosis as well as enzymatic mechanisms relevant to the recycling of plant materials in marine environments and the production

  10. Effects of age and macrophage lineage on intracellular survival and cytokine induction after infection with Rhodococcus equi.

    Science.gov (United States)

    Berghaus, Londa J; Giguère, Steeve; Sturgill, Tracy L

    2014-07-15

    Rhodococcus equi, a facultative intracellular pathogen of macrophages, causes life-threatening pneumonia in foals and in people with underlying immune deficiencies. As a basis for this study, we hypothesized that macrophage lineage and age would affect intracellular survival of R. equi and cytokine induction after infection. Monocyte-derived and bronchoalveolar macrophages from 10 adult horses and from 10 foals (sampled at 1-3 days, 2 weeks, 1 month, 3 months, and 5 months of age) were infected ex vivo with virulent R. equi. Intracellular R. equi were quantified and mRNA expression of IL-1β, IL-4, IL-6, IL-8, IL-10, IL-12 p40, IL-18, IFN-γ, and TNF-α was measured. Intracellular replication of R. equi was significantly (Pequi was significantly (P=0.002) higher in 3-month-old foals than in 3-day old foals, 2-week-old foals, 1-month-old foals, and adult horses. Expression of IL-4 mRNA was significantly higher in monocyte-derived macrophages whereas expression of IL-6, IL-18, and TNF-α was significantly higher in bronchoalveolar macrophages. Induction of IL-1β, IL-10, IL-12 p40, and IL-8 mRNA in bronchoalveolar macrophages of 1-3-day old foals was significantly higher than in older foals or adult horses. Preferential intracellular survival of R. equi in bronchoalveolar macrophages of juvenile horses may play a role in the pulmonary tropism of the pathogen and in the window of age susceptibility to infection. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Hydrophilic fluorescent nanogel thermometer for intracellular thermometry.

    Science.gov (United States)

    Gota, Chie; Okabe, Kohki; Funatsu, Takashi; Harada, Yoshie; Uchiyama, Seiichi

    2009-03-01

    The first methodology to measure intracellular temperature is described. A highly hydrophilic fluorescent nanogel thermometer developed for this purpose stays in the cytoplasm and emits stronger fluorescence at a higher temperature. Thus, intracellular temperature variations associated with biological processes can be monitored by this novel thermometer with a temperature resolution of better than 0.5 degrees C.

  12. Equivalent cleaning in a juvenile facultative and obligate cleaning wrasse: an insight into the evolution of cleaning in labrids?

    Science.gov (United States)

    Grutter, Alexandra S.; Feeney, William E.

    2016-09-01

    Species that exhibit ontogenetic variation in interspecific cleaning behaviours may offer insights into how interspecific cooperation evolves. We investigated the foraging ecology of the yellowtail tubelip wrasse ( Diproctacanthus xanthurus), a facultative cleaner as a juvenile and corallivore as an adult, and compared its juvenile ecology with that of juvenile blue-streak cleaner wrasse ( Labroides dimidiatus), a closely related and sympatric obligate cleaner. While juveniles of the two species differed in the amount of time they inspected clients, the number of client individuals and species that were cleaned and the proportion that posed did not differ, nor did the number of ectoparasitic isopods in their guts. In contrast, adult yellowtail tubelip wrasse had fewer isopods and more coral mucus in their guts than juveniles. These data support a hypothesized series of events in which juvenile cleaning acts as an evolutionary precursor to obligate cleaning and suggest that the yellowtail tubelip wrasse may present an intermediate between corallivory and cleaning.

  13. Comparative study of biological hydrogen production by pure strains and consortia of facultative and strict anaerobic bacteria.

    Science.gov (United States)

    Hiligsmann, Serge; Masset, Julien; Hamilton, Christopher; Beckers, Laurent; Thonart, Philippe

    2011-02-01

    In this paper, a simple and rapid method was developed in order to assess in comparative tests the production of binary biogas mixtures containing CO(2) and another gaseous compound such as hydrogen or methane. This method was validated and experimented for the characterisation of the biochemical hydrogen potential of different pure strains and mixed cultures of hydrogen-producing bacteria (HPB) growing on glucose. The experimental results compared the hydrogen production yield of 19 different pure strains and sludges: facultative and strict anaerobic HPB strains along with anaerobic digester sludges thermally pre-treated or not. Significant yields variations were recorded even between different strains of the same species by i.e. about 20% for three Clostridium butyricum strains. The pure Clostridium butyricum and pasteurianum strains achieved the highest yields i.e. up to 1.36 mol H(2)/mol glucose compared to the yields achieved by the sludges and the tested Escherichia and Citrobacter strains.

  14. Molecular evidence of facultative intraguild predation by Monochamus titillator larvae (Coleoptera: Cerambycidae) on members of the southern pine beetle guild

    Science.gov (United States)

    Schoeller, Erich N.; Husseneder, Claudia; Allison, Jeremy D.

    2012-11-01

    The southern pine bark beetle guild (SPBG) is arguably the most destructive group of forest insects in the southeastern USA. This guild contains five species of bark beetles (Coleoptera: Curculionidae: Scolytinae): Dendroctonus frontalis, Dendroctonus terebrans, Ips avulsus, Ips calligraphus, and Ips grandicollis. A diverse community of illicit receivers is attracted to pheromones emitted by the SPBG, including the woodborers Monochamus carolinensis and Monochamus titillator (Coleoptera: Cerambycidae). These woodborers have been traditionally classified as resource competitors; however, laboratory assays suggest that larval M. carolinensis may be facultative intraguild predators of SPBG larvae. This study used polymerase chain reaction (PCR)-based molecular gut content analyses to characterize subcortical interactions between M. titillator and members of the SPBG. The half-lives of SPBG DNA were estimated in the laboratory prior to examining these interactions in the field. A total of 271 field-collected M. titillator larvae were analyzed and 26 (9.6 %) tested positive for DNA of members of the SPBG. Of these larvae, 25 (96.2 %) tested positive for I. grandicollis and one (3.8 %) for I. calligraphus. Failure to detect D. terebrans and D. frontalis was likely due to their absence in the field. I. avulsus was present, but primers developed using adult tissues failed to amplify larval tissue. Results from this study support the hypothesis that larval Monochamus spp. are facultative intraguild predators of bark beetle larvae. Additionally, this study demonstrates the capabilities of PCR in elucidating the interactions of cryptic forest insects and provides a tool to better understand mechanisms driving southern pine beetle guild population fluctuations.

  15. Intracellular structure and nucleocytoplasmic transport.

    Science.gov (United States)

    Agutter, P S

    1995-01-01

    Intracellular movement of any solute or particle accords with one of two general schemes: either it takes place predominantly in the solution phase or it occurs by dynamic interactions with solid-state structures. If nucleocytoplasmic exchanges of macromolecules and complexes are predominantly solution-phase processes, i.e., if the former ("diffusionist") perspective applies, then the only significant structures in nucleocytoplasmic transport are the pore complexes. However, if such exchanges accord with the latter ("solid-state") perspective, then the roles of the nucleoskeleton and cytoskeleton in nucleocytoplasmic transport are potentially, at least, as important as that of the pore complexes. The role of the nucleoskeleton in mRNA transport is more difficult to evaluate than that of the cytoskeleton because it is less well characterized, and current evidence does not exclude either perspective. However, the balance of evidence favors a solid-state scheme. It is argued that ribosomal subunits are also more likely to migrate by a solid-state rather than a diffusionist mechanism, though the opposite is true of proteins and tRNAs. Moreover, recent data on the effects of viral proteins on intranuclear RNA processing and migration accord with the solid-state perspective. In view of this balance of evidence, three possible solid-state mechanisms for nucleocytoplasmic mRNA transport are described and evaluated. The explanatory advantage of solid-state models is contrasted with the heuristic advantage of diffusion theory, but it is argued that diffusion theory itself, even aided by modern computational techniques and numerical and graphical approaches, cannot account for data describing the movements of materials within the cell. Therefore, the mechanisms envisaged in a diffusionist perspective cannot be confined to diffusion alone, but must include other processes such as bulk fluid flow.

  16. A method for functional trans-complementation of intracellular Francisella tularensis.

    Directory of Open Access Journals (Sweden)

    Shaun Steele

    trans-complementation using bead-bound bacteria co-infections is a method to rapidly identify mutants that fail to modify a host response. Francisella tularensis is a facultative intracellular bacterial pathogen and is the causative agent of the disease tularemia. F. tularensis enters host cells through phagocytosis, escapes the phagosome, and replicates in the host cell cytosol while suppressing cytokine secretion [1]-[4]. Although substantial progress has been made in understanding the intracellular life cycle of F. tularensis, the F. tularensis proteins responsible for manipulating many host cell pathways are unknown. Identifying novel host-pathogen effector proteins is difficult because there is no rapid method to reliably distinguish between bacterial proteins that modify host processes and proteins that are involved in bacterial processes that are required for the bacteria to survive or replicate in the intracellular environment. The ability to identify mutants that are deficient for host-pathogen interactions is important because it can aid in prioritizing the investigation of genes of interest and in downstream experimental design. Moreover, certain mutant phenotypes, such as decreased phagosomal escape, hinder investigation of other potential phenotypes. A method to specifically complement these phenotypes would allow for further characterizations of certain F. tularensis mutants. Thus we sought to develop a method to easily identify and functionally complement mutants that are deficient for interactions with the host.

  17. The pore-forming toxin listeriolysin O is degraded by neutrophil metalloproteinase-8 and fails to mediate Listeria monocytogenes intracellular survival in neutrophils.

    Science.gov (United States)

    Arnett, Eusondia; Vadia, Stephen; Nackerman, Colleen C; Oghumu, Steve; Satoskar, Abhay R; McLeish, Kenneth R; Uriarte, Silvia M; Seveau, Stephanie

    2014-01-01

    The pore-forming toxin listeriolysin O (LLO) is a major virulence factor secreted by the facultative intracellular pathogen Listeria monocytogenes. This toxin facilitates L. monocytogenes intracellular survival in macrophages and diverse nonphagocytic cells by disrupting the internalization vesicle, releasing the bacterium into its replicative niche, the cytosol. Neutrophils are innate immune cells that play an important role in the control of infections, yet it was unknown if LLO could confer a survival advantage to L. monocytogenes in neutrophils. We report that LLO can enhance the phagocytic efficiency of human neutrophils and is unable to protect L. monocytogenes from intracellular killing. To explain the absence of L. monocytogenes survival in neutrophils, we hypothesized that neutrophil degranulation leads to the release of LLO-neutralizing molecules in the forming phagosome. In support of this, L. monocytogenes is a potent inducer of neutrophil degranulation, since its virulence factors, such as LLO, facilitate granule exocytosis. Within the first few minutes of interaction with L. monocytogenes, granules can fuse with the plasma membrane at the bacterial interaction site before closure of the phagosome. Furthermore, granule products directly degrade LLO, irreversibly inhibiting its activity. The matrix metalloproteinase-8, stored in secondary granules, was identified as an endoprotease that degrades LLO, and blocking neutrophil proteases increased L. monocytogenes intracellular survival. In conclusion, we propose that LLO degradation by matrix metalloproteinase-8 during phagocytosis protects neutrophil membranes from perforation and contributes to maintaining L. monocytogenes in a bactericidal phagosome from which it cannot escape.

  18. Stochastic resonance in an intracellular genetic perceptron

    Science.gov (United States)

    Bates, Russell; Blyuss, Oleg; Zaikin, Alexey

    2014-03-01

    Intracellular genetic networks are more intelligent than was first assumed due to their ability to learn. One of the manifestations of this intelligence is the ability to learn associations of two stimuli within gene-regulating circuitry: Hebbian-type learning within the cellular life. However, gene expression is an intrinsically noisy process; hence, we investigate the effect of intrinsic and extrinsic noise on this kind of intracellular intelligence. We report a stochastic resonance in an intracellular associative genetic perceptron, a noise-induced phenomenon, which manifests itself in noise-induced increase of response in efficiency after the learning event under the conditions of optimal stochasticity.

  19. 天山冻土中嗜冷酵母菌生物多样性%Diversity of Psychrotrophic Yeast from Permafrost Soil at the Terminus of a Glacier in the Tianshan Mountains

    Institute of Scientific and Technical Information of China (English)

    姜远丽; 郑晓吉; 史学伟; 倪永清

    2012-01-01

    采用麦芽浸膏富集平板涂布天山冻土中可培养酵母菌,通过麦芽浸膏分离培养基筛选菌株.采用酵母菌常规生理生化实验、最适生长温度、最适pH对分离菌株的生理学进行研究.以NL1和NL4为PCR扩增引物,通过26S rRNA基因D1/D2区序列同源性分析,初步确定酵母菌株的系统进化地位.分离筛选到4株嗜低温酵母菌,分属于隐球酵母(Cystofilobasidium infirmominiatum)、掷孢酵母(Sporobolomyces patagonicus)、棒孢酵母(Clavispora lusitaniae)和毕赤酵母(Pichia kluyveri).%The objective of this study was to investigate the diversity of culturable yeast strains from permafrost soils at the terminus of a glacier in the Tianshan Mountains was investigated. Isolation and molecular phylogenetic analysis were performed in order to expand our knowledge on diversity of psychrotrophic and psychrophilic yeast. In addition, efforts focused on screening for psychrophilic yeast. Psychrophilic yeast strains were detected on YEPG plates, respectively. Identity and genetic diversity of strains isolated was determined by 26S rRNA gene D1/D2 by the forward primers NL1 and the reverse NL4. The physiological tests were carried out to determine the phonotypic differences between strains showing high similarity of 26S rRNA gene sequences. In all,4 yeast stains was isolated by 26S rRNA gene D1/D2 and internal transcribed spacers of 4 genera (Cryptococcus ,Sporobolomyces ,Clavispora,Pichia). The results enrich our knowledge on the differences of psychrotrophic yeast diversity between the glacier under study and other worldwide cold habitats are also discussed.

  20. Phenotypic mutants of the intracellular actinomycete Rhodococcus equi created by in vivo Himar1 transposon mutagenesis.

    Science.gov (United States)

    Ashour, Joseph; Hondalus, Mary K

    2003-04-01

    Rhodococcus equi is a facultative intracellular opportunistic pathogen of immunocompromised people and a major cause of pneumonia in young horses. An effective live attenuated vaccine would be extremely useful in the prevention of R. equi disease in horses. Toward that end, we have developed an efficient transposon mutagenesis system that makes use of a Himar1 minitransposon delivered by a conditionally replicating plasmid for construction of R. equi mutants. We show that Himar1 transposition in R. equi is random and needs no apparent consensus sequence beyond the required TA dinucleotide. The diversity of the transposon library was demonstrated by the ease with which we were able to screen for auxotrophs and mutants with pigmentation and capsular phenotypes. One of the pigmentation mutants contained an insertion in a gene encoding phytoene desaturase, an enzyme of carotenoid biosynthesis, the pathway necessary for production of the characteristic salmon color of R. equi. We identified an auxotrophic mutant with a transposon insertion in the gene encoding a putative dual-functioning GTP cyclohydrolase II-3,4-dihydroxy-2-butanone-4-phosphate synthase, an enzyme essential for riboflavin biosynthesis. This mutant cannot grow in minimal medium in the absence of riboflavin supplementation. Experimental murine infection studies showed that, in contrast to wild-type R. equi, the riboflavin-requiring mutant is attenuated because it is unable to replicate in vivo. The mutagenesis methodology we have developed will allow the characterization of R. equi virulence mechanisms and the creation of other attenuated strains with vaccine potential.

  1. Intracellular Biopotentials During Static Extracellular Stimulation

    Science.gov (United States)

    Klee, Maurice

    1973-01-01

    Two properties of the intracellular potentials and electric fields resulting from static extracellular stimulation are obtained for arbitrarily shaped cells. First, the values of intracellular potential are shown to be bounded by the maximum and minimum values of extracellular potential on the surface of the cell. Second, the volume average of the magnitude of intracellular electric field is shown to have an upper bound given by the ratio of the magnitude of the largest extracellular potential difference on the surface of the cell to a generalized length constant λ = [σintraVcell/(σmemb Acell)]1/2, where Vcell and Acell are the volume and surface area of the cell, σintra is the intracellular conductivity (reciprocal ohms per centimeter), and σmemb is the membrane conductivity (reciprocal ohms per square centimeter). The use of the upper bound on the volume average of the magnitude of intracellular electric field as an estimate for intracellular isopotentiality is discussed and the use of the generalized length constant for electrically describing arbitrary cells is illustrated for cylindrical- and spheroidal-shaped cells. PMID:4726882

  2. Diverse gene cassettes in class 1 integrons of facultative oligotrophic bacteria of River Mahananda,West Bengal, India.

    Directory of Open Access Journals (Sweden)

    Ranadhir Chakraborty

    Full Text Available BACKGROUND: In this study a large random collection (n=2188 of facultative oligotrophic bacteria, from 90 water samples gathered in three consecutive years (2007-2009 from three different sampling sites of River Mahananda in Siliguri, West Bengal, India, were investigated for the presence of class 1 integrons and sequences of the amplification products. METHODOLOGY/PRINCIPAL FINDINGS: Replica plating method was employed for determining the antibiotic resistance profile of the randomly assorted facultative oligotrophic isolates. Genomic DNA from each isolate was analyzed by PCR for the presence of class 1 integron. Amplicons were cloned and sequenced. Numerical taxonomy and 16S rRNA gene sequence analyses were done to ascertain putative genera of the class 1 integron bearing isolates. Out of 2188 isolates, 1667 (76.19% were antibiotic-resistant comprising of both single-antibiotic resistance (SAR and multiple-antibiotic resistant (MAR, and 521 (23.81% were sensitive to all twelve different antibiotics used in this study. Ninety out of 2188 isolates produced amplicon(s of varying sizes from 0.15 to 3.45 KB. Chi-square (χ(2 test revealed that the possession of class 1 integron in sensitive, SAR and MAR is not equally probable at the 1% level of significance. Diverse antibiotic-resistance gene cassettes, aadA1, aadA2, aadA4, aadA5, dfrA1, dfrA5, dfrA7, dfrA12, dfrA16, dfrA17, dfrA28, dfrA30, dfr-IIe, blaIMP-9, aacA4, Ac-6'-Ib, oxa1, oxa10 and arr2 were detected in 64 isolates. The novel cassettes encoding proteins unrelated to any known antibiotic resistance gene function were identified in 26 isolates. Antibiotic-sensitive isolates have a greater propensity to carry gene cassettes unrelated to known antibiotic-resistance genes. The integron-positive isolates under the class Betaproteobacteria comprised of only two genera, Comamonas and Acidovorax of family Comamonadaceae, while isolates under class Gammaproteobacteria fell under the families

  3. Experimental evolution of rhizobia may lead to either extra- or intracellular symbiotic adaptation depending on the selection regime.

    Science.gov (United States)

    Marchetti, Marta; Clerissi, Camille; Yousfi, Yasmine; Gris, Carine; Bouchez, Olivier; Rocha, Eduardo; Cruveiller, Stéphane; Jauneau, Alain; Capela, Delphine; Masson-Boivin, Catherine

    2017-04-01

    Experimental evolution is a powerful approach to study the process of adaptation to new environments, including the colonization of eukaryotic hosts. Facultative endosymbionts, including pathogens and mutualists, face changing and spatially structured environments during the symbiotic process, which impose diverse selection pressures. Here, we provide evidence that different selection regimes, involving different times spent in the plant environment, can result in either intra- or extracellular symbiotic adaptations. In previous work, we introduced the symbiotic plasmid of Cupriavidus taiwanensis, the rhizobial symbiont of Mimosa pudica, into the phytopathogen Ralstonia solanacearum and selected three variants able to form root nodules on M. pudica, two (CBM212 and CBM349) being able to rudimentarily infect nodule cells and the third one (CBM356) only capable of extracellular infection of nodules. Each nodulating ancestor was further challenged to evolve using serial ex planta-in planta cycles of either 21 (three short-cycle lineages) or 42 days (three long-cycle lineages). In this study, we compared the phenotype of the 18 final evolved clones. Evolution through short and long cycles resulted in similar adaptive paths on lineages deriving from the two intracellularly infectious ancestors, CBM212 and CBM349. In contrast, only short cycles allowed a stable acquisition of intracellular infection in lineages deriving from the extracellularly infecting ancestor, CBM356. Long cycles, instead, favoured improvement of extracellular infection. Our work highlights the importance of the selection regime in shaping desired traits during host-mediated selection experiments. © 2016 John Wiley & Sons Ltd.

  4. Life History Consequences of the Facultative Expression of a Dispersal Life Stage in the Phoretic Bulb Mite (Rhizoglyphus robini.

    Directory of Open Access Journals (Sweden)

    Jacques A Deere

    Full Text Available Life history traits play an important role in population dynamics and correlate, both positively and negatively, with dispersal in a wide range of taxa. Most invertebrate studies on trade-offs between life history traits and dispersal have focused on dispersal via flight, yet much less is known about how life history trade-offs influence species that disperse by other means. In this study, we identify effects of investing in dispersal morphology (dispersal expression on life history traits in the male dimorphic bulb mite (Rhizoglyphus robini. This species has a facultative juvenile life stage (deutonymph during which individuals can disperse by phoresy. Further, adult males are either fighters (which kill other mites or benign scramblers. Here, in an experiment, we investigate the effects of investing in dispersal on size at maturity, sex and male morph ratio, and female lifetime reproductive success. We show that life history traits correlate negatively with the expression of the dispersal stage. Remarkably, all males that expressed the dispersal life stage developed into competitive fighters and none into scramblers. This suggests that alternative, male reproductive strategies and dispersal should not be viewed in isolation but considered concurrently.

  5. Gill morphometry of the facultative air-breathing loricariid fish,Hypostomus plecostomus (Walbaum) with, special emphasis on aquatic respiration.

    Science.gov (United States)

    Aparecida Perna, S; Fernandes, M N

    1996-06-01

    Gill respiratory surface area and oxygen consumption during aquatic respiration were measured in the facultative air-breathing loricariid fish,Hypostomus plecostomus. The fish did not surface to breathe atmospheric air in normoxic water; air-breathing was evoked by environmental hypoxia (water oxygen tension=35±2, mmHg) and did not show size-related threshold differences for air breathing.During gradual hypoxia, without access to atmospheric, air,H. plecostomus was found to be an oxyregulator and showed a reduced range of water oxygen tension in which the oxygen consumption remained constant in smaller fish. The critical oxygen tensions were 55 and 33 mmHg at 25°C for fish of 14-30 g and 31-80g body weight, respectively.The gill respiratory surface area (total lamellae area) is reduced, however, the lamellar frequency per mm of gill filament is high which facilitates the gas exchange. Moreover, the increase of gill respiratory surface area (b=0.666) is higher than the increase in routine VO2 (b=0.338) showing a positive relationship between the gill respiratory surface area /VO2 ratio and body mass (b=0.328); this indicates that the fish have greater gill respiratory surface area per unit of routine VO2 as they grow.

  6. Carnobacterium Pleistocaenium sp. nov.: A Novel Psychrotolerant, Facultative Anaerobe Isolated from Permafrost of the Fox Tunnel in Alaska

    Science.gov (United States)

    Pikuta, Elena V.; Marsic, Damien; Bej, Asim; Tang, Jane; Krader, Paul; Hoover, Richard B.

    2004-01-01

    A novel, psychrotolerant, facultative anaerobe, strain FTRIT1(sup T), was isolated from Pleistocene ice from the permafrost tunnel in Fox, Alaska. Gram-positive, motile, rod-shaped cells with sizes 0.6-0.7 x 0.9-1.5 micrometers were observed. Growth occurred within the pH range 6.5-9.5 and optimum at pH 7.3-7.5. The temperature range of the new isolate was 0-28 C and optimum growth occurred at 24 C. The novel isolate requires NaCl (growth absent at 0 %) and growth was observed between 0 and 5% NaCl with optimum at 0.5% (w/v). The new isolate was a catalase-negative chemoorganoheterotroph that used as substrates sugars and some products of proteolysis. The metabolic end products were: acetate, ethanol and CO2. Strain FTRl was sensitive to ampicillin, tetracycline, chloramphenicol, rifampin, kanamycin, and gentamycin. The 16S rDNA sequence analysis showed 99.8% similarity of strain FTR1 with Carnobacterium alterfunditum, but the DNA-DNA hybridization between them demonstrated 39 plus or minus 5% homology. On the basis of genotypic and phenotypic characteristics, it is proposed that the strain FTR1(sup T) (= ATCC BAA-754(sup T) = JSM 12174(sup T) is assigned to the new species of the genus Carnobacterium with proposed name Carnobacterium pleistocaenium sp. nov.

  7. Carnobacterium pleistocenium sp. nov., a novel psychrotolerant, facultative anaerobe isolated from permafrost of the Fox Tunnel in Alaska

    Science.gov (United States)

    Pilkuta, Elena V.; Marsic, Damien; Bej, Asim; Tang, Jane; Krader, Paul; Hoover, Richard B.

    2005-01-01

    A novel, psychrotolerant, facultative anaerobe, strain FTRl, was isolated from Pleistocene ice from the permafrost tunnel in Fox, Alaska. Gram-positive, motile, rod-shaped cells were observed with sizes 0(raised dot)6-0(raised dot)7 x 0(raised dot)9-1(raised dot)5 microns. Growth occurred within the pH range 6(raised dot)5-9(raised dot)5 with optimum growth at pH 7(raised dot)3-7(raised dot)5. The temperature range for growth of the novel isolate was 0-28 C and optimum growth occurred at 24 C. The novel isolate does not require NaCl; growth was observed between 0 and 5% NaCl with optimum growth at 0(raised dot)5% (w/v). The novel isolate was a catalase-negative chemoorganoheterotroph that used as substrates sugars and some products of proteolysis. The metabolic end products were acetate, ethanol and CO2. Strain FTRl was sensitive to ampicillin, tetracycline, chloramphenicol, rifampicin, kanamycin and gentamicin. 16s rRNA gene sequence analysis showed 99(raised dot)8% similarity between strain FTR1 and Carnobacterium alterfunditum, but DNA-DNA hybridization between them demonstrated 39 plus or minus 1(raised dot)5% relatedness. On the basis of genotypic and phenotypic characteristics, it is proposed that strain FTRl (= ATCC BAA-754T= JCM 12174T=CIP 108033) be assigned to the novel species Carnobacterium pleistocenium sp. nov.

  8. Anaerobic oxidation of arsenite in Mono Lake water and by a facultative, arsenite-oxidizing chemoautotroph, strain MLHE-1

    Science.gov (United States)

    Oremland, R.S.; Hoeft, S.E.; Santini, J.M.; Bano, N.; Hollibaugh, R.A.; Hollibaugh, J.T.

    2002-01-01

    Arsenite [As(III)]-enriched anoxic bottom water from Mono Lake, California, produced arsenate [As(V)] during incubation with either nitrate or nitrite. No such oxidation occurred in killed controls or in live samples incubated without added nitrate or nitrite. A small amount of biological As(III) oxidation was observed in samples amended with Fe(III) chelated with nitrolotriacetic acid, although some chemical oxidation was also evident in killed controls. A pure culture, strain MLHE-1, that was capable of growth with As(III) as its electron donor and nitrate as its electron acceptor was isolated in a defined mineral salts medium. Cells were also able to grow in nitrate-mineral salts medium by using H2 or sulfide as their electron donor in lieu of As(III). Arsenite-grown cells demonstrated dark 14CO2 fixation, and PCR was used to indicate the presence of a gene encoding ribulose-1,5-biphosphate carboxylase/oxygenase. Strain MLHE-1 is a facultative chemoautotroph, able to grow with these inorganic electron donors and nitrate as its electron acceptor, but heterotrophic growth on acetate was also observed under both aerobic and anaerobic (nitrate) conditions. Phylogenetic analysis of its 16S ribosomal DNA sequence placed strain MLHE-1 within the haloalkaliphilic Ectothiorhodospira of the ??-Proteobacteria. Arsenite oxidation has never been reported for any members of this subgroup of the Proteobacteria.

  9. Testing strength of biotic resistance against an introduced fish: inter-specific competition or predation through facultative piscivory?

    Science.gov (United States)

    Britton, J Robert

    2012-01-01

    Biotic resistance is the process where aspects of the receiving environment inhibit the establishment and invasion of an introduced species. Resistance against an introduced fish can be through strong competition and/or predation from resident fishes. Here, the biotic resistance against introduced topmouth gudgeon Pseudorasbora parva (a highly invasive fish in Europe) by resident carp Cyprinus carpio was tested in experimental mesocosms. The introduction scenario was six adult P. parva (three male, three female) on a single occasion. Resistance to their establishment was provided by three and six resident C. carpio whose effects on P. parva growth and reproduction were compared to a Control (no resident fish at the time of introduction) and treatments containing three and six P. parva. After 120 days, the growth rates of the introduced P. parva were significantly depressed in C. carpio presence and in mesocosms with three C. carpio present, significantly decreased numbers of 0+P. parva were recorded. Where six C. carpio were present, no 0+P. parva were recorded, indicating resistance strength increased with carp abundance. In contrast, there were no differences in P. parva reproduction and growth rates between the Control and treatments containing conspecifics. Stable isotope analysis (δ(15)N, δ(13)C) revealed C. carpio were feeding at one trophic level above 0+P. parva, suggesting the process of resistance was predation (facultative piscivory) rather than competition. Thus, if P. parva are to establish and invade following an introduction, they must overcome this biotic resistance from cyprinid fishes such as C. carpio.

  10. In vitro sensitivity of oral, gram-negative, facultative bacteria to the bactericidal activity of human neutrophil defensins.

    Science.gov (United States)

    Miyasaki, K T; Bodeau, A L; Ganz, T; Selsted, M E; Lehrer, R I

    1990-12-01

    Neutrophils play a major role in defending the periodontium against infection by oral, gram-negative, facultative bacteria, such as Actinobacillus actinomycetemcomitans, Eikenella corrodens, and Capnocytophaga spp. We examined the sensitivity of these bacteria to a mixture of low-molecular-weight peptides and highly purified individual defensin peptides (HNP-1, HNP-2, and HNP-3) isolated from human neutrophils. Whereas the Capnocytophaga spp. strains were killed significantly by the mixed human neutrophil peptides, the A. actinomycetemcomitans and E. corrodens strains were resistant. Killing was attributable to the defensins. The bactericidal activities of purified defensins HNP-1 and HNP-2 were equal, and both of these activities were greater than HNP-3 activity against strains of Capnocytophaga sputigena and Capnocytophaga gingivalis. The strain of Capnocytophaga ochracea was more sensitive to defensin-mediated bactericidal activity than either C. sputigena or C. gingivalis was. The three human defensins were equipotent in killing C. ochracea. C. ochracea was killed under aerobic and anaerobic conditions and over a broad pH range. Killing was most effective under hypotonic conditions but also occurred at physiologic salt concentrations. We concluded that Capnocytophaga spp. are sensitive to oxygen-independent killing by human defensins. Additional studies will be required to identify other components that may equip human neutrophils to kill A. actinomycetemcomitans, E. corrodens, and other oral gram-negative bacteria.

  11. Organization and regulation of the arsenite oxidase operon of the moderately acidophilic and facultative chemoautotrophic Thiomonas arsenitoxydans.

    Science.gov (United States)

    Slyemi, Djamila; Moinier, Danielle; Talla, Emmanuel; Bonnefoy, Violaine

    2013-11-01

    Thiomonas arsenitoxydans is an acidophilic and facultatively autotrophic bacterium that can grow by oxidizing arsenite to arsenate. A comparative genomic analysis showed that the T. arsenitoxydans aioBA cluster encoding the two subunits of arsenite oxidase is distinct from the other clusters, with two specific genes encoding a cytochrome c and a metalloregulator belonging to the ArsR/SmtB family. These genes are cotranscribed with aioBA, suggesting that these cytochromes c are involved in arsenite oxidation and that this operon is controlled by the metalloregulator. The growth of T. arsenitoxydans in the presence of thiosulfate and arsenite, or arsenate, is biphasic. Real-time PCR experiments showed that the operon is transcribed during the second growth phase in the presence of arsenite or arsenate, whereas antimonite had no effect. These results suggest that the expression of the aioBA operon of T. arsenitoxydans is regulated by the electron donor present in the medium, i.e., is induced in the presence of arsenic but is repressed by more energetic substrates. Our data indicate that the genetic organization and regulation of the aioBA operon of T. arsenitoxydans differ from those of the other arsenite oxidizers.

  12. Complete genome sequence of the facultatively chemolithoautotrophic and methylotrophic alpha Proteobacterium Starkeya novella type strain (ATCC 8093T)

    Energy Technology Data Exchange (ETDEWEB)

    Kappler, Ulrike [University of Queensland, The, Brisbane, Queensland, Australia; Davenport, Karen W. [Los Alamos National Laboratory (LANL); Beatson, Scott [University of Queensland, The, Brisbane, Queensland, Australia; Lucas, Susan [U.S. Department of Energy, Joint Genome Institute; Lapidus, Alla L. [U.S. Department of Energy, Joint Genome Institute; Copeland, A [U.S. Department of Energy, Joint Genome Institute; Berry, Kerrie W. [U.S. Department of Energy, Joint Genome Institute; Glavina Del Rio, Tijana [U.S. Department of Energy, Joint Genome Institute; Hammon, Nancy [U.S. Department of Energy, Joint Genome Institute; Dalin, Eileen [U.S. Department of Energy, Joint Genome Institute; Tice, Hope [U.S. Department of Energy, Joint Genome Institute; Pitluck, Sam [U.S. Department of Energy, Joint Genome Institute; Richardson, P M [U.S. Department of Energy, Joint Genome Institute; Bruce, David [Los Alamos National Laboratory (LANL); Goodwin, Lynne A. [Los Alamos National Laboratory (LANL); Han, Cliff [Los Alamos National Laboratory (LANL); Tapia, Roxanne [Los Alamos National Laboratory (LANL); Detter, J. Chris [U.S. Department of Energy, Joint Genome Institute; Chang, Yun-Juan [ORNL; Jeffries, Cynthia [Oak Ridge National Laboratory (ORNL); Land, Miriam L [ORNL; Hauser, Loren John [ORNL; Kyrpides, Nikos C [U.S. Department of Energy, Joint Genome Institute; Goker, Markus [DSMZ - German Collection of Microorganisms and Cell Cultures GmbH, Braunschweig, Germany; Ivanova, N [U.S. Department of Energy, Joint Genome Institute; Klenk, Hans-Peter [DSMZ - German Collection of Microorganisms and Cell Cultures GmbH, Braunschweig, Germany; Woyke, Tanja [U.S. Department of Energy, Joint Genome Institute

    2012-01-01

    Starkeya novella (Starkey 1934) Kelly et al. 2000 is a member of the family Xanthobacteraceae in the order Rhizobiales , which is thus far poorly characterized at the genome level. Cultures from this spe- cies are most interesting due to their facultatively chemolithoautotrophic lifestyle, which allows them to both consume carbon dioxide and to produce it. This feature makes S. novella an interesting model or- ganism for studying the genomic basis of regulatory networks required for the switch between con- sumption and production of carbon dioxide, a key component of the global carbon cycle. In addition, S. novella is of interest for its ability to grow on various inorganic sulfur compounds and several C1- compounds such as methanol. Besides Azorhizobium caulinodans, S. novella is only the second spe- cies in the family Xanthobacteraceae with a completely sequenced genome of a type strain. The cur- rent taxonomic classification of this group is in significant conflict with the 16S rRNA data. The ge- nomic data indicate that the physiological capabilities of the organism might have been underestimat- ed. The 4,765,023 bp long chromosome with its 4,511 protein-coding and 52 RNA genes was se- quenced as part of the DOE Joint Genome Institute Community Sequencing Program (CSP) 2008.

  13. Anaerobic biodegradation of high-molecular-weight polycyclic aromatic hydrocarbons by a facultative anaerobe Pseudomonas sp. JP1.

    Science.gov (United States)

    Liang, Lei; Song, Xiaohui; Kong, Jing; Shen, Chenghui; Huang, Tongwang; Hu, Zhong

    2014-11-01

    Polycyclic aromatic hydrocarbons (PAHs) are harmful persistent organic pollutants, while the high-molecular-weight (HMW) PAHs are even more detrimental to the environment and human health. However, microbial anaerobic degradation of HMW PAHs has rarely been reported. One facultative anaerobe Pseudomonas sp. JP1 was isolated from Shantou Bay, Shantou, China, which could degrade a variety of HMW PAHs. After 40 days cultivation with strain JP1, anaerobic biodegradation rate of benzo[a]pyrene (BaP), fluoranthene, and phenanthrene was 30, 47, and 5 %, respectively. Consumption of nitrate as the electron acceptor was confirmed by N-(1-naphthyl) ethylenediamine spectrophotometry. Supplementation of sodium sulfite, maltose, or glycine, and in a salinity of 0-20 ‰ significantly stimulated anaerobic degradation of BaP. Lastly, the anaerobic degradation metabolites of BaP by strain JP1 were investigated using GC/MS, and the degradation pathway was proposed. This study is helpful for further studies on the mechanism of anaerobic biodegradation of PAHs.

  14. Biological decomposition of Na2S2O3 into sulfur by a newly isolated facultative thermophilic alkaline desulphuricant strain

    Institute of Scientific and Technical Information of China (English)

    ZHANG JianBin; ZHANG Tong; MA Kai; ZHANG PengYan; LI Qiang; WEI XiongHui

    2009-01-01

    A facultative thermophilic alkaline desulphuricant strain named GDJ-3 was isolated from neutral soils, enriched on sulfur synthetic medium, and detected in previous work. Conventional and chemotax-onomic analyses and 16s RNA gone sequencing showed that the strain was in good agreement with Alpha proteobacterium sp. (97%) and ochrobactrum sp. (98%). The regenerative processes of the solu-tion containing sulfur compounds (SCS) from the strain through an orthogonal test were investigated to get the optimum regenerative condition. The results showed that regenerative temperature, air flow, and stirring speed of the agitator were the main three variables influencing the regenerative processes of the SCS. The optimum regenerative efficiency of the SCS from the strain was obtained when tem-perature, air flow, and stirring speed of the agitator were 318.2 K, 3.0 L/min, and zero r/min, respectively. Under this condition, when the cell concentration of the strain was adjusted to 107/mL, the concentra-tions of Na2S2O3 and Na2S in the SCS decreased from 112.68 g/L to 96.88 g/L and from 0. 87 g/L to 0.11 g/L in 9.5 h. Meanwhile, XRD spectrum shows that sulfur was formed in the regeneration process. These results suggest that the strain has potential application to the regeneration of the industrial so-lution containing sulfur compounds.

  15. Pandoraea thiooxydans sp. nov., a facultatively chemolithotrophic, thiosulfate-oxidizing bacterium isolated from rhizosphere soils of sesame (Sesamum indicum L.).

    Science.gov (United States)

    Anandham, Rangasamy; Indiragandhi, Pandiyan; Kwon, Soon Wo; Sa, Tong Min; Jeon, Che Ok; Kim, Yong Ki; Jee, Hyeong Jin

    2010-01-01

    A facultatively chemolithoautotrophic, thiosulfate-oxidizing, Gram-negative, aerobic, motile, rod-shaped bacterial strain, designated ATSB16(T), was isolated from rhizosphere soils of sesame (Sesamum indicum L.). 16S rRNA gene sequence analysis demonstrated that this strain was closely related to Pandoraea pnomenusa LMG 18087(T) (96.7 % similarity), P. pulmonicola LMG 18016(T) (96.5 %), P. apista LMG 16407(T) (96.2 %), P. norimbergensis LMG 18379(T) (96.1 %) and P. sputorum LMG 18819(T) (96.0 %). Strain ATSB16(T) shared 96.0-96.4 % sequence similarity with four unnamed genomospecies of Pandoraea. The major cellular fatty acids of the strain ATSB16(T) were C(17 : 0) cyclo (33.0 %) and C(16 : 0) (30.6 %). Q-8 was the predominant respiratory quinone. The major polar lipids were phosphatidylmethylethanolamine, diphosphatidylglycerol, phosphatidylethanolamine and two unidentified aminophospholipids. Hydroxyputrescine and putrescine were the predominant polyamines. The genomic DNA G+C content of the strain was 64.0 mol%. On the basis of the results obtained from this study, strain ATSB16(T) represents a novel species of the genus Pandoraea, for which the name Pandoraea thiooxydans sp. nov. is proposed. The type strain is ATSB16(T) (=KACC 12757(T) =LMG 24779(T)).

  16. Mitochondrial genome from the facultative anaerobe and petite-positive yeast Dekkera bruxellensis contains the NADH dehydrogenase subunit genes.

    Science.gov (United States)

    Procházka, Emanuel; Poláková, Silvia; Piskur, Jure; Sulo, Pavol

    2010-08-01

    The progenitor of the Dekkera/Brettanomyces clade separated from the Saccharomyces/Kluyveromyces clade over 200 million years ago. However, within both clades, several lineages developed similar physiological traits. Both Saccharomyces cerevisiae and Dekkera bruxellensis are facultative anaerobes; in the presence of excess oxygen and sugars, they accumulate ethanol (Crabtree effect) and they both spontaneously generate respiratory-deficient mutants (petites). In order to understand the role of respiratory metabolism, the mitochondrial DNA (mtDNA) molecules of two Dekkera/Brettanomyces species were analysed. Dekkera bruxellensis mtDNA shares several properties with S. cerevisiae, such as the large genome size (76 453 bp), and the organization of the intergenic sequences consisting of spacious AT-rich regions containing a number of hairpin GC-rich cluster-like elements. In addition to a basic set of the mitochondrial genes coding for the components of cytochrome oxidase, cytochrome b, subunits of ATPase, two rRNA subunits and 25 tRNAs, D. bruxellensis also carries genes for the NADH dehydrogenase complex. Apparently, in yeast, the loss of this complex is not a precondition to develop a petite-positive, Crabtree-positive and anaerobic nature. On the other hand, mtDNA from a petite-negative Brettanomyces custersianus is much smaller (30 058 bp); it contains a similar gene set and has only short intergenic sequences.

  17. Exploring species level taxonomy and species delimitation methods in the facultatively self-fertilizing land snail genus Rumina (gastropoda: pulmonata.

    Directory of Open Access Journals (Sweden)

    Vanya Prévot

    Full Text Available Delimiting species in facultatively selfing taxa is a challenging problem of which the terrestrial pulmonate snail genus Rumina is a good example. These snails have a mixed breeding system and show a high degree of shell and color variation. Three nominal species (R. decollata, R. saharica and R. paivae and two color morphs within R. decollata (dark and light are currently recognized. The present study aims at evaluating to what extent these entities reflect evolutionary diverging taxonomic units, rather than fixed polymorphisms due to sustained selfing. Therefore, a phylogenetic analysis of nuclear (ITS1, ITS2 and mitochondrial DNA (COI, CytB, 12S rDNA, 16S rDNA sequences was performed. Putative species in Rumina, inferred from the mitochondrial DNA phylogeny, were compared with those proposed on the basis of the COI gene by (1 DNA barcoding gap analysis, (2 Automatic Barcode Gap Discovery, (3 the species delimitation plug-in of the Geneious software, (4 the Genealogical Sorting Index, and (5 the General Mixed Yule Coalescent model. It is shown that these methods produce a variety of different species hypotheses and as such one may wonder to what extent species delimitation methods are really useful. With respect to Rumina, the data suggest at least seven species, one corresponding to R. saharica and six that are currently grouped under the name R. decollata. The species-level status of R. paivae is rejected.

  18. Isolation of a facultative anaerobic exoelectrogenic strain LZ-1 and probing electron transfer mechanism in situ by linking UV/Vis spectroscopy and electrochemistry.

    Science.gov (United States)

    Zhou, Lei; Deng, Dandan; Zhang, Yichi; Zhou, Wen; Jiang, Yujing; Liu, Ying

    2017-04-15

    A new facultative anaerobic exoelectrogenic strain LZ-1, belonging to Citrobacter freundii, has been isolated. This strain can produce current densities of 843.9 and 865.6μAcm(-2) using citrate or acetate as carbon source in a three-electrode configuration. The electricity generation performance was also analyzed in a dual-chamber MFC system, reaching a maximum power density of 1233mWm(-2). In addition to acetate and citrate, other carbon sources such as pyruvate, formate, acetate, citrate and fumarate could also be utilized to produce current by strain LZ-1. Data supports the presence of electroactive c-type cytochromes in C. freundii sp. when grown on ITO electrodes, by linking spectroscopy and electrochemistry in situ. Since facultative strains possess many desirable properties compared to anaerobic strains, strain LZ-1 represents a promising exoelectrogenic species in engineering of biological catalysts for microbial electrochemistry. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Identification of a conserved protein involved in anaerobic unsaturated fatty acid synthesis in Neiserria gonorrhoeae: implications for facultative and obligate anaerobes that lack FabA

    OpenAIRE

    Isabella, Vincent M.; Clark, Virginia L.

    2011-01-01

    Transcriptome analysis of the facultative anaerobe, Neisseria gonorrhoeae, revealed that many genes of unknown function were induced under anaerobic conditions. Mutation of one such gene, NGO1024, encoding a protein belonging to the 2-nitropropane dioxygenase-like superfamiliy of proteins, was found to result in an inability of gonococci to grow anaerobically. Anaerobic growth of an NG1024 mutant was restored upon supplementation with unsaturated fatty acids (UFA), but not with the saturated ...

  20. Internal affairs: investigating the Brucella intracellular lifestyle.

    Science.gov (United States)

    von Bargen, Kristine; Gorvel, Jean-Pierre; Salcedo, Suzana P

    2012-05-01

    Bacteria of the genus Brucella are Gram-negative pathogens of several animal species that cause a zoonotic disease in humans known as brucellosis or Malta fever. Within their hosts, brucellae reside within different cell types where they establish a replicative niche and remain protected from the immune response. The aim of this article is to discuss recent advances in the field in the specific context of the Brucella intracellular 'lifestyle'. We initially discuss the different host cell targets and their relevance during infection. As it represents the key to intracellular replication, the focus is then set on the maturation of the Brucella phagosome, with particular emphasis on the Brucella factors that are directly implicated in intracellular trafficking and modulation of host cell signalling pathways. Recent data on the role of the type IV secretion system are discussed, novel effector molecules identified and how some of them impact on trafficking events. Current knowledge on Brucella gene regulation and control of host cell death are summarized, as they directly affect intracellular persistence. Understanding how Brucella molecules interplay with their host cell targets to modulate cellular functions and establish the intracellular niche will help unravel how this pathogen causes disease.

  1. Efficient intracellular delivery and improved biocompatibility of colloidal silver nanoparticles towards intracellular SERS immuno-sensing.

    Science.gov (United States)

    Bhardwaj, Vinay; Srinivasan, Supriya; McGoron, Anthony J

    2015-06-21

    High throughput intracellular delivery strategies, electroporation, passive and TATHA2 facilitated diffusion of colloidal silver nanoparticles (AgNPs) are investigated for cellular toxicity and uptake using state-of-art analytical techniques. The TATHA2 facilitated approach efficiently delivered high payload with no toxicity, pre-requisites for intracellular applications of plasmonic metal nanoparticles (PMNPs) in sensing and therapeutics.

  2. GTPases in intracellular trafficking: an overview.

    Science.gov (United States)

    Segev, Nava

    2011-02-01

    Small GTPases that belong to the ras sub-families of Rab, Arf, and Rho, and the large GTPase dynamin, regulate intracellular trafficking. This issue of Seminars of Cell and Developmental Biology highlights topics regarding mechanisms by which these GTPases regulate the different steps of vesicular transport: vesicle formation, scission, targeting and fusion. In addition, the emerging roles of GTPases in coordination of individual transport steps as well as coordination of intracellular trafficking with other cellular processes are reviewed. Finally, common structures and mechanisms underlying the function of the ras-like GTPases and the importance of their function to human health and disease are discussed.

  3. Antimicrobial Activity of Endodontic Medicaments and Vehicles using Agar Well Diffusion Method on Facultative and Obligate Anaerobes

    Science.gov (United States)

    Bhat, Kishore G; Sogi, Suma

    2016-01-01

    , and lesser chances of developing resistance. How to cite this article Nalawade TM, Bhat KG, Sogi S. Antimicrobial Activity of Endodontic Medicaments and Vehicles using Agar Well Diffusion Method on Facultative and Obligate Anaerobes. Int J Clin Pediatr Dent 2016;9(4):335-341. PMID:28127166

  4. Trichococcus Patagoniensis sp. nov., a Facultative Anaerobe that grows at -5 C, Isolated from Penguin Guano in Chilean Patagonia

    Science.gov (United States)

    Pikuta, Elena V.; Hoover, Richard B.; Bej, Asim K.; Marsic, Damien; Whitman, William B.; Krader, Paul E.; Tang, Jane

    2006-01-01

    A novel, extremely psychrotolerant, facultative anaerobe, strain PmagGl(sup T), was isolated from guano of Magellanic penguins (Spheniscus magellanicus) collected in Chilean Patagonia. Gram-variable, motile cocci with a diameter of 1.3-2.0 micrometers were observed singularly or in pairs, short chains and irregular conglomerates. Growth occurred within the pH range 6.0-10.0, with optimum growth at pH 8.5. The temperature range for growth of the novel isolate was from -5 to 35 C, with optimum growth at 28-30 C. Strain PmagG1(sup T) did not require NaCl, as growth was observed in the presence of 0-6.5% NaCl with optimum growth at 0.5% (w/v). Strain PmagGl(sup T) was a catalase-negative chemo-organoheterotroph that used sugars and some organic acids as substrates. The metabolic end products were lactate, formate, acetate, ethanol and Con. Strain PmagG1(sup T) was sensitive to ampicillin, tetracycline, chloramphenicol, rifampicin, kanamycin and gentamicin. The G+C content of its genomic DNA was 45.8 mol%. 16S rRNA gene sequence analysis showed 100 % similarity of strain PmagG1(sup T) with Trichococcus collinsii ATCC BAA-296(sup T), but DNA-DNA hybridization between them demonstrated relatedness values of less than 45 plus or minus 1%. Another phylogenetically closely related species, Trichococcus pasteurii, showed 99.85 % similarity by 16s rRNA sequencing and DNA-DNA hybridization showed relatedness values of 47 plus or minus 1.5%. Based on genotypic and phenotypic characteristics, the novel species Trichococcus patagoniensis sp. nov. is proposed, with strain PmagG1(sup T) (=ATCC BAA-756(sup T)=JCM 12176(sup T)=CIP 108035(sup T)) as the type strain.

  5. Testing strength of biotic resistance against an introduced fish: inter-specific competition or predation through facultative piscivory?

    Directory of Open Access Journals (Sweden)

    J Robert Britton

    Full Text Available Biotic resistance is the process where aspects of the receiving environment inhibit the establishment and invasion of an introduced species. Resistance against an introduced fish can be through strong competition and/or predation from resident fishes. Here, the biotic resistance against introduced topmouth gudgeon Pseudorasbora parva (a highly invasive fish in Europe by resident carp Cyprinus carpio was tested in experimental mesocosms. The introduction scenario was six adult P. parva (three male, three female on a single occasion. Resistance to their establishment was provided by three and six resident C. carpio whose effects on P. parva growth and reproduction were compared to a Control (no resident fish at the time of introduction and treatments containing three and six P. parva. After 120 days, the growth rates of the introduced P. parva were significantly depressed in C. carpio presence and in mesocosms with three C. carpio present, significantly decreased numbers of 0+P. parva were recorded. Where six C. carpio were present, no 0+P. parva were recorded, indicating resistance strength increased with carp abundance. In contrast, there were no differences in P. parva reproduction and growth rates between the Control and treatments containing conspecifics. Stable isotope analysis (δ(15N, δ(13C revealed C. carpio were feeding at one trophic level above 0+P. parva, suggesting the process of resistance was predation (facultative piscivory rather than competition. Thus, if P. parva are to establish and invade following an introduction, they must overcome this biotic resistance from cyprinid fishes such as C. carpio.

  6. No apparent costs for facultative antibiotic production by the soil bacterium Pseudomonas fluorescens Pf0-1.

    Directory of Open Access Journals (Sweden)

    Paolina Garbeva

    Full Text Available BACKGROUND: Many soil-inhabiting bacteria are known to produce secondary metabolites that can suppress microorganisms competing for the same resources. The production of antimicrobial compounds is expected to incur fitness costs for the producing bacteria. Such costs form the basis for models on the co-existence of antibiotic-producing and non-antibiotic producing strains. However, so far studies quantifying the costs of antibiotic production by bacteria are scarce. The current study reports on possible costs, for antibiotic production by Pseudomonas fluorescens Pf0-1, a soil bacterium that is induced to produce a broad-spectrum antibiotic when it is confronted with non-related bacterial competitors or supernatants of their cultures. METHODOLOGY AND PRINCIPAL FINDINGS: We measured the possible cost of antibiotic production for Pseudomonas fluorescens Pf0-1 by monitoring changes in growth rate with and without induction of antibiotic production by supernatant of a bacterial competitor, namely Pedobacter sp.. Experiments were performed in liquid as well as on semi-solid media under nutrient-limited conditions that are expected to most clearly reveal fitness costs. Our results did not reveal any significant costs for production of antibiotics by Pseudomonas fluorescens Pf0-1. Comparison of growth rates of the antibiotic-producing wild-type cells with those of non-antibiotic producing mutants did not reveal costs of antibiotic production either. SIGNIFICANCE: Based on our findings we propose that the facultative production of antibiotics might not be selected to mitigate metabolic costs, but instead might be advantageous because it limits the risk of competitors evolving resistance, or even the risk of competitors feeding on the compounds produced.

  7. Presence and antimicrobial profile of gram-negative facultative anaerobe rods in patients with chronic periodontitis and gingivitis.

    Science.gov (United States)

    Gamboa, Fredy; García, Dabeiba-Adriana; Acosta, Adriana; Mizrahi, Deborah; Paz, Andreína; Martínez, Diana; Arévalo, Azucena; Aristizabal, Fabio; Abba, Martín

    2013-01-01

    Chronic periodontitis is a multifactorial infectious disease associated with Gram-negative anaerobes which are part of the subgingival microflora. In recent years, studies have been conducted to assess the presence of Gram-negative facultative anaerobes (Enterobacteriaceae) and their participation in the development and progression of chronic periodontitis. The aim of this study was to determine the presence of Enterobacteriaceae in patients with chronic periodontitis and gingivitis and to assess antimicrobial susceptibility of clinical isolates. A descriptive, observational study was performed including 64 patients with chronic periodontitis and 22 patients with gingivitis. Microbiological samples were taken from the gingival sulcus using paper points, which then were placed in thioglycollate broth. Samples were incubated for 4 hours at 37 degrees C and finally replated on MacConkey agar Bacteria were identified using the API-20E system (Biomerieux, France) and antimicrobial susceptibility was determined using the disk diffusion method. The evaluation of samples showed presence of 29 enterobacterial species distributed as follows: 7 in the group with gingivitis and 22 in the group with chronic periodontitis. In the chronic periodontitis group the most common species were: K. oxytoca n = 5, S. liquefaciens n = 4 and K. pneumoniae and E. coli with n = 3. The gingivitis group had the highest frequency of Erwinia sp. (n = 2). Clinical isolates showed very low sensitivity levels to beta-lactam ampicillin and amoxicillin/ clavulanic acid, 17.2% and 27.6% respectively, and higher sensitivity levels to ciprofloxacin (96.6%), amikacin (79.3%), gentamicin (68.9%) and ceftazidime, ceftriaxone, kanamycin and trimethoprimsulfa (65.5%). In conclusion, the existence of a high frequency of enterobacteria in patients with chronic periodontitis and gingivitis shows that periodontologists should pay greater attention to prevention protocols, and develop mechanical and antimicrobial

  8. Biological decomposition of Na2S2O3 into sulfur by a newly isolated facultative thermophilic alkaline desulphuricant strain

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    A facultative thermophilic alkaline desulphuricant strain named GDJ-3 was isolated from neutral soils,enriched on sulfur synthetic medium,and detected in previous work.Conventional and chemotax-onomic analyses and 16s RNA gene sequencing showed that the strain was in good agreement with Alpha proteobacterium sp.(97%) and ochrobactrum sp.(98%).The regenerative processes of the solu-tion containing sulfur compounds(SCS) from the strain through an orthogonal test were investigated to get the optimum regenerative condition.The results showed that regenerative temperature,air flow,and stirring speed of the agitator were the main three variables influencing the regenerative processes of the SCS.The optimum regenerative efficiency of the SCS from the strain was obtained when tem-perature,air flow,and stirring speed of the agitator were 318.2 K,3.0 L/min,and zero r/min,respectively.Under this condition,when the cell concentration of the strain was adjusted to 107/mL,the concentra-tions of Na2S2O3 and Na2S in the SCS decreased from 112.68 g/L to 96.88 g/L and from 0.87 g/L to 0.11 g/L in 9.5 h.Meanwhile,XRD spectrum shows that sulfur was formed in the regeneration process.These results suggest that the strain has potential application to the regeneration of the industrial so-lution containing sulfur compounds.

  9. Iron, Sulfur, Arsenic and Water: Geochemical Implications of Facultative Anoxygenic Photosynthesis in Cyanobacteria and the Slow Rise of Oxygen

    Science.gov (United States)

    Wolfe-Simon, F.; Johnston, D. T.; Girguis, P. R.; Pearson, A.; Knoll, A. H.

    2008-12-01

    Over geologic time, the global rise in atmospheric oxygen (O2) is attributed to the evolution and wide spread proliferation of oxygenic photosynthesis in cyanobacteria. However, cyanobacteria maintain a metabolic flexibility that may not always result in O2 release. Specifically, cyanobacteria can use a variety of alternative electron donors, rather than water, that are also readily oxidized. These may include sulfur, iron, and arsenic. Cyanobacteria are thus not uniquely constrained towards O2 production. Changes in the bioavailability of these key elements may have had dramatic consequences for and resulted in the slow accumulation of O2 in the atmosphere. In particular, by using facultative anoxygenic photosynthesis the cells maintain advantageous anaerobic conditions for N2-fixation. Although other types of bacteria are capable of N2-fixation, cyanobacteria singularly possess the dynamic capability of generating and surviving O2. These two processes "pull" the cells in opposite directions, metabolically speaking, around an aerobic-anaerobic continuum. Such a strategy also confers a distinct competitive advantage for cyanobacteria over photosynthetic eukaryotes, as they can endure widespread euxinia and maintain their cellular N quota. In an anoxic and/or sulfidic ocean, cyanobacteria would be expected to dominate over eukaryotic algae. Here we present Bayesian constructed phylogenetic distribution of specific genes and the metabolic role of key enzymes that form the basis of this hypothesis. We further suggest that the consequences of this proposed ecosystem structure altered the redox balance of the fluid Earth (atmosphere and oceans) and can help explain the observed long-term geochemical stasis and slow rates of eukaryotic diversification. We suggest that the underlying control for global oxygenation was a synergistic interplay between the evolution and elastic physiology of cyanobacteria as they impacted the redox state of early Earth.

  10. [Magnetic nanoparticles and intracellular delivery of biopolymers].

    Science.gov (United States)

    Kornev, A A; Dubina, M V

    2014-03-01

    The basic methods of intracellular delivery of biopolymers are present in this review. The structure and synthesis of magnetic nanoparticles, their stabilizing surfactants are described. The examples of the interaction of nanoparticles with biopolymers such as nucleic acids and proteins are considered. The final part of the review is devoted to problems physiology and biocompatibility of magnetic nanoparticles.

  11. Intracellular mechanisms of solar water disinfection

    Science.gov (United States)

    Castro-Alférez, María; Polo-López, María Inmaculada; Fernández-Ibáñez, Pilar

    2016-12-01

    Solar water disinfection (SODIS) is a zero-cost intervention measure to disinfect drinking water in areas of poor access to improved water sources, used by more than 6 million people in the world. The bactericidal action of solar radiation in water has been widely proven, nevertheless the causes for this remain still unclear. Scientific literature points out that generation of reactive oxygen species (ROS) inside microorganisms promoted by solar light absorption is the main reason. For the first time, this work reports on the experimental measurement of accumulated intracellular ROS in E. coli during solar irradiation. For this experimental achievement, a modified protocol based on the fluorescent probe dichlorodihydrofluorescein diacetate (DCFH-DA), widely used for oxidative stress in eukaryotic cells, has been tested and validated for E. coli. Our results demonstrate that ROS and their accumulated oxidative damages at intracellular level are key in solar water disinfection.

  12. Dynamics of gradient formation by intracellular shuttling

    Energy Technology Data Exchange (ETDEWEB)

    Berezhkovskii, Alexander M. [Mathematical and Statistical Computing Laboratory, Division of Computational Bioscience, Center for Information Technology, National Institutes of Health, Bethesda, Maryland 20892 (United States); Shvartsman, Stanislav Y. [Department of Chemical and Biological Engineering and Lewis-Sigler Institute for Integrative Genomics, Princeton University, Princeton, New Jersey 08544 (United States)

    2015-08-21

    A number of important cellular functions rely on the formation of intracellular protein concentration gradients. Experimental studies discovered a number of mechanisms for the formation of such gradients. One of the mechanisms relies on the intracellular shuttling of a protein that interconverts between the two states with different diffusivities, under the action of two enzymes, one of which is localized to the plasma membrane, whereas the second is uniformly distributed in the cytoplasm. Recent work reported an analytical solution for the steady state gradient in this mechanism, obtained in the framework of a one-dimensional reaction-diffusion model. Here, we study the dynamics in this model and derive analytical expressions for the Laplace transforms of the time-dependent concentration profiles in terms of elementary transcendental functions. Inverting these transforms numerically, one can obtain time-dependent concentration profiles of the two forms of the protein.

  13. Dynamics of gradient formation by intracellular shuttling

    Science.gov (United States)

    Berezhkovskii, Alexander M.; Shvartsman, Stanislav Y.

    2015-08-01

    A number of important cellular functions rely on the formation of intracellular protein concentration gradients. Experimental studies discovered a number of mechanisms for the formation of such gradients. One of the mechanisms relies on the intracellular shuttling of a protein that interconverts between the two states with different diffusivities, under the action of two enzymes, one of which is localized to the plasma membrane, whereas the second is uniformly distributed in the cytoplasm. Recent work reported an analytical solution for the steady state gradient in this mechanism, obtained in the framework of a one-dimensional reaction-diffusion model. Here, we study the dynamics in this model and derive analytical expressions for the Laplace transforms of the time-dependent concentration profiles in terms of elementary transcendental functions. Inverting these transforms numerically, one can obtain time-dependent concentration profiles of the two forms of the protein.

  14. Anoxybacillusgeothermalis sp. nov., a facultatively anaerobic, endospore-forming bacterium isolated from mineral deposits in a geothermal station.

    Science.gov (United States)

    Filippidou, Sevasti; Jaussi, Marion; Junier, Thomas; Wunderlin, Tina; Jeanneret, Nicole; Palmieri, Fabio; Palmieri, Ilona; Roussel-Delif, Ludovic; Vieth-Hillebrand, Andrea; Vetter, Alexandra; Chain, Patrick S; Regenspurg, Simona; Junier, Pilar

    2016-08-01

    A novel endospore-forming bacterium designated strain GSsed3T was isolated from deposits clogging aboveground filters from the geothermal power platform of Groß Schönebeck in northern Germany. The novel isolate was Gram-staining-positive, facultatively anaerobic, catalase-positive and oxidase-positive. Optimum growth occurred at 60 °C, 0.5 % (w/v) NaCl and pH 7-8. Analysis of the 16S rRNA gene sequence similarity indicated that strain GSsed3T belonged to the genus Anoxybacillus, and showed 99.8 % sequence similarity to Anoxybacillus rupiensis R270T, 98.2 % similarity to Anoxybacillus tepidamans GS5-97T, 97.9 % similarity to Anoxybacillus voinovskiensis TH13T, 97.7 % similarity to Anoxybacillus caldiproteolyticus DSM 15730T and 97.6 % similarity to Anoxybacillus amylolyticus MR3CT. DNA-DNA hybridization (DDH) indicated only 16 % relatedness to Anoxybacillus rupiensis DSM 17127T. Furthermore, DDH estimation based on genomes analysis indicated only 19.9 % overall nucleotide similarity to Anoxybacillus amylolyticus DSM 15939T. The major respiratory menaquinone was MK-8. The polar lipid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, one unknown phosphoglycolipid and one unknown phospholipid. The predominant cellular fatty acids were iso-C15 : 0, iso-C17 : 0, C16 : 0, iso-C16 : 0 and anteiso-C17 : 0. The peptidoglycan type was A1γ meso-Dpm-direct. The genomic DNA G+C content of the strain was 46.9 mol%. The phenotypic, genotypic and chemotaxonomic characterization indicated that strain GSsed3T differs from related species of the genus. Therefore, strain GSsed3T is considered to be a representative of a novel species of the genus Anoxybacillus, for which the name Anoxybacillus geothermalis sp. nov. is proposed. The type strain of Anoxybacillus geothermalis is GSsed3T (=CCOS808T =ATCC BAA2555T).

  15. Biology and intracellular life of chlamydia

    Directory of Open Access Journals (Sweden)

    Ranin Lazar

    2011-01-01

    Full Text Available Introduction. Chlamydiae are Gram-negative obligate intracellular bacteria. The developmental cycle of Chlamydiae is specific and different from other bacteria. The elementary body is the infectious form of the organism, responsible for attaching to the target host cell and promoting its entry. The reticulate body is the larger, metabolically active form of the organism, synthesizing deoxyribonucleic acid, ribonucleic acid and proteins. The elementary body and reticulate body represent evolutionary adaptations to extracellular and intracellular environments. Intracellular persistence of Chlamydia. Predisposition of Chlamydia to persist within the host cell has been recognized as a major factor in the pathogenesis of chlamydial disease. The persistence implies a long-term association between chlamydiae and their host cell that may not manifest as clinically recognizable disease. The ability of chlamydia to remain within one morphological state for a long time in response to exogenous factors suggests an innate ability of these organisms to persist intracellulary in a unique developmental form. Chlamydiae induce interferon γ and exhibit growth inhibition in their presence. While the high levels of interferon γ completely restrict the development of chlamydia, its low levels induce the development of morphologically aberrant intracellular forms. The persistent forms contain reduced levels of major outer membrane protein but high levels of chlamydial heat shock protein. Conclusion. Immunopathogenesis of chlamydial infection is one of the main focal points of current research into Chlamydia. Chlamydial infections are highly prevalent, usually asymptomatic and associated with serious sequelae. Screening programmes are the most important in the prevention of a long-term sequele.

  16. Fluorescent nanoparticles for intracellular sensing: A review

    Energy Technology Data Exchange (ETDEWEB)

    Ruedas-Rama, Maria J., E-mail: mjruedas@ugr.esmailto [Department of Physical Chemistry, Faculty of Pharmacy, University of Granada, Campus Cartuja, 18071, Granada (Spain); Walters, Jamie D. [Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, UK CB2 1QT (United Kingdom); Orte, Angel [Department of Physical Chemistry, Faculty of Pharmacy, University of Granada, Campus Cartuja, 18071, Granada (Spain); Hall, Elizabeth A.H., E-mail: lisa.hall@biotech.cam.ac.uk [Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QT (United Kingdom)

    2012-11-02

    Highlights: Black-Right-Pointing-Pointer Analytical applications of fluorescent nanoparticles (NPs) in intracellular sensing. Black-Right-Pointing-Pointer Critical review on performance of QDots, metal NPs, silica NPs, and polymer NPs. Black-Right-Pointing-Pointer Highlighted potential of fluorescence lifetime imaging microscopy (FLIM). - Abstract: Fluorescent nanoparticles (NPs), including semiconductor NPs (Quantum Dots), metal NPs, silica NPs, polymer NPs, etc., have been a major focus of research and development during the past decade. The fluorescent nanoparticles show unique chemical and optical properties, such as brighter fluorescence, higher photostability and higher biocompatibility, compared to classical fluorescent organic dyes. Moreover, the nanoparticles can also act as multivalent scaffolds for the realization of supramolecular assemblies, since their high surface to volume ratio allow distinct spatial domains to be functionalized, which can provide a versatile synthetic platform for the implementation of different sensing schemes. Their excellent properties make them one of the most useful tools that chemistry has supplied to biomedical research, enabling the intracellular monitoring of many different species for medical and biological purposes. In this review, we focus on the developments and analytical applications of fluorescent nanoparticles in chemical and biological sensing within the intracellular environment. The review also points out the great potential of fluorescent NPs for fluorescence lifetime imaging microscopy (FLIM). Finally, we also give an overview of the current methods for delivering of fluorescent NPs into cells, where critically examine the benefits and liabilities of each strategy.

  17. Invasion and intracellular survival by protozoan parasites.

    Science.gov (United States)

    Sibley, L David

    2011-03-01

    Intracellular parasitism has arisen only a few times during the long ancestry of protozoan parasites including in diverse groups such as microsporidians, kinetoplastids, and apicomplexans. Strategies used to gain entry differ widely from injection (e.g. microsporidians), active penetration of the host cell (e.g. Toxoplasma), recruitment of lysosomes to a plasma membrane wound (e.g. Trypanosoma cruzi), to host cell-mediated phagocytosis (e.g. Leishmania). The resulting range of intracellular niches is equally diverse ranging from cytosolic (e.g. T. cruzi) to residing within a non-fusigenic vacuole (e.g. Toxoplasma, Encephalitozoon) or a modified phagolysosome (e.g. Leishmania). These lifestyle choices influence access to nutrients, interaction with host cell signaling pathways, and detection by pathogen recognition systems. As such, intracellular life requires a repertoire of adaptations to assure entry-exit from the cell, as well as to thwart innate immune mechanisms and prevent clearance. Elucidating these pathways at the cellular and molecular level may identify key steps that can be targeted to reduce parasite survival or augment immunologic responses and thereby prevent disease.

  18. Detection of intracellular phosphatidylserine in living cells.

    Science.gov (United States)

    Calderon, Frances; Kim, Hee-Yong

    2008-03-01

    To demonstrate the intracellular phosphatidylserine (PS) distribution in neuronal cells, neuroblastoma cells and hippocampal neurons expressing green fluorescence protein (GFP)-AnnexinV were stimulated with a calcium ionophore and localization of GFP-AnnexinV was monitored by fluorescence microscopy. Initially, GFP-AnnexinV distributed evenly in the cytosol and nucleus. Raising the intracellular calcium level with ionomycin-induced translocation of cytoplasmic GFP-AnnexinV to the plasma membrane but not to the nuclear membrane, indicating that PS distributes in the cytoplasmic side of the plasma membrane. Nuclear GFP-AnnexinV subsequently translocated to the nuclear membrane, indicating PS localization in the nuclear envelope. GFP-AnnexinV also localized in a juxtanuclear organelle that was identified as the recycling endosome. However, minimal fluorescence was detected in any other subcellular organelles including mitochondria, endoplasmic reticulum, Golgi complex, and lysosomes, strongly suggesting that PS distribution in the cytoplasmic face in these organelles is negligible. Similarly, in hippocampal primary neurons PS distributed in the inner leaflet of plasma membranes of cell body and dendrites, and in the nuclear envelope. To our knowledge, this is the first demonstration of intracellular PS localization in living cells, providing an insight for specific sites of PS interaction with soluble proteins involved in signaling processes.

  19. The measurement of constitutive and facultative skin pigmentation and estimation of sun exposure in Caucasians with basal cell carcinoma and cutaneous malignant melanoma

    DEFF Research Database (Denmark)

    Lock-Andersen, J; Drzewiecki, K T; Wulf, H C

    1998-01-01

    In two identical and simultaneously performed case-control studies of basal cell carcinoma (BCC) and cutaneous malignant melanoma (CMM) with age-matched, sex-matched and residence-matched controls, skin pigmentation was measured objectively by skin reflectance spectroscopy in 145 BCC patients...... by all subjects. There were no statistically significant differences in constitutive skin pigmentation at the buttocks between BCC patients and controls (P = 0.96) or between CMM patients and controls (P = 0.13). Facultative skin pigmentation in ultraviolet-exposed sites was not significantly different...

  20. Strategies to improve intracellular drug delivery by targeted liposomes

    NARCIS (Netherlands)

    Fretz, M.M.

    2007-01-01

    Biotechnological advances increased the number of novel macromolecular drugs and new drug targets. The latter are mostly found intracellular. Unfortunately, most of the new macromolecular drugs rely on drug delivery tools for their intracellular delivery because their unfavourable physicochemical pr

  1. Intracellular ethanol accumulation in Saccharomyces cerevisiae during fermentation.

    OpenAIRE

    D'Amore, T; C.J. Panchal; Stewart, G G

    1988-01-01

    An intracellular accumulation of ethanol in Saccharomyces cerevisiae was observed during the early stages of fermentation (3 h). However, after 12 h of fermentation, the intracellular and extracellular ethanol concentrations were similar. Increasing the osmotic pressure of the medium caused an increase in the ratio of intracellular to extracellular ethanol concentrations at 3 h of fermentation. As in the previous case, the intracellular and extracellular ethanol concentrations were similar af...

  2. Milk contamination in different points of the dairy process. ii mesophilic, psychrotrophic and proteolytic microorganisms/ Contaminação do leite em diferentes pontos da produção leiteira: ii microrganismos mesófilos, psicrotróficos e proteolíticos

    Directory of Open Access Journals (Sweden)

    Viviane Vieira Gusmão

    2004-05-01

    Full Text Available Psychrotrophics microorganisms are able to multiply under refrigeration temperatures. They can produce heat-resistant extracellular enzymes that cause organoleptic and structural changes in milk and dairy products. With the objective of determining the frequency of microorganisms in the dairy process, four farms were evaluated in Londrina city – Brazil and different points were assessed. Mesophilic (48hs at 35oC, psychrotrophic (25hs at 21oC and proteolytic (72hs at 22oC microorganisms were researched, and the proteolytic predominant microflora were determined by using the Gram stain technique. The main psychrotrophic contamination points were the residual water on milk cans, on bulk tank and bad cleaned teats, and milk cans and teats were the most important sources of proteolytics. Gram negative bacilli represented 69% of the microorganisms in the refrigerated milk that means 45% of the total proteolytics microorganisms in the dairy process. Gram negative bacilli is most frequent on bad cleaned teats, teatcups, residual water in milk cans and bulk tank. The psychrotroph count in the refrigerated milk was more than the mesophilic count, showing that that psychrotrophics are the best indicatator for refrigerated milk contamination. The psycrotrophics count at all points assessed were greater than the ideal limit (10% of the mesophilic count. Thus as refrigeration at 4oC is not sufficient to control the multiplication of the psycrotrophics group, good practices should be associated to avoid or control milk contamination by psycrotrophics.Os psicrotróficos são microrganismos que têm capacidade de multiplicação em temperaturas de refrigeração. Produzem enzimas termorresistentes como proteases e lipases que promovem alterações organolépticas no leite e comprometem a produção de derivados. Com o objetivo de determinar a freqüência desses microrganismos a produção do leite, foram estudadas 04 propriedades da região de Londrina, PR

  3. Facultés de droit en crise : formation et socialisation des élites allemandes sous la République de Weimar

    Directory of Open Access Journals (Sweden)

    Marie-Bénédicte Vincent

    2006-05-01

    Full Text Available L’article se propose d’explorer l’univers des facultés de droit sous la République de Weimar, que les contemporains jugent en « crise ». Cette perception renvoie tout d’abord aux difficultés d’adaptation d’un enseignement qui est de plus en plus écartelé entre les exigences de la science (transmettre une compréhension historique de l’évolution du droit et celles de la pratique (préparer les étudiants au monde professionnel par une connaissance du droit en vigueur : l’Université apparaît ainsi comme un lieu de confrontation entre professeurs et autorités étatiques pour le contrôle du cursus juridique. Par ailleurs, dans un contexte de grande expansion des effectifs étudiants, les facultés de droit ne connaissent pas de démocratisation de leur public et restent le domaine des élites traditionnelles ; de ce fait, elles semblent en décalage avec leur époque et continuent de former un univers passéiste que conforte l’engagement des étudiants dans des corporations traditionnelles. Ce mode de socialisation explique leur radicalisation dans le camp antirépublicain dès les années 1920.

  4. Bacterial Microbiota Associated with the Glacier Ice Worm Is Dominated by Both Worm-Specific and Glacier-Derived Facultative Lineages

    Science.gov (United States)

    Murakami, Takumi; Segawa, Takahiro; Dial, Roman; Takeuchi, Nozomu; Kohshima, Shiro; Hongoh, Yuichi

    2017-01-01

    The community structure of bacteria associated with the glacier ice worm Mesenchytraeus solifugus was analyzed by amplicon sequencing of 16S rRNA genes and their transcripts. Ice worms were collected from two distinct glaciers in Alaska, Harding Icefield and Byron Glacier, and glacier surfaces were also sampled for comparison. Marked differences were observed in bacterial community structures between the ice worm and glacier surface samples. Several bacterial phylotypes were detected almost exclusively in the ice worms, and these bacteria were phylogenetically affiliated with either animal-associated lineages or, interestingly, clades mostly consisting of glacier-indigenous species. The former included bacteria that belong to Mollicutes, Chlamydiae, Rickettsiales, and Lachnospiraceae, while the latter included Arcicella and Herminiimonas phylotypes. Among these bacteria enriched in ice worm samples, Mollicutes, Arcicella, and Herminiimonas phylotypes were abundantly and consistently detected in the ice worm samples; these phylotypes constituted the core microbiota associated with the ice worm. A fluorescence in situ hybridization analysis showed that Arcicella cells specifically colonized the epidermis of the ice worms. Other bacterial phylotypes detected in the ice worm samples were also abundantly recovered from the respective habitat glaciers; these bacteria may be food for ice worms to digest or temporary residents. Nevertheless, some were overrepresented in the ice worm RNA samples; they may also function as facultative gut bacteria. Our results indicate that the community structure of bacteria associated with ice worms is distinct from that in the associated glacier and includes worm-specific and facultative, glacier-indigenous lineages. PMID:28302989

  5. Transgenic insect-resistant corn affects the fourth trophic level: effects of Bacillus thuringiensis-corn on the facultative hyperparasitoid Tetrastichus howardi

    Science.gov (United States)

    Prütz, Gernot; Brink, Andreas; Dettner, Konrad

    2004-09-01

    As hyperparasitoids may have a considerable influence on the control of herbivorous arthropods, analyzing the host-mediated impact of Bacillus thuringiensis-plants (“Bt-plants”) on hyperparasitoids is of interest. Laboratory-scale experiments were conducted in order to assess the potential effect of Bt-corn leaf material on the facultative hyperparasitoid Tetrastichus howardi (Olliff) (Hymenoptera: Eulophidae), mediated through the herbivore Chilo partellus Swinhoe (Lepidoptera: Crambidae) and its primary parasitoid Cotesia flavipes (Cameron) (Hymenoptera: Braconidae). In the Bt-group, significantly less C. flavipes larvae per host spun cocoons and pupated than in the control, and the mean fresh weight of a single C. flavipes cocoon was significantly reduced compared to the control. All C. flavipes cocoons of one host formed cocoon clusters. T. howardi females of the Bt-group parasitized significantly less cocoon clusters than in the control. Moreover, significantly fewer C. flavipes cocoons per cocoon cluster were successfully parasitized as compared to the control. As a consequence, T. howardi females of the control had more offspring than in the Bt-group. Adult female T. howardi offspring of the Bt-group weighed significantly less than in the control, but there was no significant weight difference between males of both groups. Our results suggest that transgenic insect-resistant plants could affect hyperparasitoids indirectly. However, it remains to be determined whether facultative hyperparasitoids prefer to develop as primary or secondary parasitoids under field conditions.

  6. Evidence for a facultative mutualist nutritional relationship between the green coccoid alga Bracteacoccus sp. (Chlorophyceae) and the zoosporic fungus Rhizidium phycophilum (Chytridiomycota).

    Science.gov (United States)

    Picard, Kathryn T; Letcher, Peter M; Powell, Martha J

    2013-05-01

    Symbiotic interactions between fungi and photosynthetic partners are common among derived fungal lineages. The only fungal-phototroph interactions thus far reported from the early diverging zoosporic fungi are parasitic in nature. Rhizidium phycophilum is a terrestrial, saprotrophic chytrid, which appears to be able to enter a facultative mutualism with a coccoid green alga in the absence of refractory organic material, such as pollen and chitin. Liquid and solid culturing methods were used in a series of differential fitness experiments in conjunction with microscopic analyses to characterize the interaction between R. phycophilum and the alga. The alga in this partnership is identified as a member of the genus Bracteacoccus. Under certain culturing conditions, algal cells grown in coculture with R. phycophilum were shown to grow larger and more prolifically than when cultured axenically under the same conditions. Additionally, dialysis experiments demonstrate that R. phycophilum does not parasitize Bracteacoccus sp., and can be cultured in media infused with unknown algal exudates. Rhizidium phycophilum and Bracteacoccus sp. represent the first facultative positive interaction between a zoosporic fungus and a photoautotroph and may prove a tractable system for modelling interactions between early fungi and plants. Copyright © 2013 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  7. An Invertron-Like Linear Plasmid Mediates Intracellular Survival and Virulence in Bovine Isolates of Rhodococcus equi.

    Science.gov (United States)

    Valero-Rello, Ana; Hapeshi, Alexia; Anastasi, Elisa; Alvarez, Sonsiray; Scortti, Mariela; Meijer, Wim G; MacArthur, Iain; Vázquez-Boland, José A

    2015-07-01

    We report a novel host-associated virulence plasmid in Rhodococcus equi, pVAPN, carried by bovine isolates of this facultative intracellular pathogenic actinomycete. Surprisingly, pVAPN is a 120-kb invertron-like linear replicon unrelated to the circular virulence plasmids associated with equine (pVAPA) and porcine (pVAPB variant) R. equi isolates. pVAPN is similar to the linear plasmid pNSL1 from Rhodococcus sp. NS1 and harbors six new vap multigene family members (vapN to vapS) in a vap pathogenicity locus presumably acquired via en bloc mobilization from a direct predecessor of equine pVAPA. Loss of pVAPN rendered R. equi avirulent in macrophages and mice. Mating experiments using an in vivo transconjugant selection strategy demonstrated that pVAPN transfer is sufficient to confer virulence to a plasmid-cured R. equi recipient. Phylogenetic analyses assigned the vap multigene family complement from pVAPN, pVAPA, and pVAPB to seven monophyletic clades, each containing plasmid type-specific allelic variants of a precursor vap gene carried by the nearest vap island ancestor. Deletion of vapN, the predicted "bovine-type" allelic counterpart of vapA, essential for virulence in pVAPA, abrogated pVAPN-mediated intramacrophage proliferation and virulence in mice. Our findings support a model in which R. equi virulence is conferred by host-adapted plasmids. Their central role is mediating intracellular proliferation in macrophages, promoted by a key vap determinant present in the common ancestor of the plasmid-specific vap islands, with host tropism as a secondary trait selected during coevolution with specific animal species. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  8. Azithromycin effectiveness against intracellular infections of Francisella

    Directory of Open Access Journals (Sweden)

    Mann Barbara J

    2010-04-01

    Full Text Available Abstract Background Macrolide antibiotics are commonly administered for bacterial respiratory illnesses. Azithromycin (Az is especially noted for extremely high intracellular concentrations achieved within macrophages which is far greater than the serum concentration. Clinical strains of Type B Francisella (F. tularensis have been reported to be resistant to Az, however our laboratory Francisella strains were found to be sensitive. We hypothesized that different strains/species of Francisella (including Type A may have different susceptibilities to Az, a widely used and well-tolerated antibiotic. Results In vitro susceptibility testing of Az confirmed that F. tularensis subsp. holarctica Live Vaccine Strain (LVS (Type B was not sensitive while F. philomiragia, F. novicida, and Type A F. tularensis (NIH B38 and Schu S4 strain were susceptible. In J774A.1 mouse macrophage cells infected with F. philomiragia, F. novicida, and F. tularensis LVS, 5 μg/ml Az applied extracellularly eliminated intracellular Francisella infections. A concentration of 25 μg/ml Az was required for Francisella-infected A549 human lung epithelial cells, suggesting that macrophages are more effective at concentrating Az than epithelial cells. Mutants of RND efflux components (tolC and ftlC in F. novicida demonstrated less sensitivity to Az by MIC than the parental strain, but the tolC disc-inhibition assay demonstrated increased sensitivity, indicating a complex role for the outer-membrane transporter. Mutants of acrA and acrB mutants were less sensitive to Az than the parental strain, suggesting that AcrAB is not critical for the efflux of Az in F. novicida. In contrast, F. tularensis Schu S4 mutants ΔacrB and ΔacrA were more sensitive than the parental strain, indicating that the AcrAB may be important for Az efflux in F. tularensis Schu S4. F. novicida LPS O-antigen mutants (wbtN, wbtE, wbtQ and wbtA were found to be less sensitive in vitro to Az compared to the wild

  9. Caspases rule the intracellular trafficking cartel.

    Science.gov (United States)

    Duclos, Catherine; Lavoie, Christine; Denault, Jean-Bernard

    2017-05-01

    During apoptosis, caspases feast on several hundreds of cellular proteins to orchestrate rapid cellular demise. Indeed, caspases are known to get a taste of every cellular process in one way or another, activating some, but most often shutting them down. Thus, it is not surprising that caspases proteolyze proteins involved in intracellular trafficking with particularly devastating consequences for this important process. This review article focuses on how caspases target the machinery responsible for smuggling goods within and outside the cell. © 2017 Federation of European Biochemical Societies.

  10. Intracellular pH in sperm physiology.

    Science.gov (United States)

    Nishigaki, Takuya; José, Omar; González-Cota, Ana Laura; Romero, Francisco; Treviño, Claudia L; Darszon, Alberto

    2014-08-01

    Intracellular pH (pHi) regulation is essential for cell function. Notably, several unique sperm ion transporters and enzymes whose elimination causes infertility are either pHi dependent or somehow related to pHi regulation. Amongst them are: CatSper, a Ca(2+) channel; Slo3, a K(+) channel; the sperm-specific Na(+)/H(+) exchanger and the soluble adenylyl cyclase. It is thus clear that pHi regulation is of the utmost importance for sperm physiology. This review briefly summarizes the key components involved in pHi regulation, their characteristics and participation in fundamental sperm functions such as motility, maturation and the acrosome reaction.

  11. Mechanisms of cellular invasion by intracellular parasites.

    Science.gov (United States)

    Walker, Dawn M; Oghumu, Steve; Gupta, Gaurav; McGwire, Bradford S; Drew, Mark E; Satoskar, Abhay R

    2014-04-01

    Numerous disease-causing parasites must invade host cells in order to prosper. Collectively, such pathogens are responsible for a staggering amount of human sickness and death throughout the world. Leishmaniasis, Chagas disease, toxoplasmosis, and malaria are neglected diseases and therefore are linked to socio-economical and geographical factors, affecting well-over half the world's population. Such obligate intracellular parasites have co-evolved with humans to establish a complexity of specific molecular parasite-host cell interactions, forming the basis of the parasite's cellular tropism. They make use of such interactions to invade host cells as a means to migrate through various tissues, to evade the host immune system, and to undergo intracellular replication. These cellular migration and invasion events are absolutely essential for the completion of the lifecycles of these parasites and lead to their for disease pathogenesis. This review is an overview of the molecular mechanisms of protozoan parasite invasion of host cells and discussion of therapeutic strategies, which could be developed by targeting these invasion pathways. Specifically, we focus on four species of protozoan parasites Leishmania, Trypanosoma cruzi, Plasmodium, and Toxoplasma, which are responsible for significant morbidity and mortality.

  12. Mucolipins: Intracellular TRPML1-3 channels.

    Science.gov (United States)

    Cheng, Xiping; Shen, Dongbiao; Samie, Mohammad; Xu, Haoxing

    2010-05-17

    The mucolipin family of Transient Receptor Potential (TRPML) proteins is predicted to encode ion channels expressed in intracellular endosomes and lysosomes. Loss-of-function mutations of human TRPML1 cause type IV mucolipidosis (ML4), a childhood neurodegenerative disease. Meanwhile, gain-of-function mutations in the mouse TRPML3 result in the varitint-waddler (Va) phenotype with hearing and pigmentation defects. The broad spectrum phenotypes of ML4 and Va appear to result from certain aspects of endosomal/lysosomal dysfunction. Lysosomes, traditionally believed to be the terminal "recycling center" for biological "garbage", are now known to play indispensable roles in intracellular signal transduction and membrane trafficking. Studies employing animal models and cell lines in which TRPML genes have been genetically disrupted or depleted have uncovered roles of TRPMLs in multiple cellular functions including membrane trafficking, signal transduction, and organellar ion homeostasis. Physiological assays of mammalian cell lines in which TRPMLs are heterologously overexpressed have revealed the channel properties of TRPMLs in mediating cation (Ca(2+)/Fe(2+)) efflux from endosomes and lysosomes in response to unidentified cellular cues. This review aims to summarize these recent advances in the TRPML field and to correlate the channel properties of endolysosomal TRPMLs with their biological functions. We will also discuss the potential cellular mechanisms by which TRPML deficiency leads to neurodegeneration.

  13. Stochastic models of intracellular calcium signals

    Energy Technology Data Exchange (ETDEWEB)

    Rüdiger, Sten, E-mail: sten.ruediger@physik.hu-berlin.de

    2014-01-10

    Cellular signaling operates in a noisy environment shaped by low molecular concentrations and cellular heterogeneity. For calcium release through intracellular channels–one of the most important cellular signaling mechanisms–feedback by liberated calcium endows fluctuations with critical functions in signal generation and formation. In this review it is first described, under which general conditions the environment makes stochasticity relevant, and which conditions allow approximating or deterministic equations. This analysis provides a framework, in which one can deduce an efficient hybrid description combining stochastic and deterministic evolution laws. Within the hybrid approach, Markov chains model gating of channels, while the concentrations of calcium and calcium binding molecules (buffers) are described by reaction–diffusion equations. The article further focuses on the spatial representation of subcellular calcium domains related to intracellular calcium channels. It presents analysis for single channels and clusters of channels and reviews the effects of buffers on the calcium release. For clustered channels, we discuss the application and validity of coarse-graining as well as approaches based on continuous gating variables (Fokker–Planck and chemical Langevin equations). Comparison with recent experiments substantiates the stochastic and spatial approach, identifies minimal requirements for a realistic modeling, and facilitates an understanding of collective channel behavior. At the end of the review, implications of stochastic and local modeling for the generation and properties of cell-wide release and the integration of calcium dynamics into cellular signaling models are discussed.

  14. Gene Therapy for HIV Infections: Intracellular Immunization

    Directory of Open Access Journals (Sweden)

    Alain Piché

    1999-01-01

    Full Text Available Despite significant advances in the treatment of human immunodeficiency virus (HIV infection in the past 10 years, it remains an incurable disease. The inability of traditional drug-based therapies to inhibit HIV replication effectively for extended periods of time has stimulated intense research to develop novel approaches for this disease. Current understanding of HIV molecular biology and pathogenesis has opened the way for the development of gene therapy strategies for HIV infections. In this context, a number of intracellular immunization-based strategies have been evaluated, and some of them have reached the stage of phase I/II human clinical trials. These strategies include the use of single-chain antibodies, capsid-targeted viral inactivation, transdominant negative mutants, ribozymes, antisense oligonucleotides and RNA decoys. While a number of issues remain to be studied before intracellular immunization can be applied to the treatment of HIV infections, the significant progress already made in this field is likely to lead to clinical applications.

  15. Intracellular accumulation of norfloxacin in Mycobacterium smegmatis.

    Science.gov (United States)

    Corti, S; Chevalier, J; Cremieux, A

    1995-11-01

    To evaluate the intracellular accumulation of norfloxacin in mycobacteria, two methods were used with Mycobacterium smegmatis. A radiometric method (K. V. Cundy, C. E. Fasching, K. E. Willard, and L. R. Peterson, J. Antimicrob. Chemother. 28:491-497, 1991) was used without great modification, but the fluorometric method (P. G. S. Mortimer and L. J. V. Piddock, J. Antimicrob. Chemother. 28:639-653, 1991) was changed considerably. Indeed, adsorption of the quinolone to the bacterial surface was characterized by measuring the level of accumulation of 0 degree C. Taking into account the adsorption, the pH of the washing buffer was increased from 7.0 to 9.0 to improve the desorption of norfloxacin from the cell surface. Both the fluorometric method, with the technical improvement, and the radiometric method could be used to estimate the intracellular accumulation of norfloxacin, which resulted from the difference between the whole uptake measured at 37 degrees C and the adsorption measured at 0 degrees C. A total of 35 ng of norfloxacin per mg of cells (dry weight) penetrated into the M. smegmatis cell, and the steady state was achieved in 5 min. Use of inhibitors of the proton motive force revealed that transport of norfloxacin was energy independent. Thus, the same mechanisms of quinolone accumulation that occur in eubacteria seem to occur in mycobacteria, at least in M. smegmatis.

  16. Intracellular signaling by diffusion: can waves of hydrogen peroxide transmit intracellular information in plant cells?

    DEFF Research Database (Denmark)

    Vestergaard, Christian L.; Flyvbjerg, Henrik; Møller, Ian Max

    2012-01-01

    Amplitude- and frequency-modulated waves of Ca(2+) ions transmit information inside cells. Reactive Oxygen Species (ROS), specifically hydrogen peroxide, have been proposed to have a similar role in plant cells. We consider the feasibility of such an intracellular communication system in view...

  17. 一种兼氧 FM BR 技术处理食品废水%A Facultative FMBR Technology for Treatment of Food Processing Wastewater

    Institute of Scientific and Technical Information of China (English)

    廖志民; 刘华琼; 刘宇森; 屈杨; 陶晶晶; 白洁; 余航

    2014-01-01

    文章介绍了一种兼氧膜生物反应器的兼氧FMBR技术处理深圳某食品加工废水.该兼氧膜生物反应器具有工艺简单、同步脱氮、不排放有机剩余污泥、节能高效、出水可回用等优点,适宜处理高浓度有机废水.实验结果表明,该工艺能有效去除CODCr 、N H3-N、T P和SS质量等污染物,处理后出水CODCr 、N H3-N、T P和SS质量浓度分别至50、2、0.5和10 mg/L ,去除率分别达98%、98%、87%和99%,满足GB8579-96污水综合排放标准.该工艺还实现了无人值守,且吨水占地面积小,单位处理成本低,经济实用.%This paper introduces the facultative FMBR technology of a facultative aerobic membrane bioreactor for treating the wastewater resulted by food processing in Shenzhen .The facultative aerobic membrane bioreactor as reported in this article has advantages such as simple processing ,simultaneous nitrogen ,without emitting organic sludge ,energy efficiency ,and the capability of recycling water ,which would be appropriate to be applied to treat high concentration organic wastewater .As the experimental results reveal ,this technology can effectively remove pollutants like CODCr ,NH3 -N ,TP and SS ,with the effluent concentration of 50 mg/L ,2 mg/L ,0 .5 mg/L ,and 10 mg/L respectively after the treatment ,therefore ,the removal rate would be 98% ,98% ,87% ,and 99% respec-tively ,which fully satisfies the "Integrated Wastewater Discharge Standard" (GB 8579 -96) .Additionally ,this technology enables unattended automation ,smaller covering areas for tons of water ,and lower unit processing costs ,w hich are economical and practical .

  18. Cytoskeletal network morphology regulates intracellular transport dynamics

    CERN Document Server

    Ando, David; Huang, Kerwyn Casey; Gopinathan, Ajay

    2016-01-01

    Intracellular transport is essential for maintaining proper cellular function in most eukaryotic cells, with perturbations in active transport resulting in several types of disease. Efficient delivery of critical cargos to specific locations is accomplished through a combination of passive diffusion and active transport by molecular motors that ballistically move along a network of cytoskeletal filaments. Although motor-based transport is known to be necessary to overcome cytoplasmic crowding and the limited range of diffusion within reasonable time scales, the topological features of the cytoskeletal network that regulate transport efficiency and robustness have not been established. Using a continuum diffusion model, we observed that the time required for cellular transport was minimized when the network was localized near the nucleus. In simulations that explicitly incorporated network spatial architectures, total filament mass was the primary driver of network transit times. However, filament traps that r...

  19. [Measurement of intracellular pH].

    Science.gov (United States)

    Hanaoka, K; Imai, M; Yoshitomi, K

    1992-09-01

    Since various cellular processes depend on changes in pH, the regulation of intracellular pH (pHi) is important both for the individual cell and for the organism. The mechanisms of the regulation of pHi can be investigated by monitoring pHi. In this report, we discuss the four major techniques available for measuring pHi, which are 1) Distribution of weak acids and bases, 2) pH-sensitive microelectrodes, 3) pH-sensitive dyes, and 4) Nuclear magnetic resonance. Among four techniques, the advantage of the microelectrode approach is that it can monitor membrane potential at the same time and be applied to a single cell. The dye technique is a relative new developing technique, which has lots of advantages. It is easy to use, and is capable of monitoring rapid pHi changes, and being applied to a smaller cell, or a single cell.

  20. Myometrial oxytocin receptor expression and intracellular pathways.

    Science.gov (United States)

    Yulia, A; Johnson, M R

    2014-06-01

    Oxytocin (OT) signalling plays a fundamental role in the mechanisms of parturition. OT is one of the most frequently used drugs in obstetrics, promoting uterine contractions for labor induction and augmentation and to prevent postpartum hemorrhage (PPH). Expression of the oxytocin receptor (OTR) in the human myometrium is tightly regulated during pregnancy and its levels have been shown to peak upon labour onset and to fall sharply in advanced labour and the postpartum period, when the uterus become refractive to OT. However, uterine sensitivity to OT varies between pregnant women, probably reflecting differences in their myometrial OTR expression. Control of OTR expression is mediated by a combination of steroid hormone stimulation, stretch, and inflammation. This review summarises current knowledge regarding the complex regulation of myometrial OTR expression and its associated intracellular signaling pathways.

  1. Intracellular calcium release modulates polycystin-2 trafficking

    Directory of Open Access Journals (Sweden)

    Miyakawa Ayako

    2013-02-01

    Full Text Available Abstract Background Polycystin-2 (PC2, encoded by the gene that is mutated in autosomal dominant polycystic kidney disease (ADPKD, functions as a calcium (Ca2+ permeable ion channel. Considerable controversy remains regarding the subcellular localization and signaling function of PC2 in kidney cells. Methods We investigated the subcellular PC2 localization by immunocytochemistry and confocal microscopy in primary cultures of human and rat proximal tubule cells after stimulating cytosolic Ca2+ signaling. Plasma membrane (PM Ca2+ permeability was evaluated by Fura-2 manganese quenching using time-lapse fluorescence microscopy. Results We demonstrated that PC2 exhibits a dynamic subcellular localization pattern. In unstimulated human or rat proximal tubule cells, PC2 exhibited a cytosolic/reticular distribution. Treatments with agents that in various ways affect the Ca2+ signaling machinery, those being ATP, bradykinin, ionomycin, CPA or thapsigargin, resulted in increased PC2 immunostaining in the PM. Exposing cells to the steroid hormone ouabain, known to trigger Ca2+ oscillations in kidney cells, caused increased PC2 in the PM and increased PM Ca2+ permeability. Intracellular Ca2+ buffering with BAPTA, inositol 1,4,5-trisphosphate receptor (InsP3R inhibition with 2-aminoethoxydiphenyl borate (2-APB or Ca2+/Calmodulin-dependent kinase inhibition with KN-93 completely abolished ouabain-stimulated PC2 translocation to the PM. Conclusions These novel findings demonstrate intracellular Ca2+-dependent PC2 trafficking in human and rat kidney cells, which may provide new insight into cyst formations in ADPKD.

  2. Intracellular mechanisms of lymphoid cell activation.

    Science.gov (United States)

    Fresa, K; Hameed, M; Cohen, S

    1989-01-01

    Activation of lymphocytes for proliferation is associated with the appearance of an intracellular factor (ADR) that can induce DNA synthesis in isolated quiescent nuclei. ADR plays a role in the sequence of intracellular events leading to activation for IL-2-mediated proliferation. Because of the nature of the defining assay, the locus of ADR action appears to be near the terminal end of the transduction pathway. Interestingly, although lymphocytes from aged individuals respond poorly to proliferative stimuli, they appear to produce normal to above-normal levels of ADR. In contrast, their nuclei are only poorly responsive to stimulation by ADR. Preparations rich in ADR activity have proteolytic activity as well. In addition, aprotinin, as well as a variety of other protease inhibitors, suppresses ADR-induced DNA synthesis in a dose-dependent manner. ADR activity can be removed from active extracts by absorption with aprotinin-conjugated agarose beads, and can be removed from the beads by elution at pH 5.0. This latter suggests that ADR itself is a protease. However, its endogenous substrate is not yet known. We have also detected an inhibitor of ADR activity in the cytoplasm of resting lymphocytes. This is a heat-stable protein of approximately 60,000 Da. In addition to suppressing the interaction of ADR with quiescent nuclei, the inhibitor can suppress DNA synthetic activity of replicative nuclei isolated from mitogen-activated lymphocytes. Interestingly, these preparations had little or no activity on replicative nuclei derived from several neoplastic cell lines. The resistance of tumor cell nuclei to spontaneously occurring cytoplasmic inhibitory factors such as the one described here may provide one explanation for the loss of growth control in neoplastic cells.

  3. Traffic jams II: an update of diseases of intracellular transport.

    Science.gov (United States)

    Aridor, Meir; Hannan, Lisa A

    2002-11-01

    As more details emerge on the mechanisms that mediate and control intracellular transport, the molecular basis for variety of human diseases has been revealed. In turn, disease pathology and physiology shed light on the intricate controls that regulate intracellular transport to assure proper cellular and tissue function and homeostasis. We previously listed a number of diseases that are the result of defects in intracellular transport, or cause defects in intracellular transport. (Aridor M, Hannan LA. Traffic Jam: A compendium of human diseases that affect intracellular transport processes. Traffic 2000; 1: 836-851). This Toolbox updates the previous list to include additional disorders that were recently identified to be related to intracellular trafficking. In the time since we have published our first list there have been significant advances in understanding of the molecular basis of these defects. Such advances will pave the way to future effective therapeutics.

  4. The measurement of constitutive and facultative skin pigmentation and estimation of sun exposure in Caucasians with basal cell carcinoma and cutaneous malignant melanoma

    DEFF Research Database (Denmark)

    Lock-Andersen, J; Drzewiecki, K T; Wulf, H C

    1998-01-01

    In two identical and simultaneously performed case-control studies of basal cell carcinoma (BCC) and cutaneous malignant melanoma (CMM) with age-matched, sex-matched and residence-matched controls, skin pigmentation was measured objectively by skin reflectance spectroscopy in 145 BCC patients...... by all subjects. There were no statistically significant differences in constitutive skin pigmentation at the buttocks between BCC patients and controls (P = 0.96) or between CMM patients and controls (P = 0.13). Facultative skin pigmentation in ultraviolet-exposed sites was not significantly different...... between BCC patients and controls except that women patients had higher pigmentation at the lateral side of the upper arm. For CMM, men patients had higher pigmentation at the lateral side of the upper arm. Self-estimations of sun exposure did not show differences between patients and controls...

  5. Isolation and Identification of Psychrotrophic Bacteria in Normal Temperature Soils and Their Properties of Cold-Adapted Enzymes Production%常温土壤中耐冷茵的分离、鉴定及产酶分析

    Institute of Scientific and Technical Information of China (English)

    易浪波; 舒琅; 唐云涛; 朱越; 杜次; 彭清忠

    2012-01-01

    Objective: To screen cold-adapted microorganisms from normal temperature soils,and identify the isolated strains preliminarily and assay their cold-active enzyme production. Methods: Cold-adapted microorganisms were isolated from normal temperature soils at the campus of Jishou university by enrichment culture at 2℃. The isolated strains were identified by bacterial morphological,physiological and biochemical characteristics,and using phylogenetic relationships based on 16S rRNA gene sequences. The cold-adapted enzyme production of these bacteria was analyzed by substrate hydrolysis experiments. Results: Six strains of psychrotrophic bacteria,named afterJSBP-1~JSBP-6,were isolated from the normal temperature soil samples,and identified as Pseudomonas sp.,Jan-thinobacterium sp. And Arthrobacter sp. Respectively by morphological,physiological and biochemical characteristics,and the phylogenetic analysis of 16S rRNA gene. JSBP-1 had the capability of producing more cold-adapted protease,JSBP-2 and JSBP-6 had the capability of producing more cold-adapted amylase at 5℃ and \\5℃ respectively,and JSBP-5 could produce more cold-active lipase only at 5℃. Conclusion: There are a certain amount of cold-adapted microorganisms in normal temperature soils,of which Pseudomonas sp. Is one of predominant populations. These microorganisms are potential resources of producing cold-adapted enzymes.%目的:从常温土壤中筛选冷适应微生物,并进行初步鉴定和产低温酶分析.方法:采集吉首大学校园内土壤样品,通过低温富集培养筛选冷适应微生物;通过形态观察、生理生化特性检测和基于16S rRNA基因序列的系统发育分析,对分离的菌株进行初步鉴定;利用平板筛选法检测其产低温酶特性.结果:分离获得6株耐冷细菌JSBP-1~JSBP-6,初步鉴定其分属假单胞菌属(Pseudomonas)、紫色杆菌属(Janthinobacterium)和节杆菌属(Arthrobacter);在5℃和15C培养条件下,菌株JSBP-1

  6. 一株耐冷反硝化菌的分离鉴定及其反硝化特性%Identification and Denitrification Characteristics of a Strain of Psychrotrophic Denitrifying Bacteria

    Institute of Scientific and Technical Information of China (English)

    李军; 闫爽; 邓娴; 王立军; 张雅南; 杨宏宇

    2012-01-01

    A psychrotrophic denitrifying bacteria (designated Y2)with high efficiency degradation of nitrate nitrogen was isolated from the submerged mud in winter water at 151 which was used to treat high-nitrate wastewater at low temperature. According to the results of morphological observation,physiological and biochemical test .sequence analyses of the 16S rDNA, strain Y2 was identified as Brevibacterium. The NO," -N removal rate of Y2 can reach 51. 6% in denitrification medium at 151 in 3 days,and 99. 3% in 7 days. The optimum carbon source and the optimum nitrogen source of Y2 were sodium acetate and potassium nitrate. The suitable molar ratio of carbon/nitrogen for denitrification of the bacteria was 3= 1,the optimum pH value was 7.5, and the optimum temperature was 25 t. It showed Y2 was suitable for the nitrate elimination of ni-trates at low temperature.%目的 筛选分离出一株具有高效降解硝态氮的耐冷反硝化菌,以提高低温下硝酸盐废水处理的生物反硝化速率.方法 在15℃条件下从冬季淹水泥中分离纯化出一株耐冷反硝化细菌Y2,通过对该菌株的形态观察、生理生化试验以及16S rRNA序列分析,对菌株进行鉴定;通过批试验研究不同因素对菌株单位生物量脱氮率的影响.结果 初步确定Y2菌株为黄色短杆菌(Brevibacterium).该菌株能在15℃下在反硝化培养基中3d内对NO3- -N的去除率达到51.6%,7d能达到99.3%,该菌的最适碳源为乙酸钠、最适氮源为硝酸钾,最适碳氮比为3:1、最适pH值为7.5、最适温度25℃.结论 菌株Y2在低温下表现出良好的反硝化性能,NO3- -N质量浓度达300 mg/L时,去除率可达99.8%,适用于低温硝酸盐废水处理.

  7. Biochemical characterization of a bifunctional acetaldehyde-alcohol dehydrogenase purified from a facultative anaerobic bacterium Citrobacter sp. S-77.

    Science.gov (United States)

    Tsuji, Kohsei; Yoon, Ki-Seok; Ogo, Seiji

    2016-03-01

    Acetaldehyde-alcohol dehydrogenase (ADHE) is a bifunctional enzyme consisting of two domains of an N-terminal acetaldehyde dehydrogenase (ALDH) and a C-terminal alcohol dehydrogenase (ADH). The enzyme is known to be important in the cellular alcohol metabolism. However, the role of coenzyme A-acylating ADHE responsible for ethanol production from acetyl-CoA remains uncertain. Here, we present the purification and biochemical characterization of an ADHE from Citrobacter sp. S-77 (ADHE(S77)). Interestingly, the ADHE(S77) was unable to be solubilized from membrane with detergents either 1% Triton X-100 or 1% Sulfobetaine 3-12. However, the enzyme was easily dissociated from membrane by high-salt buffers containing either 1.0 M NaCl or (NH(4))(2)SO(4) without detergents. The molecular weight of a native protein was estimated as approximately 400 kDa, consisting of four identical subunits of 96.3 kDa. Based on the specific activity and kinetic analysis, the ADHES77 tended to have catalytic reaction towards acetaldehyde elimination rather than acetaldehyde formation. Our experimental observation suggests that the ADHES77 may play a pivotal role in modulating intracellular acetaldehyde concentration. Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  8. Extracellular Matrix Stiffness and Architecture Govern Intracellular Rheology in Cancer

    OpenAIRE

    Baker, Erin L.; Bonnecaze, Roger T.; Zaman, Muhammad H.

    2009-01-01

    Little is known about the complex interplay between the extracellular mechanical environment and the mechanical properties that characterize the dynamic intracellular environment. To elucidate this relationship in cancer, we probe the intracellular environment using particle-tracking microrheology. In three-dimensional (3D) matrices, intracellular effective creep compliance of prostate cancer cells is shown to increase with increasing extracellular matrix (ECM) stiffness, whereas modulating E...

  9. Decrease of U(VI immobilization capability of the facultative anaerobic strain Paenibacillus sp. JG-TB8 under anoxic conditions due to strongly reduced phosphatase activity.

    Directory of Open Access Journals (Sweden)

    Thomas Reitz

    Full Text Available Interactions of a facultative anaerobic bacterial isolate named Paenibacillus sp. JG-TB8 with U(VI were studied under oxic and anoxic conditions in order to assess the influence of the oxygen-dependent cell metabolism on microbial uranium mobilization and immobilization. We demonstrated that aerobically and anaerobically grown cells of Paenibacillus sp. JG-TB8 accumulate uranium from aqueous solutions under acidic conditions (pH 2 to 6, under oxic and anoxic conditions. A combination of spectroscopic and microscopic methods revealed that the speciation of U(VI associated with the cells of the strain depend on the pH as well as on the aeration conditions. At pH 2 and pH 3, uranium was exclusively bound by organic phosphate groups provided by cellular components, independently on the aeration conditions. At higher pH values, a part (pH 4.5 or the total amount (pH 6 of the dissolved uranium was precipitated under oxic conditions in a meta-autunite-like uranyl phosphate mineral phase without supplying an additional organic phosphate substrate. In contrast to that, under anoxic conditions no mineral formation was observed at pH 4.5 and pH 6, which was clearly assigned to decreased orthophosphate release by the cells. This in turn was caused by a suppression of the indigenous phosphatase activity of the strain. The results demonstrate that changes in the metabolism of facultative anaerobic microorganisms caused by the presence or absence of oxygen can decisively influence U(VI biomineralization.

  10. On the nature of facultative and constitutive CAM: environmental and developmental control of CAM expression during early growth of Clusia, Kalanchöe, and Opuntia.

    Science.gov (United States)

    Winter, Klaus; Garcia, Milton; Holtum, Joseph A M

    2008-01-01

    The capacity to induce crassulacean acid metabolism developmentally (constitutive CAM) and to up-regulate CAM expression in response to drought stress (facultative CAM) was studied in whole shoots of seven species by measuring net CO(2) gas exchange for up to 120 day-night cycles during early growth. In Clusia rosea, CAM was largely induced developmentally. Well-watered seedlings began their life cycle as C(3) plants and developed net dark CO(2) fixation indicative of CAM after the initiation of the fourth leaf pair following the cotyledons. Thereafter, CAM activity increased progressively and drought stress led to only small additional, reversible increases in dark CO(2) fixation. In contrast, CAM expression was overwhelmingly under environmental control in seedlings and mature plants of Clusia pratensis. C(3)-type CO(2) exchange was maintained under well-watered conditions, but upon drought stress, CO(2) exchange shifted, in a fully reversible manner, to a CAM-type pattern. Clusia minor showed CO(2) exchange reponses intermediate to those of C. rosea and C. pratensis. Clusia cretosa operated in the C(3) mode at all times. Notably, reversible stress-induced increases of dark CO(2) fixation were also observed during the developmental progression to pronounced CAM in young Kalanchoë daigremontiana and Kalanchoë pinnata, two species considered constitutive CAM species. Drought-induced up-regulation of CAM was even detected in young cladodes of a cactus, Opuntia ficus-indica, an archetypal constitutive CAM species. Evidently, the defining characteristics of constitutive and facultative CAM are shared, to variable degrees, by all CAM species.

  11. La stratégie de l'OMS « Vers l'Unité Pour la Santé » et la responsabilité sociale des facultés de médecine

    National Research Council Canada - National Science Library

    C. Boelen

    2003-01-01

    ... : les décideurs politiques, les gestionnaires des services de santé, les professions de santé, les institutions universitaires, dont les facultés de médecine, et la société civile. Cette unité...

  12. Strategies of Intracellular Pathogens for Obtaining Iron from the Environment

    Directory of Open Access Journals (Sweden)

    Nidia Leon-Sicairos

    2015-01-01

    Full Text Available Most microorganisms are destroyed by the host tissues through processes that usually involve phagocytosis and lysosomal disruption. However, some organisms, called intracellular pathogens, are capable of avoiding destruction by growing inside macrophages or other cells. During infection with intracellular pathogenic microorganisms, the element iron is required by both the host cell and the pathogen that inhabits the host cell. This minireview focuses on how intracellular pathogens use multiple strategies to obtain nutritional iron from the intracellular environment in order to use this element for replication. Additionally, the implications of these mechanisms for iron acquisition in the pathogen-host relationship are discussed.

  13. Intracellular signaling by diffusion: can waves of hydrogen peroxide transmit intracellular information in plant cells?

    DEFF Research Database (Denmark)

    Vestergaard, Christian L.; Flyvbjerg, Henrik; Møller, Ian Max

    2012-01-01

    Amplitude- and frequency-modulated waves of Ca(2+) ions transmit information inside cells. Reactive Oxygen Species (ROS), specifically hydrogen peroxide, have been proposed to have a similar role in plant cells. We consider the feasibility of such an intracellular communication system in view...... of the physical and biochemical conditions in plant cells. As model system, we use a H(2)O(2) signal originating at the plasma membrane (PM) and spreading through the cytosol. We consider two maximally simple types of signals, isolated pulses and harmonic oscillations. First we consider the basic limits...... which diffusion-mediated signaling is possible. We show that purely diffusive transmission of intracellular information by H(2)O(2) over a distance of 1 μm (typical distance between organelles, which may function as relay stations) is possible at frequencies well above 1 Hz, which is the highest...

  14. Intracellular signaling by diffusion: can waves of hydrogen peroxide transmit intracellular information in plant cells?

    DEFF Research Database (Denmark)

    Vestergaard, Christian Lyngby; Flyvbjerg, Henrik; Møller, Ian Max

    2012-01-01

    Amplitude- and frequency-modulated waves of Ca2+ ions transmit information inside cells. Reactive Oxygen Species (ROS), specifically hydrogen peroxide, have been proposed to have a similar role in plant cells. We consider the feasibility of such an intracellular communication system in view...... of the physical and biochemical conditions in plant cells. As model system, we use a H2O2 signal originating at the plasma membrane (PM) and spreading through the cytosol. We consider two maximally simple types of signals, isolated pulses and harmonic oscillations. First we consider the basic limits...... diffusion-mediated signaling is possible. We show that purely diffusive transmission of intracellular information by H2O2 over a distance of 1 μm (typical distance between organelles, which may function as relay stations) is possible at frequencies well above 1 Hz, which is the highest frequency observed...

  15. Duodenal Intracellular Bicarbonate and the 'CF Paradox'

    Directory of Open Access Journals (Sweden)

    Kaunitz JD

    2001-07-01

    Full Text Available HCO(3(- secretion, which is believed to neutralize acid within the mucus gel, is the most studied duodenal defense mechanism. In general, HCO(3(- secretion rate and mucosal injury susceptibility correlate closely. Recent studies suggest that luminal acid can lower intracellular pH (pH(i of duodenal epithelial cells and that HCO(3(- secretion is unchanged during acid stress. Furthermore, peptic ulcers are rare in cystic fibrosis (CF, although, with impaired HCO(3(- secretion, increased ulcer prevalence is predicted, giving rise to the 'CF Paradox'. We thus tested the hypothesis that duodenal epithelial cell protection occurs as the result of pH(i regulation rather than by neutralization of acid by HCO(3(- in the pre-epithelial mucus. Cellular acidification during luminal acid perfusion, and unchanged HCO(3(- secretion during acid stress are inconsistent with pre-epithelial acid neutralization by secreted HCO(3(-. Furthermore, inhibition of HCO(3(- secretion by 5-nitro-2-(3-phenylpropylamino benzoic acid (NPPB despite preservation of pH(i and protection from acid-induced injury further question the pre-epithelial acid neutralization hypothesis. This decoupling of HCO(3(- secretion and injury susceptibility by NPPB (and possibly by CF further suggest that cellular buffering, rather than HCO(3(- exit into the mucus, is of primary importance for duodenal mucosal protection, and may account for the lack of peptic ulceration in CF patients.

  16. Intracellular Signals of T Cell Costimulation

    Institute of Scientific and Technical Information of China (English)

    Jianxun Song; Fengyang Tylan Lei; Xiaofang Xiong; Rizwanul Haque

    2008-01-01

    Ligation of T cell receptor (TCR) alone is insufficient to induce full activation of T lymphocytes. Additional ligand-receptor interactions (costimulation) on antigen presenting cells (APCs) and T cells are required. T cell costimulation has been shown to be essential for eliciting efficient T cell responses, involving all phases during T cell development. However, the mechanisms by which costimulation affects the function of T cells still need to be elucidated. In recent years, advances have been made in studies of costimulation as potential therapies in cancer, infectious disease as well as autoimmune disease. In this review, we discussed intracellular costimulation signals that regulate T cell proliferation, cell cycle progression, cytokine production, survival, and memory development. In general, the pathway of phosphoinositide-3 kinase (PBK)/protein kinase B (PKB, also known as Akt)/nuclear factor κB (NF-κB) might be central to many costimulatory effects. Through these pathways, costimulation controls T-cell expansion and proliferation by maintenance of survivin and aurora B expression, and sustains long-term T-cell survival and memory development by regulating the expression of bci-2 family members. Cellular & Molecular Immunology.2008;5(4):239-247.

  17. On the Computing Potential of Intracellular Vesicles.

    Science.gov (United States)

    Mayne, Richard; Adamatzky, Andrew

    2015-01-01

    Collision-based computing (CBC) is a form of unconventional computing in which travelling localisations represent data and conditional routing of signals determines the output state; collisions between localisations represent logical operations. We investigated patterns of Ca2+-containing vesicle distribution within a live organism, slime mould Physarum polycephalum, with confocal microscopy and observed them colliding regularly. Vesicles travel down cytoskeletal 'circuitry' and their collisions may result in reflection, fusion or annihilation. We demonstrate through experimental observations that naturally-occurring vesicle dynamics may be characterised as a computationally-universal set of Boolean logical operations and present a 'vesicle modification' of the archetypal CBC 'billiard ball model' of computation. We proceed to discuss the viability of intracellular vesicles as an unconventional computing substrate in which we delineate practical considerations for reliable vesicle 'programming' in both in vivo and in vitro vesicle computing architectures and present optimised designs for both single logical gates and combinatorial logic circuits based on cytoskeletal network conformations. The results presented here demonstrate the first characterisation of intracelluar phenomena as collision-based computing and hence the viability of biological substrates for computing.

  18. On the Computing Potential of Intracellular Vesicles.

    Directory of Open Access Journals (Sweden)

    Richard Mayne

    Full Text Available Collision-based computing (CBC is a form of unconventional computing in which travelling localisations represent data and conditional routing of signals determines the output state; collisions between localisations represent logical operations. We investigated patterns of Ca2+-containing vesicle distribution within a live organism, slime mould Physarum polycephalum, with confocal microscopy and observed them colliding regularly. Vesicles travel down cytoskeletal 'circuitry' and their collisions may result in reflection, fusion or annihilation. We demonstrate through experimental observations that naturally-occurring vesicle dynamics may be characterised as a computationally-universal set of Boolean logical operations and present a 'vesicle modification' of the archetypal CBC 'billiard ball model' of computation. We proceed to discuss the viability of intracellular vesicles as an unconventional computing substrate in which we delineate practical considerations for reliable vesicle 'programming' in both in vivo and in vitro vesicle computing architectures and present optimised designs for both single logical gates and combinatorial logic circuits based on cytoskeletal network conformations. The results presented here demonstrate the first characterisation of intracelluar phenomena as collision-based computing and hence the viability of biological substrates for computing.

  19. Imaging and controlling intracellular reactions: Lysosome transport as a function of diameter and the intracellular synthesis of conducting polymers

    Science.gov (United States)

    Payne, Christine

    2014-03-01

    Eukaryotic cells are the ultimate complex environment with intracellular chemical reactions regulated by the local cellular environment. For example, reactants are sequestered into specific organelles to control local concentration and pH, motor proteins transport reactants within the cell, and intracellular vesicles undergo fusion to bring reactants together. Current research in the Payne Lab in the School of Chemistry and Biochemistry at Georgia Tech is aimed at understanding and utilizing this complex environment to control intracellular chemical reactions. This will be illustrated using two examples, intracellular transport as a function of organelle diameter and the intracellular synthesis of conducting polymers. Using single particle tracking fluorescence microscopy, we measured the intracellular transport of lysosomes, membrane-bound organelles, as a function of diameter as they underwent transport in living cells. Both ATP-dependent active transport and diffusion were examined. As expected, diffusion scales with the diameter of the lysosome. However, active transport is unaffected suggesting that motor proteins are insensitive to cytosolic drag. In a second example, we utilize intracellular complexity, specifically the distinct micro-environments of different organelles, to carry out chemical reactions. We show that catalase, found in the peroxisomes of cells, can be used to catalyze the polymerization of the conducting polymer PEDOT:PSS. More importantly, we have found that a range of iron-containing biomolecules are suitable catalysts with different iron-containing biomolecules leading to different polymer properties. These experiments illustrate the advantage of intracellular complexity for the synthesis of novel materials.

  20. Methane oxidation with in situ enhanced facultative bacteria from aged-refuse%矿化垃圾中兼性营养菌原位强化甲烷氧化

    Institute of Scientific and Technical Information of China (English)

    赵天涛; 张云茹; 张丽杰; 全学军; 彭绪亚

    2012-01-01

    引言好氧甲烷氧化菌在新陈代谢上具有独一无二的特性:它们能够利用甲烷和其他一碳化合物作为唯一碳源和能源.这类微生物最典型的特点是利用甲烷单加氧酶( MMO,methane monooxygenase)催化甲烷氧化为甲醇[1].长时间来,所有的甲烷氧化菌都被认为是专一营养的,即它们无法利用含有碳碳键的化合物生长.%Facultative methanotrophs can utilize methane as well as multi-carbon compounds, including organic acids and carbohydrates. Facultative methanotrophs from aged-refuse were enhanced in situ to overcome the limitations of methane oxidation by existing landfill covers. Methanotrophs from oligotrophic aged-refuse had a better environmental tolerance by analysis of scanning electron microscope (SEM). These bacteria could enrich quickly in the presence of carbon source. However, the methane-oxidation capability could not be improved if only adding carbohydrates or nitrate mineral salts (NMS) medium alone. Compound acclimation of facultative methanotrophs were carried out by NMS medium and glucose/ starch. After a delay period of 7-9 d, methane consumption came into a logarithmic growth period, which indicated that facultative methanotrophs had strong biological activity and high substrate competitive advantage. Metabolic pathways of facultative methanotrophs were modified by adding NMS medium, and methane could be utilized despite the presence of other carbon sources. The activity of facultative methanotrophs was enhanced by low concentration of chloroform. Oxidation rate of methane reached 0. 114 ml·d-1·g-1 as the concentration of chloroform was 50 mg·L-1. The problems about the engineering application of obligate methanotrophs were overcome due to the discovery of the new method, and the results would have important implication for understanding the methane-oxidizing bacteria and the factors controlling methane fluxes in the environment.

  1. Antimicrobial susceptibilities of aerobic and facultative gram-negative bacilli isolated from Chinese patients with urinary tract infections between 2010 and 2014.

    Science.gov (United States)

    Yang, Qiwen; Zhang, Hui; Wang, Yao; Xu, Zhipeng; Zhang, Ge; Chen, Xinxin; Xu, Yingchun; Cao, Bin; Kong, Haishen; Ni, Yuxing; Yu, Yunsong; Sun, Ziyong; Hu, Bijie; Huang, Wenxiang; Wang, Yong; Wu, Anhua; Feng, Xianju; Liao, Kang; Luo, Yanping; Hu, Zhidong; Chu, Yunzhuo; Lu, Juan; Su, Jianrong; Gui, Bingdong; Duan, Qiong; Zhang, Shufang; Shao, Haifeng; Badal, Robert E

    2017-03-06

    The objective of this study was to investigate the distribution and susceptibility of aerobic and facultative Gram-negative bacilli isolated from Chinese patients with UTIs collected within 48 h (community acquired, CA) or after 48 h (hospital acquired, HA) of hospital admission. From 2010 to 2014, the minimum inhibitory concentrations (MICs) of 12 antibiotics for 4,332 aerobic and facultative Gram-negative bacilli, sampled in 21 hospitals in 16 cities, were determined by the broth microdilution method. Enterobacteriaceae composed 88.5% of the total isolates, with Escherichia coli (E. coli) (63.2%) the most commonly isolated species, followed by Klebsiella pneumoniae (K. pneumoniae) (12.2%). Non-Enterobacteriaceae accounted for only 11.5% of all isolates and included mainly Pseudomonas aeruginosa (P. aeruginosa) (6.9%) and Acinetobacter baumannii (A. baumannii) (3.3%). Among the antimicrobial agents tested, the susceptibility rates of E.coli to the two carbapenems, ertapenem and imipenem as well as amikacin and piperacillin-tazobactam ranged from 92.5 to 98.7%. Against K. pneumonia, the most potent antibiotics were imipenem (92.6% susceptibility), amikacin (89.2% susceptibility) and ertapenem (87.9% susceptibility). Although non-Enterobacteriaceae did not show high susceptibilities to the 12 common antibiotics, amikacin exhibited the highest in vitro activity against P. aeruginosa over the 5-year study period, followed by piperacillin-tazobactam, imipenem, ceftazidime, cefepime, ciprofloxacin, and levofloxacin. The Extended Spectrum Beta-Lactamase (ESBL) rates decreased slowly during the 5 years in E. coli from 68.6% in 2010 to 59.1% in 2014, in K. pneumoniae from 59.7 to 49.2%, and in Proteus mirabilis (P. mirabilis) from 40.0 to 26.1%. However, the ESBL rates were different in 5 regions of China (Northeast, North, East, South and Middle-China). E. coli and K. pneumonia were the major pathogens causing UTIs and carbapenems and amikacin retained the highest

  2. Meta-analysis of effects of inoculation with homofermentative and facultative heterofermentative lactic acid bacteria on silage fermentation, aerobic stability, and the performance of dairy cows.

    Science.gov (United States)

    Oliveira, André S; Weinberg, Zwi G; Ogunade, Ibukun M; Cervantes, Andres A P; Arriola, Kathy G; Jiang, Yun; Kim, Donghyeon; Li, Xujiao; Gonçalves, Mariana C M; Vyas, Diwakar; Adesogan, Adegbola T

    2017-06-01

    Forages are usually inoculated with homofermentative and facultative heterofermentative lactic acid bacteria (LAB) to enhance lactic acid fermentation of forages, but effects of such inoculants on silage quality and the performance of dairy cows are unclear. Therefore, we conducted a meta-analysis to examine the effects of LAB inoculation on silage quality and preservation and the performance of dairy cows. A second objective was to examine the factors affecting the response to silage inoculation with LAB. The studies that met the selection criteria included 130 articles that examined the effects of LAB inoculation on silage quality and 31 articles that investigated dairy cow performance responses. The magnitude of the effect (effect size) was evaluated using raw mean differences (RMD) between inoculated and uninoculated treatments. Heterogeneity was explored by meta-regression and subgroup analysis using forage type, LAB species, LAB application rate, and silo scale (laboratory or farm-scale) as covariates for the silage quality response and forage type, LAB species, diet type [total mixed ration (TMR) or non-TMR], and the level of milk yield of the control cows as covariates for the performance responses. Inoculation with LAB (≥10(5) cfu/g as fed) markedly increased silage fermentation and dry matter recovery in temperate and tropical grasses, alfalfa, and other legumes. However, inoculation did not improve the fermentation of corn, sorghum, or sugarcane silages. Inoculation with LAB reduced clostridia and mold growth, butyric acid production, and ammonia-nitrogen in all silages, but it had no effect on aerobic stability. Silage inoculation (≥10(5) cfu/g as fed) increased milk yield and the response had low heterogeneity. However, inoculation had no effect on diet digestibility and feed efficiency. Inoculation with LAB improved the fermentation of grass and legume silages and the performance of dairy cows but did not affect the fermentation of corn, sorghum

  3. Targeting intracellular Staphylococcus aureus to lower recurrence of orthopaedic infection.

    Science.gov (United States)

    Dusane, Devendra H; Kyrouac, Douglas; Petersen, Iris; Bushrow, Luke; Calhoun, Jason H; Granger, Jeffrey F; Phieffer, Laura S; Stoodley, Paul

    2017-09-08

    Staphylococcus aureus is often found in orthopaedic infections and may be protected from commonly prescribed antibiotics by forming biofilms or growing intracellularly within osteoblasts. To investigate the effect of non-antibiotic compounds in conjunction with antibiotics to clear intracellular and biofilm forming S. aureus causing osteomyelitis. SAOS-2 osteoblast-like cell lines were infected with S. aureus BB1279. Antibiotics (vancomycin, VAN; and dicloxacillin, DICLOX), bacterial efflux pump inhibitors (piperine, PIP; carbonyl cyanide m-chlorophenyl hydrazone, CCCP) and bone morphogenetic protein (BMP-2) were evaluated individually and in combination to kill intracellular bacteria. We present direct evidence that after gentamicin killed extracellular planktonic bacteria and antibiotics had been stopped, seeding from the infected osteoblasts grew as biofilms. VAN was ineffective in treating the intracellular bacteria even at 10x MIC; however in presence of PIP or CCCP the intracellular S. aureus was significantly reduced. Bacterial efflux pump inhibitors (PIP and CCCP) were effective in enhancing permeability of antibiotics within the osteoblasts and facilitated killing of intracellular S. aureus. Confocal laser scanning microscopy (CLSM) showed increased uptake of propidium iodide within osteoblasts in presence of PIP and CCCP. BMP-2 had no effect on growth of S. aureus either alone or in combination with antibiotics. Combined application of antibiotics and natural agents could help in the treatment of osteoblast infected intracellular bacteria and biofilms associated with osteomyelitis. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  4. Extracellular matrix stiffness and architecture govern intracellular rheology in cancer.

    Science.gov (United States)

    Baker, Erin L; Bonnecaze, Roger T; Zaman, Muhammad H

    2009-08-19

    Little is known about the complex interplay between the extracellular mechanical environment and the mechanical properties that characterize the dynamic intracellular environment. To elucidate this relationship in cancer, we probe the intracellular environment using particle-tracking microrheology. In three-dimensional (3D) matrices, intracellular effective creep compliance of prostate cancer cells is shown to increase with increasing extracellular matrix (ECM) stiffness, whereas modulating ECM stiffness does not significantly affect the intracellular mechanical state when cells are attached to two-dimensional (2D) matrices. Switching from 2D to 3D matrices induces an order-of-magnitude shift in intracellular effective creep compliance and apparent elastic modulus. However, for a given matrix stiffness, partial blocking of beta1 integrins mitigates the shift in intracellular mechanical state that is invoked by switching from a 2D to 3D matrix architecture. This finding suggests that the increased cell-matrix engagement inherent to a 3D matrix architecture may contribute to differences observed in viscoelastic properties between cells attached to 2D matrices and cells embedded within 3D matrices. In total, our observations show that ECM stiffness and architecture can strongly influence the intracellular mechanical state of cancer cells.

  5. New perspective in the assessment of total intracellular magnesium

    Directory of Open Access Journals (Sweden)

    Azzurra Sargenti

    2014-01-01

    Full Text Available Magnesium (Mg is essential for biological processes, but its cellular homeostasis has not been thoroughly elucidated, mainly because of the inadequacy of the available techniques to map intracellular Mg distribution. Recently, particular interest has been raised by a new family of fluorescent probes, diaza-18-crown-hydroxyquinoline (DCHQ, that shows remarkably high affinity and specificity for Mg, thus permitting the detection of the total intracellular Mg. The data obtained by fluori- metric and cytofluorimetric assays performed with DCHQ5 are in good agreement with atomic absorption spectroscopy, confirming that DCHQ5 probe allows both qualitative and quantitative determination of total intracellular Mg.

  6. Human neutrophils dump Candida glabrata after intracellular killing.

    Science.gov (United States)

    Essig, Fabian; Hünniger, Kerstin; Dietrich, Stefanie; Figge, Marc Thilo; Kurzai, Oliver

    2015-11-01

    Interaction between fungal pathogens and human phagocytes can lead to remarkably variable outcomes, ranging from intracellular killing to prolonged survival and replication of the pathogen in the host cell. Using live cell imaging we observed primary human neutrophils that release phagocytosed Candida glabrata yeast cells after intracellular killing. This process, for which we propose the name "dumping", adds a new outcome to phagocyte-fungus interaction which may be of potential immunological importance as it allows professional antigen presenting cells to take up and process neutrophil-inactivated pathogens that in their viable state are able to evade intracellular degradation in these cells.

  7. Intracellular Acidosis Enhances the Excitability of Working Muscle

    Science.gov (United States)

    Pedersen, Thomas H.; Nielsen, Ole B.; Lamb, Graham D.; Stephenson, D. George

    2004-08-01

    Intracellular acidification of skeletal muscles is commonly thought to contribute to muscle fatigue. However, intracellular acidosis also acts to preserve muscle excitability when muscles become depolarized, which occurs with working muscles. Here, we show that this process may be mediated by decreased chloride permeability, which enables action potentials to still be propagated along the internal network of tubules in a muscle fiber (the T system) despite muscle depolarization. These results implicate chloride ion channels in muscle function and emphasize that intracellular acidosis of muscle has protective effects during muscle fatigue.

  8. Intracellular acidosis enhances the excitability of working muscle.

    Science.gov (United States)

    Pedersen, Thomas H; Nielsen, Ole B; Lamb, Graham D; Stephenson, D George

    2004-08-20

    Intracellular acidification of skeletal muscles is commonly thought to contribute to muscle fatigue. However, intracellular acidosis also acts to preserve muscle excitability when muscles become depolarized, which occurs with working muscles. Here, we show that this process may be mediated by decreased chloride permeability, which enables action potentials to still be propagated along the internal network of tubules in a muscle fiber (the T system) despite muscle depolarization. These results implicate chloride ion channels in muscle function and emphasize that intracellular acidosis of muscle has protective effects during muscle fatigue.

  9. Genome sequence of Phaeobacter daeponensis type strain (DSM 23529(T)), a facultatively anaerobic bacterium isolated from marine sediment, and emendation of Phaeobacter daeponensis.

    Science.gov (United States)

    Dogs, Marco; Teshima, Hazuki; Petersen, Jörn; Fiebig, Anne; Chertkov, Olga; Dalingault, Hajnalka; Chen, Amy; Pati, Amrita; Goodwin, Lynne A; Chain, Patrick; Detter, John C; Ivanova, Natalia; Lapidus, Alla; Rohde, Manfred; Gronow, Sabine; Kyrpides, Nikos C; Woyke, Tanja; Simon, Meinhard; Göker, Markus; Klenk, Hans-Peter; Brinkhoff, Thorsten

    2013-10-16

    TF-218(T) is the type strain of the species Phaeobacter daeponensis Yoon et al. 2007, a facultatively anaerobic Phaeobacter species isolated from tidal flats. Here we describe the draft genome sequence and annotation of this bacterium together with previously unreported aspects of its phenotype. We analyzed the genome for genes involved in secondary metabolite production and its anaerobic lifestyle, which have also been described for its closest relative Phaeobacter caeruleus. The 4,642,596 bp long genome of strain TF-218(T) contains 4,310 protein-coding genes and 78 RNA genes including four rRNA operons and consists of five replicons: one chromosome and four extrachromosomal elements with sizes of 276 kb, 174 kb, 117 kb and 90 kb. Genome analysis showed that TF-218(T) possesses all of the genes for indigoidine biosynthesis, and on specific media the strain showed a blue pigmentation. We also found genes for dissimilatory nitrate reduction, gene-transfer agents, NRPS/ PKS genes and signaling systems homologous to the LuxR/I system.

  10. Influence of habitat and climate variables on arbuscular mycorrhizal fungus community distribution, as revealed by a case study of facultative plant epiphytism under semiarid conditions.

    Science.gov (United States)

    Torrecillas, E; Torres, P; Alguacil, M M; Querejeta, J I; Roldán, A

    2013-12-01

    In semiarid Mediterranean ecosystems, epiphytic plant species are practically absent, and only some species of palm trees can support epiphytes growing in their lower crown area, such as Phoenix dactylifera L. (date palm). In this study, we focused on Sonchus tenerrimus L. plants growing as facultative epiphytes in P. dactylifera and its terrestrial forms growing in adjacent soils. Our aim was to determine the possible presence of arbuscular mycorrhizal fungi (AMF) in these peculiar habitats and to relate AMF communities with climatic variations. We investigated the AMF community composition of epiphytic and terrestrial S. tenerrimus plants along a temperature and precipitation gradient across 12 localities. Epiphytic roots were colonized by AMF, as determined by microscopic observation; all of the epiphytic and terrestrial samples analyzed showed AMF sequences from taxa belonging to the phylum Glomeromycota, which were grouped in 30 AMF operational taxonomic units. The AMF community composition was clearly different between epiphytic and terrestrial root samples, and this could be attributable to dispersal constraints and/or the contrasting environmental and ecophysiological conditions prevailing in each habitat. Across sites, the richness and diversity of terrestrial AMF communities was positively correlated with rainfall amount during the most recent growing season. In contrast, there was no significant correlation between climate variables and AMF richness and diversity for epiphytic AMF communities, which suggests that the composition of AMF communities in epiphytic habitats appears to be largely determined by the availability and dispersion of fungal propagules from adjacent terrestrial habitats.

  11. Does a facultative precocious life cycle predispose the marine trematode Proctoeces cf. lintoni to inbreeding and genetic differentiation among host species?

    Science.gov (United States)

    Valdivia, I M; Criscione, C D; Cárdenas, L; Durán, C P; Oliva, M E

    2014-03-01

    Intraspecific variability in parasite life cycle complexity (number of hosts and species of hosts in the life cycle) may have an impact how parasite genetic variation is partitioned among individual parasites, host individuals or host species within a given area. Among digenean trematodes, a three-host life cycle is common. However, a few species are precocious and may reach sexual maturity in what is typically regarded as the second intermediate host. The objective of this study was to determine whether a precocious life cycle predisposes digeneans to possible inbreeding or genetic subdivision among host species. As a study system, we used the digenean Proctoeces cf. lintoni whose metacercariae precociously mature (facultative) without a cyst wall in the gonads of multiple sympatric species of keyhole limpets (Fissurella spp.), typically regarded as the second intermediate hosts. Genotyped parasites were collected from four species of limpets and the clingfish Sicyases sanguineus, the third and final host where sexual maturity occurs. We found very high microsatellite diversity, Hardy-Weinberg equilibrium over all genotyped individuals, and little to no genetic structuring among parasites collected from the different host species. The fact that metacercariae do not encyst in the keyhole limpets, coupled with the high mixing potential of an aquatic environment, likely promote panmixia in local populations of P. cf. lintoni.

  12. Genome sequence of Phaeobacter daeponensis type strain (DSM 23529T), a facultatively anaerobic bacterium isolated from marine sediment, and emendation of Phaeobacter daeponensis

    Science.gov (United States)

    Dogs, Marco; Teshima, Hazuki; Petersen, Jörn; Fiebig, Anne; Chertkov, Olga; Dalingault, Hajnalka; Chen, Amy; Pati, Amrita; Goodwin, Lynne A.; Chain, Patrick; Detter, John C.; Ivanova, Natalia; Lapidus, Alla; Rohde, Manfred; Gronow, Sabine; Kyrpides, Nikos C.; Woyke, Tanja; Simon, Meinhard; Göker, Markus; Klenk, Hans-Peter; Brinkhoff, Thorsten

    2013-01-01

    TF-218T is the type strain of the species Phaeobacter daeponensis Yoon et al. 2007, a facultatively anaerobic Phaeobacter species isolated from tidal flats. Here we describe the draft genome sequence and annotation of this bacterium together with previously unreported aspects of its phenotype. We analyzed the genome for genes involved in secondary metabolite production and its anaerobic lifestyle, which have also been described for its closest relative Phaeobacter caeruleus. The 4,642,596 bp long genome of strain TF-218T contains 4,310 protein-coding genes and 78 RNA genes including four rRNA operons and consists of five replicons: one chromosome and four extrachromosomal elements with sizes of 276 kb, 174 kb, 117 kb and 90 kb. Genome analysis showed that TF-218T possesses all of the genes for indigoidine biosynthesis, and on specific media the strain showed a blue pigmentation. We also found genes for dissimilatory nitrate reduction, gene-transfer agents, NRPS/ PKS genes and signaling systems homologous to the LuxR/I system. PMID:24501652

  13. The abundance of pink-pigmented facultative methylotrophs in the root zone of plant species in invaded coastal sage scrub habitat.

    Science.gov (United States)

    Irvine, Irina C; Brigham, Christy A; Suding, Katharine N; Martiny, Jennifer B H

    2012-01-01

    Pink-pigmented facultative methylotrophic bacteria (PPFMs) are associated with the roots, leaves and seeds of most terrestrial plants and utilize volatile C(1) compounds such as methanol generated by growing plants during cell division. PPFMs have been well studied in agricultural systems due to their importance in crop seed germination, yield, pathogen resistance and drought stress tolerance. In contrast, little is known about the PPFM abundance and diversity in natural ecosystems, let alone their interactions with non-crop species. Here we surveyed PPFM abundance in the root zone soil of 5 native and 5 invasive plant species along ten invasion gradients in Southern California coastal sage scrub habitat. PPFMs were present in every soil sample and ranged in abundance from 10(2) to 10(5) CFU/g dry soil. This abundance varied significantly among plant species. PPFM abundance was 50% higher in the root zones of annual or biennial species (many invasives) than perennial species (all natives). Further, PPFM abundance appears to be influenced by the plant community beyond the root zone; pure stands of either native or invasive species had 50% more PPFMs than mixed species stands. In sum, PPFM abundance in the root zone of coastal sage scrub plants is influenced by both the immediate and surrounding plant communities. The results also suggest that PPFMs are a good target for future work on plant-microorganism feedbacks in natural ecosystems.

  14. Evaluation of agricultural reuse practices and relevant guidelines for the alba rancho WWTP (primary and secondary facultative ponds) in Cochabamba, Bolivia.

    Science.gov (United States)

    Zabalaga, J; Amy, G; von Münch, E

    2007-01-01

    Many cities in developing countries are experiencing high population growth, which is generating chaotic and unplanned development, reducing land areas available for agriculture, and polluting surface and groundwater. Consequently, the reuse of untreated or partially treated wastewater for agricultural irrigation is increasing in arid and semi-arid regions in developing countries. Cochabamba city in Bolivia also has a high population growth. The climatic characteristics and the lack of clean water sources in this city are forcing the agriculture sector to use treated and untreated wastewater for irrigation. We investigated the effluent quality of the Alba Rancho Wastewater Treatment Plant (WWTP) in Cochabamba, Bolivia, and the existing effluent reuse practices for irrigation of fodder crops in the surrounding agricultural land (La Mayca area). The plant uses primary and secondary facultative ponds, and does not achieve the required effluent quality (according to Bolivian environmental law) for effluent BOD, COD, TDS and faecal coliforms. This paper also includes a brief comparison of guidelines for wastewater reuse in agriculture from several developing and developed countries, comparing the parameters measured as pollution indicators. It appears that for developed countries, the main concern is the health risk that reuse can cause to the farmers and consumers. For developing countries on the other hand, pollution reduction is currently the major aim in their guidelines and standards.

  15. Facultative slave-making ants Formica sanguinea label their slaves with own recognition cues instead of employing the strategy of chemical mimicry.

    Science.gov (United States)

    Włodarczyk, Tomasz; Szczepaniak, Lech

    2017-01-01

    Slave-making ant species use the host workforce to ensure normal colony functioning. Slaves are robbed as pupae from their natal nest and after eclosion, assume the parasite colony as their own. A possible factor promoting the successful integration of slaves into a foreign colony is congruence with the slave-makers in terms of cuticular hydrocarbons, which are known to play the role of recognition cues in social insects. Such an adaptation is observed in the obligate slave-making ant species, which are chemically adjusted to their slaves. To date, however, no reports have been available on facultative slave-making species, which represent an earlier stage of the evolution of slavery. Such an example is Formica sanguinea, which exploit F. fusca colonies as their main source of a slave workforce. Our results show that F. sanguinea ants have a distinct cuticular hydrocarbon profile, which contains compounds not present in free-living F. fusca ants from potential target nests. Moreover, enslaved F. fusca ants acquire hydrocarbons from their slave-making nestmates to such an extent that they become chemically differentiated from free-living, conspecific ants. Our study shows that F. sanguinea ants promote their own recognition cues in their slaves, rather than employing the strategy of chemical mimicry. Possible reasons why F. sanguinea is not chemically well adjusted to its main host species are discussed in this paper. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Net ion fluxes in the facultative air-breather Hoplosternum littorale (tamoata) and the obligate air-breather Arapaima gigas (pirarucu) exposed to different Amazonian waters.

    Science.gov (United States)

    Baldisserotto, Bernardo; Copatti, Carlos E; Gomes, Levy C; Chagas, Edsandra C; Brinn, Richard P; Roubach, Rodrigo

    2008-12-01

    Fishes that live in the Amazon environment may be exposed to several kinds of water: black water (BW), acidic black water (pH 3.5) (ABW) and white water (WW), among others. The aim of the present study was to analyze net ion fluxes in the facultative air-breather Hoplosternum littorale (tamoata) and the obligate air-breather Arapaima gigas (pirarucu) exposed to different types of water. Fishes were acclimated in well water and later placed in individual chambers containing one type of water for ion flux measurements. After 4 h, the water in the chambers was replaced by a different type of water. The transfer of both species to ABW (independent of previous water exposure) increased net ion loss. Tamoatas transferred from ABW to BW or WW presented a net ion influx, but pirarucus showed only small changes on net ion efflux. These results allow us to conclude that tamoatas and pirarucus present differences in terms of ion regulation but that the general aspects of the ion flux are similar: (1) exposure to ABW led to net ion loss; (2) transfer from BW to WW or vice-versa induced only minor changes on net ion fluxes. These observations demonstrate that any osmoregulatory difficulties encountered by either species during changes between these latter two waters can be easily overcome.

  17. The abundance of pink-pigmented facultative methylotrophs in the root zone of plant species in invaded coastal sage scrub habitat.

    Directory of Open Access Journals (Sweden)

    Irina C Irvine

    Full Text Available Pink-pigmented facultative methylotrophic bacteria (PPFMs are associated with the roots, leaves and seeds of most terrestrial plants and utilize volatile C(1 compounds such as methanol generated by growing plants during cell division. PPFMs have been well studied in agricultural systems due to their importance in crop seed germination, yield, pathogen resistance and drought stress tolerance. In contrast, little is known about the PPFM abundance and diversity in natural ecosystems, let alone their interactions with non-crop species. Here we surveyed PPFM abundance in the root zone soil of 5 native and 5 invasive plant species along ten invasion gradients in Southern California coastal sage scrub habitat. PPFMs were present in every soil sample and ranged in abundance from 10(2 to 10(5 CFU/g dry soil. This abundance varied significantly among plant species. PPFM abundance was 50% higher in the root zones of annual or biennial species (many invasives than perennial species (all natives. Further, PPFM abundance appears to be influenced by the plant community beyond the root zone; pure stands of either native or invasive species had 50% more PPFMs than mixed species stands. In sum, PPFM abundance in the root zone of coastal sage scrub plants is influenced by both the immediate and surrounding plant communities. The results also suggest that PPFMs are a good target for future work on plant-microorganism feedbacks in natural ecosystems.

  18. Variation in Heavy Metal Accumulation and Genetic Diversity at a Regional Scale Among Metallicolous and Non-Metallicolous Populations of the Facultative Metallophyte Biscutella laevigata subsp. laevigata.

    Science.gov (United States)

    Pošćić, Filip; Fellet, Guido; Vischi, Massimo; Casolo, Valentino; Schat, Henk; Marchiol, Luca

    2015-01-01

    Biscutella laevigata is a facultative metallophyte, with populations on non-metalliferous and metalliferous soils. Some of its metallicolous populations have been shown to hyperaccumulate thallium or lead in nature. Only Tl hyperaccumulation has been experimentally confirmed. We aimed to compare the patterns of metal (hyper)accumulation and genetic diversity among populations of B. laevigata subsp. laevigata in NE Italy. None of the populations exhibited foliar hyperaccumulation of Cu, Zn, or Pb. The root-to-shoot accumulation rates for these metals were unchanged or decreased rather than enhanced in the metallicolous populations, in comparison with the non-metallicolous ones. Hyperaccumulation of Tl was confined to the population of the Cave del Predil mine. This population was genetically very distinct from the others, as demonstrated by AFLP-based cluster analysis. The two other mine populations did not surpass the threshold for Tl hyperaccumulation, but showed enhanced foliar Tl concentrations and root-to-shoot translocation rates, in comparison with the non-metallicolous populations. Genetic analysis suggested that adaptation to metalliferous soil must have been independently evolved in the metallicolous populations.

  19. Decreases in body temperature and body mass constitute pre-hibernation remodelling in the Syrian golden hamster, a facultative mammalian hibernator.

    Science.gov (United States)

    Chayama, Yuichi; Ando, Lisa; Tamura, Yutaka; Miura, Masayuki; Yamaguchi, Yoshifumi

    2016-04-01

    Hibernation is an adaptive strategy for surviving during periods with little or no food availability, by profoundly reducing the metabolic rate and the core body temperature (T b). Obligate hibernators (e.g. bears, ground squirrels, etc.) hibernate every winter under the strict regulation of endogenous circannual rhythms, and they are assumed to undergo adaptive remodelling in autumn, the pre-hibernation period, prior to hibernation. However, little is known about the nature of pre-hibernation remodelling. Syrian hamsters (Mesocricetus auratus) are facultative hibernators that can hibernate irrespective of seasons when exposed to prolonged short photoperiod and cold ambient temperature (SD-Cold) conditions. Their T b set point reduced by the first deep torpor (DT) and then increased gradually after repeated cycles of DT and periodic arousal (PA), and finally recovered to the level observed before the prolonged SD-Cold in the post-hibernation period. We also found that, before the initiation of hibernation, the body mass of animals decreased below a threshold, indicating that hibernation in this species depends on body condition. These observations suggest that Syrian hamsters undergo pre-hibernation remodelling and that T b and body mass can be useful physiological markers to monitor the remodelling process during the pre-hibernation period.

  20. Comparing mannose binding lectin genetic diversity in intracellular ...

    African Journals Online (AJOL)

    SERVER

    2007-09-05

    Sep 5, 2007 ... mannose binding lectin level for susceptibility to intracellular pathogens. Key words: .... The distribution of alleles and genotypes between groups were compared using ... Frequency of promoter variants and position +4 (Table.

  1. Identification and evaluation of semiochemicals for the biological control of the beetle Omorgus suberosus (F.) (Coleoptera: Trogidae), a facultative predator of eggs of the sea turtle Lepidochelys olivacea (Eschscholtz)

    OpenAIRE

    Cortez, Vieyle; José R. Verdú; Ortiz, Antonio J.; Halffter, Gonzalo

    2017-01-01

    The beetle Omorgus suberosus (F.) is a facultative predator of eggs of the olive ridley turtle Lepidochelys olivacea (Eschscholtz). Laboratory and field investigations were conducted in order to characterize volatile attractants of O. suberosus and to explore the potential for application of these volatiles in a selective mass trapping method. Headspace sorptive extraction (HSSE) coupled to thermo-desorption gas chromatography–mass spectrometry (TD-GC-MS) analysis of the volatile constituents...

  2. FLIPR assays of intracellular calcium in GPCR drug discovery

    DEFF Research Database (Denmark)

    Hansen, Kasper Bø; Bräuner-Osborne, Hans

    2009-01-01

    Fluorescent dyes sensitive to changes in intracellular calcium have become increasingly popular in G protein-coupled receptor (GPCR) drug discovery for several reasons. First of all, the assays using the dyes are easy to perform and are of low cost compared to other assays. Second, most non-Galph...... making them obtainable even for academic groups. Here, we present a protocol for measuring changes in intracellular calcium levels in living mammalian cells based on the fluorescent calcium binding dye, fluo-4....

  3. Study of neurotoxic intracellular calcium signalling triggered by amyloids.

    Science.gov (United States)

    Villalobos, Carlos; Caballero, Erica; Sanz-Blasco, Sara; Núñez, Lucía

    2012-01-01

    Neurotoxicity in Alzheimer's disease (AD) is associated to dishomeostasis of intracellular Ca(2+) induced by amyloid β peptide (Aβ) species. Understanding of the effects of Aβ on intracellular Ca(2+) homeostasis requires preparation of the different Aβ assemblies including oligomers and fibrils and the testing of their effects on cytosolic and mitochondrial Ca(2+) in neurons. Procedures for cerebellar granule cell culture, preparation of Aβ species as well as fluorescence and bioluminescence imaging of cytosolic and mitochondrial Ca(2+) in neurons are described.

  4. Intracellular concentrations determine the cytotoxicity of adefovir, cidofovir and tenofovir.

    Science.gov (United States)

    Zhang, Xun; Wang, Ruduan; Piotrowski, Mary; Zhang, Hui; Leach, Karen L

    2015-02-01

    Lack of in vitro to in vivo translation is a major challenge in safety prediction during early drug discovery.One of the most common in vitro assays to evaluate the probability of a compound to cause adverse effects is a cytotoxicity assay. Cytotoxicity of a compound is often measured by dose–response curves assuming the administered doses and intracellular exposures are equal at the time of measurement.However, this may not be true for compounds with low membrane permeability or those which are substrates for drug transporters as intracellular concentrations are determined both by passive permeability and active uptake through drug transporters. We show here that three antiviral drugs, adefovir, cidofovir and tenofovir exhibit significantly increased cytotoxicity in HEK293 cells transfected with organic anion transporter (OAT) 1 and 3 compared to a lack of cytotoxicity in HEK293 wildtype cells. A further look at the media and intracellular drug concentrations showed that 24 h after dosing, all three drugs had higher intracellular drug concentrations than that of media in the HEK-OAT1 cells whereas the intracellular drug concentrations in the wildtype cells were much lower than the administered doses. Comparing cytotoxicity IC(50) values of adefovir, cidofovir and tenofovir based on administered doses and measured intracellular concentrations in HEK-OAT1 cells revealed that intracellular drug concentrations have significant impact on calculated IC(50) values. Tenofovir showed much less intrinsic cytotoxicity than adefovir and cidofovir using intracellular concentrations rather than media concentration. Our data suggest that for low permeable drugs or drugs that are substrates for drug transporters, the choice of cellular model is critical for providing an accurate determination of cytotoxicity.

  5. In vivo metabolite compartmentalization probed using intracellular GdDTPA

    DEFF Research Database (Denmark)

    Peters, David Alberg; Rowland, Ian

    Fast trans-membrane water exchange enables in- tracellular relaxation enhancement of water by contrast agents in the extracellular space. For me- tabolites not in fast exchange across membranes, intracellular metabolite relaxation enhancement will only occur if the contrast agent and metabolite a...... are in the same compartment. Extracellular contrast has utilized electroporation methods to deliver GdDTPA into the cytosol of rat muscle in vivo in order to probe the intracellular compart- mentalization of MR-visible metabolites....

  6. Intra-cellular Staphylococcus aureus alone causes infection in vivo

    Directory of Open Access Journals (Sweden)

    T Hamza

    2013-07-01

    Full Text Available Chronic and recurrent bone infections occur frequently but have not been explained. Staphylococcus aureus (S. aureus is often found among chronic and recurrent infections and may be responsible for such infections. One possible reason is that S. aureus can internalize and survive within host cells and by doing so, S. aureus can evade both host defense mechanisms and most conventional antibiotic treatments. In this study, we hypothesized that intra-cellular S. aureus could induce infections in vivo. Osteoblasts were infected with S. aureus and, after eliminating extra-cellular S. aureus, inoculated into an open fracture rat model. Bacterial cultures and radiographic observations at post-operative day 21 confirmed local bone infections in animals inoculated with intra-cellular S. aureus within osteoblasts alone. We present direct in vivo evidence that intra-cellular S. aureus could be sufficient to induce bone infection in animals; we found that intra-cellular S. aureus inoculation of as low as 102 colony forming units could induce severe bone infections. Our data may suggest that intra-cellular S. aureus can “hide” in host cells during symptom-free periods and, under certain conditions, they may escape and lead to infection recurrence. Intra-cellular S. aureus therefore could play an important role in the pathogenesis of S. aureus infections, especially those chronic and recurrent infections in which disease episodes may be separated by weeks, months, or even years.

  7. Compromised Lactobacillus helveticus starter activity in the presence of facultative heterofermentative Lactobacillus casei DPC6987 results in atypical eye formation in Swiss-type cheese.

    Science.gov (United States)

    O'Sullivan, Daniel J; McSweeney, Paul L H; Cotter, Paul D; Giblin, Linda; Sheehan, Jeremiah J

    2016-04-01

    Nonstarter lactic acid bacteria are commonly implicated in undesirable gas formation in several varieties, including Cheddar, Dutch-, and Swiss-type cheeses, primarily due to their ability to ferment a wide variety of substrates. This effect can be magnified due to factors that detrimentally affect the composition or activity of starter bacteria, resulting in the presence of greater than normal amounts of fermentable carbohydrates and citrate. The objective of this study was to determine the potential for a facultatively heterofermentative Lactobacillus (Lactobacillus casei DPC6987) isolated from a cheese plant environment to promote gas defects in the event of compromised starter activity. A Swiss-type cheese was manufactured, at pilot scale and in triplicate, containing a typical starter culture (Streptococcus thermophilus and Lactobacillus helveticus) together with propionic acid bacteria. Lactobacillus helveticus populations were omitted in certain vats to mimic starter failure. Lactobacillus casei DPC6987 was added to each experimental vat at 4 log cfu/g. Cheese compositional analysis and X-ray computed tomography revealed that the failure of starter bacteria, in this case L. helveticus, coupled with the presence of a faculatively heterofermentative Lactobacillus (L. casei) led to excessive eye formation during ripening. The availability of excess amounts of lactose, galactose, and citrate during the initial ripening stages likely provided the heterofermentative L. casei with sufficient substrates for gas formation. The accrual of these fermentable substrates was notable in cheeses lacking the L. helveticus starter population. The results of this study are commercially relevant, as they demonstrate the importance of viability of starter populations and the control of specific nonstarter lactic acid bacteria to ensure appropriate eye formation in Swiss-type cheese.

  8. Effects of nitrite exposure on functional haemoglobin levels, bimodal respiration, and swimming performance in the facultative air-breathing fish Pangasianodon hypophthalmus.

    Science.gov (United States)

    Lefevre, Sjannie; Jensen, Frank B; Huong, Do T T; Wang, Tobias; Phuong, Nguyen T; Bayley, Mark

    2011-07-01

    In this study we investigated nitrite (NO₂⁻) effects in striped catfish, a facultative air-breather. Fish were exposed to 0, 0.4, and 0.9 mM nitrite for 0, 1, 2, 4, and 7 days, and levels of functional haemoglobin, methaemoglobin (metHb) and nitrosyl haemoglobin (HbNO) were assessed using spectral deconvolution. Plasma concentrations of nitrite, nitrate, chloride, potassium, and sodium were also measured. Partitioning of oxygen consumption was determined to reveal whether elevated metHb (causing functional hypoxia) induced air-breathing. The effects of nitrite on maximum oxygen uptake (MO(2max)) and critical swimming speed (U(crit)) were also assessed. Striped catfish was highly tolerant to nitrite exposure, as reflected by a 96 h LC₅₀ of 1.65 mM and a moderate nitrite uptake into the blood. Plasma levels of nitrite reached a maximum after 1 day of exposure, and then decreased, never exceeding ambient levels. MetHb, HbNO and nitrate (a nitrite detoxification product) also peaked after 1 day and then decreased. Only high levels of nitrite and metHb caused reductions in MO(2max) and U(crit). The response of striped catfish contrasts with that seen in most other fish species and discloses efficient mechanisms of combating nitrite threats. Furthermore, even though striped catfish is an efficient air-breather, this species has the ability to sustain aerobic scope and swimming performance without air-breathing, even when faced with nitrite-induced reductions in blood oxygen carrying capacity. Our study is the first to confirm that high levels of nitrite and metHb reduce MO(2max) and thereby aerobic scope, while more moderate elevations fail to do so. Further studies are needed to elucidate the mechanisms underlying the low nitrite accumulation in striped catfish.

  9. Effects of nitrite exposure on functional haemoglobin levels, bimodal respiration, and swimming performance in the facultative air-breathing fish Pangasianodon hypophthalmus

    Energy Technology Data Exchange (ETDEWEB)

    Lefevre, Sjannie, E-mail: sjannie.lefevre@biology.au.dk [Zoophysiology, Department of Biological Sciences, Aarhus University, Aarhus (Denmark); Jensen, Frank B. [Department of Biology, University of Southern Denmark, Odense (Denmark); Huong, Do.T.T. [College of Aquaculture and Fisheries, Can Tho University, Can Tho City (Viet Nam); Wang, Tobias [Zoophysiology, Department of Biological Sciences, Aarhus University, Aarhus (Denmark); Phuong, Nguyen T. [College of Aquaculture and Fisheries, Can Tho University, Can Tho City (Viet Nam); Bayley, Mark [Zoophysiology, Department of Biological Sciences, Aarhus University, Aarhus (Denmark)

    2011-07-15

    In this study we investigated nitrite (NO{sub 2}{sup -}) effects in striped catfish, a facultative air-breather. Fish were exposed to 0, 0.4, and 0.9 mM nitrite for 0, 1, 2, 4, and 7 days, and levels of functional haemoglobin, methaemoglobin (metHb) and nitrosyl haemoglobin (HbNO) were assessed using spectral deconvolution. Plasma concentrations of nitrite, nitrate, chloride, potassium, and sodium were also measured. Partitioning of oxygen consumption was determined to reveal whether elevated metHb (causing functional hypoxia) induced air-breathing. The effects of nitrite on maximum oxygen uptake (MO{sub 2max}) and critical swimming speed (U{sub crit}) were also assessed. Striped catfish was highly tolerant to nitrite exposure, as reflected by a 96 h LC{sub 50} of 1.65 mM and a moderate nitrite uptake into the blood. Plasma levels of nitrite reached a maximum after 1 day of exposure, and then decreased, never exceeding ambient levels. MetHb, HbNO and nitrate (a nitrite detoxification product) also peaked after 1 day and then decreased. Only high levels of nitrite and metHb caused reductions in MO{sub 2max} and U{sub crit}. The response of striped catfish contrasts with that seen in most other fish species and discloses efficient mechanisms of combating nitrite threats. Furthermore, even though striped catfish is an efficient air-breather, this species has the ability to sustain aerobic scope and swimming performance without air-breathing, even when faced with nitrite-induced reductions in blood oxygen carrying capacity. Our study is the first to confirm that high levels of nitrite and metHb reduce MO{sub 2max} and thereby aerobic scope, while more moderate elevations fail to do so. Further studies are needed to elucidate the mechanisms underlying the low nitrite accumulation in striped catfish.

  10. Effects of food availability on yolk androgen deposition in the black-legged kittiwake (Rissa tridactyla, a seabird with facultative brood reduction.

    Directory of Open Access Journals (Sweden)

    Z M Benowitz-Fredericks

    Full Text Available In birds with facultative brood reduction, survival of the junior chick is thought to be regulated primarily by food availability. In black-legged kittiwakes (Rissa tridactyla where parents and chicks are provided with unlimited access to supplemental food during the breeding season, brood reduction still occurs and varies interannually. Survival of the junior chick is therefore affected by factors in addition to the amount of food directly available to them. Maternally deposited yolk androgens affect competitive dynamics within a brood, and may be one of the mechanisms by which mothers mediate brood reduction in response to a suite of environmental and physiological cues. The goal of this study was to determine whether food supplementation during the pre-lay period affected patterns of yolk androgen deposition in free-living kittiwakes in two years (2003 and 2004 that varied in natural food availability. Chick survival was measured concurrently in other nests where eggs were not collected. In both years, supplemental feeding increased female investment in eggs by increasing egg mass. First-laid ("A" eggs were heavier but contained less testosterone and androstenedione than second-laid ("B" eggs across years and treatments. Yolk testosterone was higher in 2003 (the year with higher B chick survival across treatments. The difference in yolk testosterone levels between eggs within a clutch varied among years and treatments such that it was relatively small when B chick experienced the lowest and the highest survival probabilities, and increased with intermediate B chick survival probabilities. The magnitude of testosterone asymmetry in a clutch may allow females to optimize fitness by either predisposing a brood for reduction or facilitating survival of younger chicks.

  11. The Biosynthesis of Deuterium Labeled Amino Acids Using a Strain of Facultative Methylotrophic Bacterium Вrevibacterium Methylicum 5662 With RuMP Cycle of Carbon Assimilation

    Directory of Open Access Journals (Sweden)

    Oleg Mosin

    2015-03-01

    Full Text Available We used Gram-positive aerobic facultative methylotrophic bacterium, Brevibacterium methylicum, L-phenylalanine producer with ribulose-5-monophosphate (RuMP cycle for carbon assimilation for microbiological preparation of [2H]phenylalanine via conversion of low molecular weight substrates ([U-2H]MeOH and 2H2O. For this purpose, the cells of the methylotroph with improved growth characteristics were used on minimal salt media M9 supplemented with 2 % (v/v [U-2H]MeOH and increasing gradient of 2Н2O concentration from 0; 24,5; 49,0; 73,5 up to 98 % (v/v 2Н2O. L-phenylalanine was isolated from the growth medium after adding 5 M 2HCl (in 2Н2О, pH = 2,0 by extraction with isopropanol and subsequent crystallization in ethanol (output 0,65 g/l. Alanine, valine, and leucine/isoleucine were produced and accumulated exogenously in amounts of 5–6 mol in addition to the main product of biosynthesis. The method allows to obtain [2Н]amino acids with different levels of deuterium enrichment, depending on 2Н2O concentration in growth media, from 17 atom% 2Н (2 deuterium atoms (on the growth medium with 24,5 % (v/v 2Н2О up to 75 atom% 2Н (6 deuterium atoms (on the growth medium with 98 % (v/v 2Н2О with introduction of deuterium to benzyl С6Н5СН2-fragment of molecule that is confirmed with the data of electron impact (EI mass spectrometry analysis of methyl ethers of N-5-dimethylamino(naphthalene-1-sulfochloride [2H]amino acids after the separation by reverse-phase HPLC.

  12. [Microbial synthesis of deuterium labelled L-phenylalanine with different levels of isotopic enrichment by facultative methylotrophic bacterium Brevibacterium methylicum with RMP assimilation of carbon].

    Science.gov (United States)

    Mosin, O V; Shvets, V I; Skladnev, D A; Ignatov, I

    2014-01-01

    The preparative microbial synthesis of amino acids labelled with stable isotopes, including deuterium ( 2 H), suitable for biomedical applications by methylotrophic bacteria was studied using L-phenylalanine as example. This amino acid is secreted by Gram-negative aerobic facultative methylotrophic bacteria Brevibacterium methylicum, assimilating methanol via ribulose-5-monophosphate (RMP) cycle of assimilation of carbon, The data on adaptation of L-phenylalanine secreted by methylotrophic bacterium В. methylicum to the maximal concentration of deuterium in the growth medium with 98% 2 Н 2 O and 2% [ 2 Н]methanol, and biosynthesis of deuterium labelled L-phenylalanine With different levels of enrichment are presented. The strain was adapted by means of plating initial cells on firm (2% agarose) minimal growth media with an increasing gradient of 2 Н 2 O concentration from 0; 24.5; 49.0; 73.5 up to 98% 2 Н 2 O followed by subsequent selection of separate colonies stable to the action of 2 Н 2 O. These colonies were capable to produce L-phenylalanine. L-phenylalanine was extracted from growth medium by extraction with isopropanol with the subsequent crystallization in ethanol (output 0.65 g/l). The developed method of microbial synthesis allows to obtain deuterium labelled L-phenylalanine with different levels of isotopic enrichment, depending on concentration of 2 Н 2 O in growth media, from 17% (on growth medium with 24,5% 2 Н 2 O) up to 75% (on growth medium with 98% 2 Н 2 O) of deuterium in the molecule that is confirmed with the data of the electron impact (EI) mass- spectrometry analysis of methyl ethers of N-dimethylamino(naphthalene)-5-sulfochloride (dansyl) phenylalanine in these experimental conditions.

  13. Exploring anti-bacterial compounds against intracellular Legionella.

    Directory of Open Access Journals (Sweden)

    Christopher F Harrison

    Full Text Available Legionella pneumophila is a ubiquitous fresh-water bacterium which reproduces within its erstwhile predators, environmental amoeba, by subverting the normal pathway of phagocytosis and degradation. The molecular mechanisms which confer resistance to amoeba are apparently conserved and also allow replication within macrophages. Thus, L. pneumophila can act as an 'accidental' human pathogen and cause a severe pneumonia known as Legionnaires' disease. The intracellular localisation of L. pneumophila protects it from some antibiotics, and this fact must be taken into account to develop new anti-bacterial compounds. In addition, the intracellular lifestyle of L. pneumophila may render the bacteria susceptible to compounds diminishing bacterial virulence and decreasing intracellular survival and replication of this pathogen. The development of a single infection cycle intracellular replication assay using GFP-producing L. pneumophila and Acanthamoebacastellanii amoeba is reported here. This fluorescence-based assay allows for continuous monitoring of intracellular replication rates, revealing the effect of bacterial gene deletions or drug treatment. To examine how perturbations of the host cell affect L. pneumophila replication, several known host-targeting compounds were tested, including modulators of cytoskeletal dynamics, vesicle scission and Ras GTPase localisation. Our results reveal a hitherto unrealized potential antibiotic property of the β-lactone-based Ras depalmitoylation inhibitor palmostatin M, but not the closely related inhibitor palmostatin B. Further characterisation indicated that this compound caused specific growth inhibition of Legionella and Mycobacterium species, suggesting that it may act on a common bacterial target.

  14. Advances in genetic manipulation of obligate intracellular bacterial pathogens

    Directory of Open Access Journals (Sweden)

    Paul eBeare

    2011-05-01

    Full Text Available Infections by obligate intracellular bacterial pathogens result in significant morbidity and mortality worldwide. These bacteria include Chlamydia spp., which causes millions of cases of sexually transmitted disease and blinding trachoma annually, and members of the α-proteobacterial genera Anaplasma, Ehrlichia, Orientia and Rickettsia, agents of serious human illnesses including epidemic typhus. Coxiella burnetii, the agent of human Q fever, has also been considered a prototypical obligate intracellular bacterium, but recent host cell-free (axenic growth has rescued it from obligatism. The historic genetic intractability of obligate intracellular bacteria has severely limited molecular dissection of their unique lifestyles and virulence factors involved in pathogenesis. Host cell restricted growth is a significant barrier to genetic transformation that can make simple procedures for free-living bacteria, such as cloning, exceedingly difficult. Low transformation efficiency requiring long term culture in host cells to expand small transformant populations is another obstacle. Despite numerous technical limitations, the last decade has witnessed significant gains in genetic manipulation of obligate intracellular bacteria including allelic exchange. Continued development of genetic tools should soon enable routine mutation and complementation strategies for virulence factor discovery and stimulate renewed interest in these refractory pathogens. In this review, we discuss the technical challenges associated with genetic transformation of obligate intracellular bacteria and highlight advances made with individual genera.

  15. Intracellular calcium levels can regulate Importin-dependent nuclear import

    Energy Technology Data Exchange (ETDEWEB)

    Kaur, Gurpreet; Ly-Huynh, Jennifer D.; Jans, David A., E-mail: David.Jans@monash.edu

    2014-07-18

    Highlights: • High intracellular calcium inhibits Impα/β1- or Impβ1-dependent nuclear protein import. • The effect of Ca{sup 2+} on nuclear import does not relate to changes in the nuclear pore. • High intracellular calcium can result in mislocalisation of Impβ1, Ran and RCC1. - Abstract: We previously showed that increased intracellular calcium can modulate Importin (Imp)β1-dependent nuclear import of SRY-related chromatin remodeling proteins. Here we extend this work to show for the first time that high intracellular calcium inhibits Impα/β1- or Impβ1-dependent nuclear protein import generally. The basis of this relates to the mislocalisation of the transport factors Impβ1 and Ran, which show significantly higher nuclear localization in contrast to various other factors, and RCC1, which shows altered subnuclear localisation. The results here establish for the first time that intracellular calcium modulates conventional nuclear import through direct effects on the nuclear transport machinery.

  16. Intracellular Ca-carbonate biomineralization is widespread in cyanobacteria.

    Science.gov (United States)

    Benzerara, Karim; Skouri-Panet, Feriel; Li, Jinhua; Férard, Céline; Gugger, Muriel; Laurent, Thierry; Couradeau, Estelle; Ragon, Marie; Cosmidis, Julie; Menguy, Nicolas; Margaret-Oliver, Isabel; Tavera, Rosaluz; López-García, Purificación; Moreira, David

    2014-07-29

    Cyanobacteria have played a significant role in the formation of past and modern carbonate deposits at the surface of the Earth using a biomineralization process that has been almost systematically considered induced and extracellular. Recently, a deep-branching cyanobacterial species, Candidatus Gloeomargarita lithophora, was reported to form intracellular amorphous Ca-rich carbonates. However, the significance and diversity of the cyanobacteria in which intracellular biomineralization occurs remain unknown. Here, we searched for intracellular Ca-carbonate inclusions in 68 cyanobacterial strains distributed throughout the phylogenetic tree of cyanobacteria. We discovered that diverse unicellular cyanobacterial taxa form intracellular amorphous Ca-carbonates with at least two different distribution patterns, suggesting the existence of at least two distinct mechanisms of biomineralization: (i) one with Ca-carbonate inclusions scattered within the cell cytoplasm such as in Ca. G. lithophora, and (ii) another one observed in strains belonging to the Thermosynechococcus elongatus BP-1 lineage, in which Ca-carbonate inclusions lie at the cell poles. This pattern seems to be linked with the nucleation of the inclusions at the septum of the cells, showing an intricate and original connection between cell division and biomineralization. These findings indicate that intracellular Ca-carbonate biomineralization by cyanobacteria has been overlooked by past studies and open new perspectives on the mechanisms and the evolutionary history of intra- and extracellular Ca-carbonate biomineralization by cyanobacteria.

  17. Intracellular Ca-carbonate biomineralization is widespread in cyanobacteria

    Science.gov (United States)

    Benzerara, Karim; Skouri-Panet, Feriel; Li, Jinhua; Férard, Céline; Gugger, Muriel; Laurent, Thierry; Couradeau, Estelle; Ragon, Marie; Cosmidis, Julie; Menguy, Nicolas; Margaret-Oliver, Isabel; Tavera, Rosaluz; López-García, Purificación; Moreira, David

    2014-07-01

    Cyanobacteria have played a significant role in the formation of past and modern carbonate deposits at the surface of the Earth using a biomineralization process that has been almost systematically considered induced and extracellular. Recently, a deep-branching cyanobacterial species, Candidatus Gloeomargarita lithophora, was reported to form intracellular amorphous Ca-rich carbonates. However, the significance and diversity of the cyanobacteria in which intracellular biomineralization occurs remain unknown. Here, we searched for intracellular Ca-carbonate inclusions in 68 cyanobacterial strains distributed throughout the phylogenetic tree of cyanobacteria. We discovered that diverse unicellular cyanobacterial taxa form intracellular amorphous Ca-carbonates with at least two different distribution patterns, suggesting the existence of at least two distinct mechanisms of biomineralization: (i) one with Ca-carbonate inclusions scattered within the cell cytoplasm such as in Ca. G. lithophora, and (ii) another one observed in strains belonging to the Thermosynechococcus elongatus BP-1 lineage, in which Ca-carbonate inclusions lie at the cell poles. This pattern seems to be linked with the nucleation of the inclusions at the septum of the cells, showing an intricate and original connection between cell division and biomineralization. These findings indicate that intracellular Ca-carbonate biomineralization by cyanobacteria has been overlooked by past studies and open new perspectives on the mechanisms and the evolutionary history of intra- and extracellular Ca-carbonate biomineralization by cyanobacteria.

  18. Exploring Anti-Bacterial Compounds against Intracellular Legionella

    Science.gov (United States)

    Harrison, Christopher F.; Kicka, Sébastien; Trofimov, Valentin; Berschl, Kathrin; Ouertatani-Sakouhi, Hajer; Ackermann, Nikolaus; Hedberg, Christian; Cosson, Pierre; Soldati, Thierry; Hilbi, Hubert

    2013-01-01

    Legionella pneumophila is a ubiquitous fresh-water bacterium which reproduces within its erstwhile predators, environmental amoeba, by subverting the normal pathway of phagocytosis and degradation. The molecular mechanisms which confer resistance to amoeba are apparently conserved and also allow replication within macrophages. Thus, L. pneumophila can act as an ‘accidental’ human pathogen and cause a severe pneumonia known as Legionnaires’ disease. The intracellular localisation of L. pneumophila protects it from some antibiotics, and this fact must be taken into account to develop new anti-bacterial compounds. In addition, the intracellular lifestyle of L. pneumophila may render the bacteria susceptible to compounds diminishing bacterial virulence and decreasing intracellular survival and replication of this pathogen. The development of a single infection cycle intracellular replication assay using GFP-producing L. pneumophila and Acanthamoebacastellanii amoeba is reported here. This fluorescence-based assay allows for continuous monitoring of intracellular replication rates, revealing the effect of bacterial gene deletions or drug treatment. To examine how perturbations of the host cell affect L. pneumophila replication, several known host-targeting compounds were tested, including modulators of cytoskeletal dynamics, vesicle scission and Ras GTPase localisation. Our results reveal a hitherto unrealized potential antibiotic property of the β-lactone-based Ras depalmitoylation inhibitor palmostatin M, but not the closely related inhibitor palmostatin B. Further characterisation indicated that this compound caused specific growth inhibition of Legionella and Mycobacterium species, suggesting that it may act on a common bacterial target. PMID:24058631

  19. Quantifying intracellular hydrogen peroxide perturbations in terms of concentration

    Science.gov (United States)

    Huang, Beijing K.; Sikes, Hadley D.

    2014-01-01

    Molecular level, mechanistic understanding of the roles of reactive oxygen species (ROS) in a variety of pathological conditions is hindered by the difficulties associated with determining the concentration of various ROS species. Here, we present an approach that converts fold-change in the signal from an intracellular sensor of hydrogen peroxide into changes in absolute concentration. The method uses extracellular additions of peroxide and an improved biochemical measurement of the gradient between extracellular and intracellular peroxide concentrations to calibrate the intracellular sensor. By measuring peroxiredoxin activity, we found that this gradient is 650-fold rather than the 7–10-fold that is widely cited. The resulting calibration is important for understanding the mass-action kinetics of complex networks of redox reactions, and it enables meaningful characterization and comparison of outputs from endogenous peroxide generating tools and therapeutics across studies. PMID:25460730

  20. Modulation of mammalian apoptotic pathways by intracellular protozoan parasites.

    Science.gov (United States)

    Rodrigues, V; Cordeiro-da-Silva, A; Laforge, M; Ouaissi, A; Silvestre, R; Estaquier, J

    2012-03-01

    During intracellular parasitic infections, pathogens and host cells take part in a complex web of events that are crucial for the outcome of the infection. Modulation of host cell apoptosis by pathogens attracted the attention of scientists during the last decade. Apoptosis is an efficient mechanism used by the host to control infection and limit pathogen multiplication and dissemination. In order to ensure completion of their complex life cycles and to guarantee transmission between different hosts, intracellular parasites have developed mechanisms to block apoptosis and sustain the viability of their host cells. Here, we review how some of the most prominent intracellular protozoan parasites modulate the main mammalian apoptotic pathways by emphasizing the advances from the last decade, which have begun to dissect this dynamic and complex interaction.

  1. Novel antibody-antibiotic conjugate eliminates intracellular S. aureus.

    Science.gov (United States)

    Lehar, Sophie M; Pillow, Thomas; Xu, Min; Staben, Leanna; Kajihara, Kimberly K; Vandlen, Richard; DePalatis, Laura; Raab, Helga; Hazenbos, Wouter L; Morisaki, J Hiroshi; Kim, Janice; Park, Summer; Darwish, Martine; Lee, Byoung-Chul; Hernandez, Hilda; Loyet, Kelly M; Lupardus, Patrick; Fong, Rina; Yan, Donghong; Chalouni, Cecile; Luis, Elizabeth; Khalfin, Yana; Plise, Emile; Cheong, Jonathan; Lyssikatos, Joseph P; Strandh, Magnus; Koefoed, Klaus; Andersen, Peter S; Flygare, John A; Wah Tan, Man; Brown, Eric J; Mariathasan, Sanjeev

    2015-11-19

    Staphylococcus aureus is considered to be an extracellular pathogen. However, survival of S. aureus within host cells may provide a reservoir relatively protected from antibiotics, thus enabling long-term colonization of the host and explaining clinical failures and relapses after antibiotic therapy. Here we confirm that intracellular reservoirs of S. aureus in mice comprise a virulent subset of bacteria that can establish infection even in the presence of vancomycin, and we introduce a novel therapeutic that effectively kills intracellular S. aureus. This antibody-antibiotic conjugate consists of an anti-S. aureus antibody conjugated to a highly efficacious antibiotic that is activated only after it is released in the proteolytic environment of the phagolysosome. The antibody-antibiotic conjugate is superior to vancomycin for treatment of bacteraemia and provides direct evidence that intracellular S. aureus represents an important component of invasive infections.

  2. Quantifying intracellular hydrogen peroxide perturbations in terms of concentration

    Directory of Open Access Journals (Sweden)

    Beijing K. Huang

    2014-01-01

    Full Text Available Molecular level, mechanistic understanding of the roles of reactive oxygen species (ROS in a variety of pathological conditions is hindered by the difficulties associated with determining the concentration of various ROS species. Here, we present an approach that converts fold-change in the signal from an intracellular sensor of hydrogen peroxide into changes in absolute concentration. The method uses extracellular additions of peroxide and an improved biochemical measurement of the gradient between extracellular and intracellular peroxide concentrations to calibrate the intracellular sensor. By measuring peroxiredoxin activity, we found that this gradient is 650-fold rather than the 7–10-fold that is widely cited. The resulting calibration is important for understanding the mass-action kinetics of complex networks of redox reactions, and it enables meaningful characterization and comparison of outputs from endogenous peroxide generating tools and therapeutics across studies.

  3. Cadmium induces transcription independently of intracellular calcium mobilization.

    Directory of Open Access Journals (Sweden)

    Brooke E Tvermoes

    Full Text Available BACKGROUND: Exposure to cadmium is associated with human pathologies and altered gene expression. The molecular mechanisms by which cadmium affects transcription remain unclear. It has been proposed that cadmium activates transcription by altering intracellular calcium concentration ([Ca(2+](i and disrupting calcium-mediated intracellular signaling processes. This hypothesis is based on several studies that may be technically problematic; including the use of BAPTA chelators, BAPTA-based fluorescent sensors, and cytotoxic concentrations of metal. METHODOLOGY/PRINCIPAL FINDING: In the present report, the effects of cadmium on [Ca(2+](i under non-cytotoxic and cytotoxic conditions was monitored using the protein-based calcium sensor yellow cameleon (YC3.60, which was stably expressed in HEK293 cells. In HEK293 constitutively expressing YC3.60, this calcium sensor was found to be insensitive to cadmium. Exposing HEK293::YC3.60 cells to non-cytotoxic cadmium concentrations was sufficient to induce transcription of cadmium-responsive genes but did not affect [Ca(2+](i mobilization or increase steady-state mRNA levels of calcium-responsive genes. In contrast, exposure to cytotoxic concentrations of cadmium significantly reduced intracellular calcium stores and altered calcium-responsive gene expression. CONCLUSIONS/SIGNIFICANCE: These data indicate that at low levels, cadmium induces transcription independently of intracellular calcium mobilization. The results also support a model whereby cytotoxic levels of cadmium activate calcium-responsive transcription as a general response to metal-induced intracellular damage and not via a specific mechanism. Thus, the modulation of intracellular calcium may not be a primary mechanism by which cadmium regulates transcription.

  4. Connecting extracellular metabolomic measurements to intracellular flux states in yeast

    Directory of Open Access Journals (Sweden)

    Herrgård Markus J

    2009-03-01

    Full Text Available Abstract Background Metabolomics has emerged as a powerful tool in the quantitative identification of physiological and disease-induced biological states. Extracellular metabolome or metabolic profiling data, in particular, can provide an insightful view of intracellular physiological states in a noninvasive manner. Results We used an updated genome-scale metabolic network model of Saccharomyces cerevisiae, iMM904, to investigate how changes in the extracellular metabolome can be used to study systemic changes in intracellular metabolic states. The iMM904 metabolic network was reconstructed based on an existing genome-scale network, iND750, and includes 904 genes and 1,412 reactions. The network model was first validated by comparing 2,888 in silico single-gene deletion strain growth phenotype predictions to published experimental data. Extracellular metabolome data measured in response to environmental and genetic perturbations of ammonium assimilation pathways was then integrated with the iMM904 network in the form of relative overflow secretion constraints and a flux sampling approach was used to characterize candidate flux distributions allowed by these constraints. Predicted intracellular flux changes were consistent with published measurements on intracellular metabolite levels and fluxes. Patterns of predicted intracellular flux changes could also be used to correctly identify the regions of the metabolic network that were perturbed. Conclusion Our results indicate that integrating quantitative extracellular metabolomic profiles in a constraint-based framework enables inferring changes in intracellular metabolic flux states. Similar methods could potentially be applied towards analyzing biofluid metabolome variations related to human physiological and disease states.

  5. Autophagic clearance of bacterial pathogens: molecular recognition of intracellular microorganisms.

    Science.gov (United States)

    Pareja, Maria Eugenia Mansilla; Colombo, Maria I

    2013-01-01

    Autophagy is involved in several physiological and pathological processes. One of the key roles of the autophagic pathway is to participate in the first line of defense against the invasion of pathogens, as part of the innate immune response. Targeting of intracellular bacteria by the autophagic machinery, either in the cytoplasm or within vacuolar compartments, helps to control bacterial proliferation in the host cell, controlling also the spreading of the infection. In this review we will describe the means used by diverse bacterial pathogens to survive intracellularly and how they are recognized by the autophagic molecular machinery, as well as the mechanisms used to avoid autophagic clearance.

  6. Role of intracellular calcium in contraction of internal anal sphincter

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    @@ INTRODUCTION Internal anal sphincter (IAS) is a continuation of the smooth circular muscle layer thickened at the rectum, innervated by vegetative nerve. IAS is a special smooth muscle, which is different from colonic smooth muscle in physiology and pharmaology[1]. It was found that contraction of gastric smooth muscle depends on the influx of extracellular calcium and release of intracellular calcium[2]. In present study, we observed and compared the effects of extra- and intracellular calcium on the contraction of IAS and colonic smooth muscle.

  7. In vitro and ex vivo strategies for intracellular delivery

    Science.gov (United States)

    Stewart, Martin P.; Sharei, Armon; Ding, Xiaoyun; Sahay, Gaurav; Langer, Robert; Jensen, Klavs F.

    2016-10-01

    Intracellular delivery of materials has become a critical component of genome-editing approaches, ex vivo cell-based therapies, and a diversity of fundamental research applications. Limitations of current technologies motivate development of next-generation systems that can deliver a broad variety of cargo to diverse cell types. Here we review in vitro and ex vivo intracellular delivery approaches with a focus on mechanisms, challenges and opportunities. In particular, we emphasize membrane-disruption-based delivery methods and the transformative role of nanotechnology, microfluidics and laboratory-on-chip technology in advancing the field.

  8. Biochemical and ultrastructural studies suggest that the effects of thapsigargin on human platelets are mediated by changes in intracellular calcium but not by intracellular histamine

    DEFF Research Database (Denmark)

    Saxena, S P; McNicol, A; Becker, A B;

    1992-01-01

    The involvement of intracellular histamine in thapsigargin (Tg)-induced platelet aggregation was studied. Platelet aggregation induced by 0.25 and 0.5 microM Tg was not accompanied by a rise in intracellular histamine but a significant (p <0.01) increase in the level of intracellular histamine wa...

  9. Intracellular localization of VAMP-1 protein in human neutrophils.

    Science.gov (United States)

    Nabokina, S M

    2001-02-01

    We studied the intracellular localization of vesicle-associated membrane protein VAMP-1 in human neutrophils. VAMP-1 was associated with membranes of gelatinase and specific secretory granules rapidly mobilized during exocytosis. VAMP-1 probably acts as a component of the SNARE complex during exocytosis of gelatinase and specific granules in human neutrophils.

  10. Imaging atrial arrhythmic intracellular calcium in intact heart.

    Science.gov (United States)

    Xie, Wenjun; Santulli, Gaetano; Guo, Xiaoxiao; Gao, Melanie; Chen, Bi-Xing; Marks, Andrew R

    2013-11-01

    Abnormalities in intracellular Ca(2+) signaling have been proposed to play an essential role in the pathophysiology of atrial arrhythmias. However, a direct observation of intracellular Ca(2+) in atrial myocytes during atrial arrhythmias is lacking. Here, we have developed an ex vivo model of simultaneous Ca(2+) imaging and electrocardiographic recording in cardiac atria. Using this system we were able to record atrial arrhythmic intracellular Ca(2+) activities. Our results indicate that atrial arrhythmias can be tightly linked to intracellular Ca(2+) waves and Ca(2+) alternans. Moreover, we applied this strategy to analyze Ca(2+) signals in the hearts of WT and knock-in mice harboring a 'leaky' type 2 ryanodine receptor (RyR2-R2474S). We showed that sarcoplasmic reticulum (SR) Ca(2+) leak increases the susceptibility to Ca(2+) alternans and Ca(2+) waves increasing the incidence of atrial arrhythmias. Reduction of SR Ca(2+) leak via RyR2 by acute treatment with S107 reduced both Ca(2+) alternans and Ca(2+) waves, and prevented atrial arrhythmias.

  11. Intracellular water motion decreases in apoptotic macrophages after caspase activation.

    Science.gov (United States)

    Hortelano, S; García-Martín, M L; Cerdán, S; Castrillo, A; Alvarez, A M; Boscá, L

    2001-10-01

    Triggering of the macrophage cell line RAW 264.7 with lipopolysaccharide and interferon-gamma promoted apoptosis that was prevented by inhibitors of type 2 nitric oxide synthase or caspase. Using (1)H NMR analysis, we have investigated the changes of the intracellular transverse relaxation time (T(2)) and apparent diffusion coefficient (ADC) as parameters reflecting the rotational and translational motions of water in apoptotic macrophages. T(2) values decreased significantly from 287 to 182 ms in cells treated for 18 h with NO-donors. These changes of T(2) were prevented by caspase inhibitors and were not due to mitochondrial depolarization or microtubule depolymerization. The decrease of the intracellular values of T(2) and ADC in apoptotic macrophages was observed after caspase activation, but preceded phosphatidylserine exposure and nucleosomal DNA cleavage. The changes of water motion were accompanied by an enhancement of the hydrophobic properties of the intracellular milieu, as detected by fluorescent probes. These results indicate the occurrence of an alteration in the physicochemical properties of intracellular water during the course of apoptosis.

  12. Intracellular pH measurements using perfluorocarbon nanoemulsions.

    Science.gov (United States)

    Patrick, Michael J; Janjic, Jelena M; Teng, Haibing; O'Hear, Meredith R; Brown, Cortlyn W; Stokum, Jesse A; Schmidt, Brigitte F; Ahrens, Eric T; Waggoner, Alan S

    2013-12-11

    We report the synthesis and formulation of unique perfluorocarbon (PFC) nanoemulsions enabling intracellular pH measurements in living cells via fluorescent microscopy and flow cytometry. These nanoemulsions are formulated to readily enter cells upon coincubation and contain two cyanine-based fluorescent reporters covalently bound to the PFC molecules, specifically Cy3-PFC and CypHer5-PFC conjugates. The spectral and pH-sensing properties of the nanoemulsions were characterized in vitro and showed the unaltered spectral behavior of dyes after formulation. In rat 9L glioma cells loaded with nanoemulsion, the local pH of nanoemulsions was longitudinally quantified using optical microscopy and flow cytometry and displayed a steady decrease in pH to a level of 5.5 over 3 h, indicating rapid uptake of nanoemulsion to acidic compartments. Overall, these reagents enable real-time optical detection of intracellular pH in living cells in response to pharmacological manipulations. Moreover, recent approaches for in vivo cell tracking using magnetic resonance imaging (MRI) employ intracellular PFC nanoemulsion probes to track cells using (19)F MRI. However, the intracellular fate of these imaging probes is poorly understood. The pH-sensing nanoemulsions allow the study of the fate of the PFC tracer inside the labeled cell, which is important for understanding the PFC cell loading dynamics, nanoemulsion stability and cell viability over time.

  13. Deciphering the Intracellular Fate of Propionibacterium acnes in Macrophages

    Directory of Open Access Journals (Sweden)

    Natalie Fischer

    2013-01-01

    Full Text Available Propionibacterium acnes is a Gram-positive bacterium that colonizes various niches of the human body, particularly the sebaceous follicles of the skin. Over the last years a role of this common skin bacterium as an opportunistic pathogen has been explored. Persistence of P. acnes in host tissue has been associated with chronic inflammation and disease development, for example, in prostate pathologies. This study investigated the intracellular fate of P. acnes in macrophages after phagocytosis. In a mouse model of P. acnes-induced chronic prostatic inflammation, the bacterium could be detected in prostate-infiltrating macrophages at 2 weeks postinfection. Further studies performed in the human macrophage cell line THP-1 revealed intracellular survival and persistence of P. acnes but no intracellular replication or escape from the host cell. Confocal analyses of phagosome acidification and maturation were performed. Acidification of P. acnes-containing phagosomes was observed at 6 h postinfection but then lost again, indicative of cytosolic escape of P. acnes or intraphagosomal pH neutralization. No colocalization with the lysosomal markers LAMP1 and cathepsin D was observed, implying that the P. acnes-containing phagosome does not fuse with lysosomes. Our findings give first insights into the intracellular fate of P. acnes; its persistency is likely to be important for the development of P. acnes-associated inflammatory diseases.

  14. Biomineralization Patterns of Intracellular Carbonatogenesis in Cyanobacteria: Molecular Hypotheses

    Directory of Open Access Journals (Sweden)

    Jinhua Li

    2016-02-01

    Full Text Available The recent discovery of intracellular carbonatogenesis in several cyanobacteria species has challenged the traditional view that this process was extracellular and not controlled. However, a detailed analysis of the size distribution, chemical composition and 3-D-arrangement of carbonates in these cyanobacteria is lacking. Here, we characterized these features in Candidatus Gloeomargarita lithophora C7 and Candidatus Synechococcus calcipolaris G9 by conventional transmission electron microscopy, tomography, ultramicrotomy, and scanning transmission X-ray microscopy (STXM. Both Ca. G. lithophora C7 and Ca. S. calcipolaris G9 formed numerous polyphosphate granules adjacent or engulfing Ca-carbonate inclusions when grown in phosphate-rich solutions. Ca-carbonates were scattered within Ca. G. lithophora C7 cells under these conditions, but sometimes arranged in one or several chains. In contrast, Ca-carbonates formed at cell septa in Ca. S. calcipolaris G9 and were segregated equally between daughter cells after cell division, arranging as distorted disks at cell poles. The size distribution of carbonates evolved from a positively to a negatively skewed distribution as particles grew. Conventional ultramicrotomy did not preserve Ca-carbonates explaining partly why intracellular calcification has been overlooked in the past. All these new observations allow discussing with unprecedented insight some nucleation and growth processes occurring in intracellularly calcifying cyanobacteria with a particular emphasis on the possible involvement of intracellular compartments and cytoskeleton.

  15. Galectin-3 guides intracellular trafficking of some human serotransferrin glycoforms

    DEFF Research Database (Denmark)

    Carlsson, Carl Michael; Bengtson, Per; Cucak, Helena

    2013-01-01

    these transferrin glycoforms differently after preloading with exogenously added galectin-3. In all, this study provides the first evidence of a functional role for transferrin glycans, in intracellular trafficking after uptake. Moreover, the galectin-3 bound glycoform increased in cancer, suggesting...

  16. Intracellular pH gradients in migrating cells

    DEFF Research Database (Denmark)

    Martin, Christine; Pedersen, Stine Helene Falsig; Schwab, Albrecht;

    2011-01-01

    might function as such unevenly distributed regulators as they modulate the interaction of focal adhesion proteins and components of the cytoskeleton in vitro. However, an intracellular pH (pH(i)) gradient reflecting a spatial asymmetry of protons has not been shown so far. One major regulator of p...

  17. CONTRIBUTIONS OF INTRACELLULAR IONS TO Kv CHANNEL VOLTAGE SENSOR DYNAMICS.

    Directory of Open Access Journals (Sweden)

    Samuel eGoodchild

    2012-06-01

    Full Text Available Voltage sensing domains of Kv channels control ionic conductance through coupling of the movement of charged residues in the S4 segment to conformational changes at the cytoplasmic region of the pore domain, that allow K+ ions to flow. Conformational transitions within the voltage sensing domain caused by changes in the applied voltage across the membrane field are coupled to the conducting pore region and the gating of ionic conductance. However, several other factors not directly linked to the voltage dependent movement of charged residues within the voltage sensor impact the dynamics of the voltage sensor, such as inactivation, ionic conductance, intracellular ion identity and block of the channel by intracellular ligands. The effect of intracellular ions on voltage sensor dynamics is of importance in the interpretation of gating current measurements and the physiology of pore/voltage sensor coupling. There is a significant amount of variability in the reported kinetics of voltage sensor deactivation kinetics of Kv channels attributed to different mechanisms such as open state stabilization, immobilization and relaxation processes of the voltage sensor. Here we separate these factors and focus on the causal role that intracellular ions can play in allosterically modulating the dynamics of Kv voltage sensor deactivation kinetics. These considerations are of critical importance in understanding the molecular determinants of the complete channel gating cycle from activation to deactivation.

  18. Manipulation of Host Cholesterol by Obligate Intracellular Bacteria

    Directory of Open Access Journals (Sweden)

    Dhritiman Samanta

    2017-05-01

    Full Text Available Cholesterol is a multifunctional lipid that plays important metabolic and structural roles in the eukaryotic cell. Despite having diverse lifestyles, the obligate intracellular bacterial pathogens Chlamydia, Coxiella, Anaplasma, Ehrlichia, and Rickettsia all target cholesterol during host cell colonization as a potential source of membrane, as well as a means to manipulate host cell signaling and trafficking. To promote host cell entry, these pathogens utilize cholesterol-rich microdomains known as lipid rafts, which serve as organizational and functional platforms for host signaling pathways involved in phagocytosis. Once a pathogen gains entrance to the intracellular space, it can manipulate host cholesterol trafficking pathways to access nutrient-rich vesicles or acquire membrane components for the bacteria or bacteria-containing vacuole. To acquire cholesterol, these pathogens specifically target host cholesterol metabolism, uptake, efflux, and storage. In this review, we examine the strategies obligate intracellular bacterial pathogens employ to manipulate cholesterol during host cell colonization. Understanding how obligate intracellular pathogens target and use host cholesterol provides critical insight into the host-pathogen relationship.

  19. Cell-penetrating antimicrobial peptides - prospectives for targeting intracellular infections

    DEFF Research Database (Denmark)

    Bahnsen, Jesper S; Franzyk, Henrik; Sayers, Edward J;

    2015-01-01

    . TPk showed the highest antibacterial activity. SA-3 exhibited selective disruption of liposomes mimicking Gram-positive and Gram-negative membranes. CONCLUSION: PK-12-KKP is an unlikely candidate for targeting intracellular bacteria, as the eukaryotic cell-penetrating ability is poor. SA-3, affected...

  20. Intracellular pH in rat pancreatic ducts

    DEFF Research Database (Denmark)

    Novak, I; Hug, M; Greger, R

    1997-01-01

    In order to study the mechanism of H+ and HCO3- transport in a HCO3- secreting epithelium, pancreatic ducts, we have measured the intracellular pH (pHi) in this tissue using the pH sensitive probe BCECF. We found that exposures of ducts to solutions containing acetate/acetic acid or NH4+/NH3 buff...

  1. Modulating cancer cell survival by targeting intracellular cholesterol transport.

    Science.gov (United States)

    Kuzu, Omer F; Gowda, Raghavendra; Noory, Mohammad A; Robertson, Gavin P

    2017-08-08

    Demand for cholesterol is high in certain cancers making them potentially sensitive to therapeutic strategies targeting cellular cholesterol homoeostasis. A potential approach involves disruption of intracellular cholesterol transport, which occurs in Niemann-Pick disease as a result of acid sphingomyelinase (ASM) deficiency. Hence, a class of lysosomotropic compounds that were identified as functional ASM inhibitors (FIASMAs) might exhibit chemotherapeutic activity by disrupting cancer cell cholesterol homoeostasis. Here, the chemotherapeutic utility of ASM inhibition was investigated. The effect of FIASMAs on intracellular cholesterol levels, cholesterol homoeostasis, cellular endocytosis and signalling cascades were investigated. The in vivo efficacy of ASM inhibition was demonstrated using melanoma xenografts and a nanoparticle formulation was developed to overcome dose-limiting CNS-associated side effects of certain FIASMAs. Functional ASM inhibitors inhibited intracellular cholesterol transport leading to disruption of autophagic flux, cellular endocytosis and receptor tyrosine kinase signalling. Consequently, major oncogenic signalling cascades on which cancer cells were reliant for survival were inhibited. Two tested ASM inhibitors, perphenazine and fluphenazine that are also clinically used as antipsychotics, were effective in inhibiting xenografted tumour growth. Nanoliposomal encapsulation of the perphenazine enhanced its chemotherapeutic efficacy while decreasing CNS-associated side effects. This study suggests that disruption of intracellular cholesterol transport by targeting ASM could be utilised as a potential chemotherapeutic approach for treating cancer.

  2. Monitoring intracellular oxidative events using dynamic spectral unmixing microscopy

    Science.gov (United States)

    There is increasing interest in using live-cell imaging to monitor not just individual intracellular endpoints, but to investigate the interplay between multiple molecular events as they unfold in real time within the cell. A major impediment to simultaneous acquisition of multip...

  3. Nutrient salvaging and metabolism by the intracellular pathogen Legionella pneumophila.

    Science.gov (United States)

    Fonseca, Maris V; Swanson, Michele S

    2014-01-01

    The Gram-negative bacterium Legionella pneumophila is ubiquitous in freshwater environments as a free-swimming organism, resident of biofilms, or parasite of protozoa. If the bacterium is aerosolized and inhaled by a susceptible human host, it can infect alveolar macrophages and cause a severe pneumonia known as Legionnaires' disease. A sophisticated cell differentiation program equips L. pneumophila to persist in both extracellular and intracellular niches. During its life cycle, L. pneumophila alternates between at least two distinct forms: a transmissive form equipped to infect host cells and evade lysosomal degradation, and a replicative form that multiplies within a phagosomal compartment that it has retooled to its advantage. The efficient changeover between transmissive and replicative states is fundamental to L. pneumophila's fitness as an intracellular pathogen. The transmission and replication programs of L. pneumophila are governed by a number of metabolic cues that signal whether conditions are favorable for replication or instead trigger escape from a spent host. Several lines of experimental evidence gathered over the past decade establish strong links between metabolism, cellular differentiation, and virulence of L. pneumophila. Herein, we focus on current knowledge of the metabolic components employed by intracellular L. pneumophila for cell differentiation, nutrient salvaging and utilization of host factors. Specifically, we highlight the metabolic cues that are coupled to bacterial differentiation, nutrient acquisition systems, and the strategies utilized by L. pneumophila to exploit host metabolites for intracellular replication.

  4. Bioresponsive poly(amidoamine)s designed for intracellular protein delivery

    NARCIS (Netherlands)

    Coue, G.M.J.P.C.; Freese, C.; Unger, R.E.; Kirkpatrick, C.J.; Engbersen, J.F.J.

    2013-01-01

    Poly(amidoamine)s with bioreducible disulfide linkages in the main chain (SS-PAAs) and pH-responsive, negatively charged citraconate groups in the sidechain have been designed for effective intracellular delivery and release of proteins with a net positive charge at neutral pH. Using lysozyme as a c

  5. Intracellular particle tracking as a tool for tumor cell characterization

    NARCIS (Netherlands)

    Li, Yixuan; Duits, Michel H.G.; Schnekenburger, Jürgen

    2009-01-01

    We studied the dynamics of two types of intracellular probe particles, ballistically injected latex spheres and endogenous granules, in tumor cell lines of differerent metastatic potential: breast tumor cells (MCF-7 malignant, MCF-10A benign) and pancreas adenocarcinoma (PaTu8988T malignant, PaTu898

  6. Legionella pneumophila transcriptome during intracellular multiplication in human macrophages

    Directory of Open Access Journals (Sweden)

    Sebastien P Faucher

    2011-04-01

    Full Text Available Legionella pneumophila is the causative agent of Legionnaires’ disease, an acute pulmonary infection. L. pneumophila is able to infect and multiply in both phagocytic protozoa, such as Acanthamoeba castellanii, and mammalian professional phagocytes. The best-known L. pneumophila virulence determinant is the Icm/Dot Type IVB secretion system (TFBSS, which is used to translocate more than 150 effector proteins to host cells. While the transcriptional response of Legionella to the intracellular environment of A. castellanii has been investigated, much less is known about the Legionella transcriptional response inside human macrophages. In this study, the transcriptome of L. pneumophila was monitored during exponential and post-exponential phase in rich AYE broth as well as during infection of human cultured macrophages. This was accomplished with microarrays and an RNA amplification procedure called SCOTS to detect small amounts of mRNA from low numbers of intracellular bacteria. Among the genes induced intracellularly are those involved in amino acid biosynthetic pathways leading to L-arginine, L-histidine and L-proline as well as many transport systems involved in amino acid and iron uptake. Gene involved in catabolism of glycerol is also induced during intracellular growth and could be used as a carbon source. The genes encoding the Icm/Dot system are not differentially expressed inside cells compared to control bacteria grown in rich broth, but the genes encoding several translocated effectors are strongly induced. Moreover, we used the transcriptome data to predict previously unrecognized Icm/Dot effector genes based on their expression pattern and confirmed translocation for three candidates. This study provides a comprehensive view of how L. pneumophila responds to the human macrophage intracellular environment.

  7. Enhanced intracellular delivery and antibacterial efficacy of enrofloxacin-loaded docosanoic acid solid lipid nanoparticles against intracellular Salmonella

    Science.gov (United States)

    Xie, Shuyu; Yang, Fei; Tao, Yanfei; Chen, Dongmei; Qu, Wei; Huang, Lingli; Liu, Zhenli; Pan, Yuanhu; Yuan, Zonghui

    2017-01-01

    Enrofloxacin-loaded docosanoic acid solid lipid nanoparticles (SLNs) with different physicochemical properties were developed to enhance activity against intracellular Salmonella. Their cellular uptake, intracellular elimination and antibacterial activity were studied in RAW 264.7 cells. During the experimental period, SLN-encapsulated enrofloxacin accumulated in the cells approximately 27.06–37.71 times more efficiently than free drugs at the same extracellular concentration. After incubation for 0.5 h, the intracellular enrofloxacin was enhanced from 0.336 to 1.147 μg/mg of protein as the sizes of nanoparticles were increased from 150 to 605 nm, and from 0.960 to 1.147 μg/mg of protein when the charge was improved from −8.1 to −24.9 mv. The cellular uptake was more significantly influenced by the size than it was by the charge, and was not affected by whether the charge was positive or negative. The elimination of optimal SLN-encapsulated enrofloxacin from the cells was significantly slower than that of free enrofloxacin after removing extracellular drug. The inhibition effect against intracellular Salmonella CVCC541 of 0.24 and 0.06 μg/mL encapsulated enrofloxacin was stronger than 0.6 μg/mL free drug after all of the incubation periods and at 48 h, respectively. Docosanoic acid SLNs are thus considered as a promising carrier for intracellular bacterial treatment. PMID:28112240

  8. Analysing intracellular deformation of polymer capsules using structured illumination microscopy

    Science.gov (United States)

    Chen, Xi; Cui, Jiwei; Sun, Huanli; Müllner, Markus; Yan, Yan; Noi, Ka Fung; Ping, Yuan; Caruso, Frank

    2016-06-01

    Understanding the behaviour of therapeutic carriers is important in elucidating their mechanism of action and how they are processed inside cells. Herein we examine the intracellular deformation of layer-by-layer assembled polymer capsules using super-resolution structured illumination microscopy (SIM). Spherical- and cylindrical-shaped capsules were studied in three different cell lines, namely HeLa (human epithelial cell line), RAW264.7 (mouse macrophage cell line) and differentiated THP-1 (human monocyte-derived macrophage cell line). We observed that the deformation of capsules was dependent on cell line, but independent of capsule shape. This suggests that the mechanical forces, which induce capsule deformation during cell uptake, vary between cell lines, indicating that the capsules are exposed to higher mechanical forces in HeLa cells, followed by RAW264.7 and then differentiated THP-1 cells. Our study demonstrates the use of super-resolution SIM in analysing intracellular capsule deformation, offering important insights into the cellular processing of drug carriers in cells and providing fundamental knowledge of intracellular mechanobiology. Furthermore, this study may aid in the design of novel drug carriers that are sensitive to deformation for enhanced drug release properties.Understanding the behaviour of therapeutic carriers is important in elucidating their mechanism of action and how they are processed inside cells. Herein we examine the intracellular deformation of layer-by-layer assembled polymer capsules using super-resolution structured illumination microscopy (SIM). Spherical- and cylindrical-shaped capsules were studied in three different cell lines, namely HeLa (human epithelial cell line), RAW264.7 (mouse macrophage cell line) and differentiated THP-1 (human monocyte-derived macrophage cell line). We observed that the deformation of capsules was dependent on cell line, but independent of capsule shape. This suggests that the mechanical forces

  9. Genetic Heterogeneity and Identification of Its Physiological Property of Psychrotrophic Lactic Acid Bacteria from the Intestinal Tract of Cold-water Fishes from the Eerqisi River, Xinjiang%新疆额尔齐斯河流域冷水鱼肠道耐冷乳酸菌遗传分析及生理特征鉴定

    Institute of Scientific and Technical Information of China (English)

    李佳益; 孙海龙; 张敏; 蔺朋武; 倪永清

    2016-01-01

    目的:从新疆额尔齐斯河流域冷水鱼肠道中筛选耐冷乳酸菌,研究其遗传多样性,为低温保藏及其它技术研究提供优良菌株。方法:利用MRS、M17、LSA、ELLIKER、CQ五种培养基从8种冷水鱼肠道内分离筛选出耐冷菌株,鉴定菌株表型特征并对其理化性质进行测定。建立16S基因系统发育树探究菌株进化关系,在16S系统发育关系的基础上再进行rep PCR实验,使用引物为(GTG)5。结果:分离得到90株耐冷菌,最终确定有25株为耐低温乳酸菌,菌株的最适生长温度在28-32℃之间。系统发育分析结果表明:25株耐低温乳酸菌隶属于3个属,其中肠球属(Enterococcus)9株,乳杆属(Lactobacillus)15株,明串珠属(Leuconostoc)1株。结论:新疆额尔齐斯河流域冷水鱼肠道中分离的耐冷乳酸菌主要归为三个属,分离种属单一,需要进一步从冷水鱼肠道中大量分离采集菌株,继续探索研究其物种多样性和遗传多样性。%Objective:The purpose of this study was to select cold-adapted lactic acid bacteria (LAB) from the intestinal tract of cold-water fishes from the Eerqisi river, Xinjiang. The genetic diversity of lactic acid bacteria was studied, which provided the excellent species for low temperature preservation and technology research. Methods: By using culture medium MRS,M17,LSA,ELLIKER and CQ, isolation of lactic acid bacteria (LAB) from 8 intestinal tract of cold-water fishes was carried out. Taxonomic identity and genetic diversity of strains isolated were determined by partial 16S rRNA gene sequences and rep-PCR with primers named (GTG)5. Results: A total of 90 psychrotrophic isolates were obtained. Among them, 25 isolates had characteristics of LAB and showed the optimal growth temperature ranging from 28 to 32℃. The phylogenesis result showed that 25 strains belonged to 3 Genuses, Lactococcus (15strains), Enterococcus (9strains), Leuconostoc

  10. 南极嗜冷杆菌脂肪酶的原核可溶性表达优化及酶学性能表征%Optimized soluble expression and characterization of a cold-adapted lipase from psychrotrophic bacterium in Escherichia coli

    Institute of Scientific and Technical Information of China (English)

    刘弘忍; 王亮亮; 何琦阳; 王飞; 李迅

    2016-01-01

    Lipase is any enzyme that catalyzes the hydrolysis of lipids, and is the subclass of the esterases. Given its ex⁃cellent functions, lipase plays an indispensable role in many industrial areas such as food processing and biodiesel prepa⁃ration. In this paper, in order to effectively improve the ulilization of lipase from microbes, a psychrophilic lipase gene from antarctic psychrotrophic bacterium, Psychrobacter sp. 7195 ( Lip7195) was cloned into the commercial plasmid pTrc99a by the methods of polymerase chain reaction ( PCR) and DNA restriction enzyme digestion as well as ligation. The recombinant lipase was chemically induced in the presence of isopropyl β⁃D⁃Thiogalactoside ( IPTG) and heterolo⁃gously expressed in Escherichia coli. The proportion of the soluble recombinant lipase was significantly enhanced by a se⁃ries of modifications at various levels including optimizing codon usage, incorporating polycationic amino acid tags. The engineered recombinant lipase showed the maximum activities at 40℃ and pH 9.0. The specific activity was determined as 10.9 U/mg. The thermal stability assay exhibited that Lip7195 was fairly thermally stable at the temperatures ranging from 30 to 40℃, and at the pH values from 8.0 to 10.0. The recombinant Lip7195 was chemically activated in the pres⁃ence of metal ion Co2+ under the assay conditions. The results indicate that these modifications can effectively decrease the formation of inclusion body in the protein biosynthesis process in E. coli. Apart from the decrease in the inclusion body of the desired protein, the biochemical property and catalytic ability of recombinant Lip7195 can be intactly main⁃tained after the relevant modifications.%脂肪酶广泛应用于食品加工、生物柴油制备等领域。为了有效提高微生物脂肪酶的可利用度,将来源于南极嗜冷杆菌属( antarctic psychrotrophic bacterium, Psychrobacter sp�7195)的嗜冷脂肪酶( Lip7195)在大肠杆

  11. Determination of the sedimentation constants for total suspended solids and the algal component in a full-scale primary facultative pond operating at high wind velocities under tropical conditions.

    Science.gov (United States)

    Saraiva, L B; Ribeiro Meneses, C G; de Souza Melo, H N; Calado Araújo, A L; Pearson, H

    2005-01-01

    This study evaluated the amount, distribution and sedimentation constant of solids in a full-scale primary facultative pond operating mostly under high wind conditions and the contribution made by the algal biomass. Solids deposition rates were measured using sedimentation traps placed in the inlet and outlet zones of the pond. Most sludge accumulation occurred, not surprisingly, in the inlet zone A1 with a sludge volume of 9072.m3 accumulating over an operating time of approximately 3 years. However, sludge deposition within this zone was uneven and affected by wind action. Mean proportionality constant (K) values for solids sedimentation were 3.02 and 5.70 for depths of 50 cm and 100 cm respectively for A1. In contrast in zone A3, (the outlet zone), reduced K values of 1.38 and 3.22 were obtained for depths of 50 cm and 100 cm respectively. The algal sedimentation constant varied from 0.8 d(-1) in zone A1 to 0.02 d(-1) in A3. These data suggest that in this large facultative pond the wind, blowing predominantly from the direction of the outlets towards the pond inlets, had a greater influence on solids deposition than the bulk hydraulic flow and also kept the pond completely mixed for most of the time.

  12. Pico gauges for minimally invasive intracellular hydrostatic pressure measurements

    DEFF Research Database (Denmark)

    Knoblauch, Jan; Mullendore, Daniel L.; Jensen, Kaare Hartvig

    2014-01-01

    in the tip of microcapillaries, which we call pico gauges. The production of pico gauges can be accomplished with standard laboratory equipment, and measurements are comparably easy to conduct. Example pressure measurements are performed on cells that are difficult or impossible to measure with other methods.......Intracellular pressure has a multitude of functions in cells surrounded by a cell wall or similar matrix in all kingdoms of life. The functions include cell growth, nastic movements, and penetration of tissue by parasites. The precise measurement of intracellular pressure in the majority of cells......, however, remains difficult or impossible due to their small size and/or sensitivity to manipulation. Here, we report on a method that allows precise measurements in basically any cell type over all ranges of pressure. It is based on the compression of nanoliter and picoliter volumes of oil entrapped...

  13. Monitoring the intracellular calcium response to a dynamic hypertonic environment

    Science.gov (United States)

    Huang, Xiaowen; Yue, Wanqing; Liu, Dandan; Yue, Jianbo; Li, Jiaqian; Sun, Dong; Yang, Mengsu; Wang, Zuankai

    2016-03-01

    The profiling of physiological response of cells to external stimuli at the single cell level is of importance. Traditional approaches to study cell responses are often limited by ensemble measurement, which is challenging to reveal the complex single cell behaviors under a dynamic environment. Here we report the development of a simple microfluidic device to investigate intracellular calcium response to dynamic hypertonic conditions at the single cell level in real-time. Interestingly, a dramatic elevation in the intracellular calcium signaling is found in both suspension cells (human leukemic cell line, HL-60) and adherent cells (lung cancer cell line, A549), which is ascribed to the exposure of cells to the hydrodynamic stress. We also demonstrate that the calcium response exhibits distinct single cell heterogeneity as well as cell-type-dependent responses to the same stimuli. Our study opens up a new tool for tracking cellular activity at the single cell level in real time for high throughput drug screening.

  14. Regulation of dopamine transporter trafficking by intracellular amphetamine

    DEFF Research Database (Denmark)

    Kahlig, Kristopher M; Lute, Brandon J; Wei, Yuqiang

    2006-01-01

    -induced cell surface DAT redistribution may result in long-lasting changes in DA homeostasis. The molecular mechanism by which AMPH induces trafficking is not clear. Because AMPH is a substrate, we do not know whether extracellular AMPH stimulates trafficking through its interaction with DAT and subsequent...... alteration in DAT function, thereby triggering intracellular signaling or whether AMPH must be transported and then act intracellularly. In agreement with our previous studies, extracellular AMPH caused cytosolic redistribution of the wild-type human DAT (WT-hDAT). However, AMPH did not induce cytosolic...... redistribution in an uptake-impaired hDAT (Y335A-hDAT) that still binds AMPH. The divalent cation zinc (Zn(2+)) inhibits WT-hDAT activity, but it restores Y335A-hDAT uptake. Coadministration of Zn(2+) and AMPH consistently reduced WT-hDAT trafficking but stimulated cytosolic redistribution of Y335A...

  15. Intracellular transport proteins: classification, structure and function of kinesins

    Directory of Open Access Journals (Sweden)

    Agnieszka Chudy

    2011-09-01

    Full Text Available Correct cell functioning, division and morphogenesis rely on efficient intracellular transport. Apart from dyneins and myosins, kinesins are the main proteins responsible for intracellular movement. Kinesins are a large, diverse group of motor proteins, which based on phylogenetic similarity were classified into fourteen families. Among these families, due to the location of their motor domains, three groups have been characterized: N-, C- and M-kinesin. As molecular motors, kinesins transport various molecules and vesicles mainly towards the microtubule plus end (from the cell body participating in anterograde transport, although there are also kinesins involved in retrograde transport (C-kinesins. Kinesins are also involved in spindle formation, chromosome segregation, and spermatogenesis. Because of their great importance for the correct functioning of cells, mutations in kinesin coding genes may lead to such neurodegenerative diseases as dominant hereditary spastic paraplegia or Charcot-Marie-Tooth disease.

  16. Characterization of a Mycobacterium intracellulare Variant Strain by Molecular Techniques

    Science.gov (United States)

    Menendez, M. C.; Palenque, E.; Navarro, M. C.; Nuñez, M. C.; Rebollo, M. J.; Garcia, M. J.

    2001-01-01

    This paper describes a Mycobacterium intracellulare variant strain causing an unusual infection. Several isolates obtained from an immunocompromised patient were identified as members of the Mycobacterium avium complex (MAC) by the commercial AccuProbe system and biochemical standard identification. Further molecular approaches were undertaken for a more accurate characterization of the bacteria. Up to seven different genomic sequences were analyzed, ranging from conserved mycobacterial genes such as 16S ribosomal DNA to MAC-specific genes such as mig (macrophage-induced gene). The results obtained identify the isolates as a variant of M. intracellulare, an example of the internal variability described for members of the MAC, particularly within that species. The application of other molecular approaches is recommended for more accurate identification of bacteria described as MAC members. PMID:11724827

  17. Non-contact intracellular binding of chloroplasts in vivo

    Science.gov (United States)

    Li, Yuchao; Xin, Hongbao; Liu, Xiaoshuai; Li, Baojun

    2015-06-01

    Non-contact intracellular binding and controllable manipulation of chloroplasts in vivo was demonstrated using an optical fiber probe. Launching a 980-nm laser beam into a fiber, which was placed about 3 μm above the surface of a living plant (Hydrilla verticillata) leaf, enabled stable binding of different numbers of chloroplasts, as well as their arrangement into one-dimensional chains and two-dimensional arrays inside the leaf without damaging the chloroplasts. Additionally, the formed chloroplast chains were controllably transported inside the living cells. The optical force exerted on the chloroplasts was calculated to explain the experimental results. This method provides a flexible method for studying intracellular organelle interaction with highly organized organelle-organelle contact in vivo in a non-contact manner.

  18. Intracellular and extracellular Abeta, a tale of two neuropathologies.

    Science.gov (United States)

    Cuello, A Claudio

    2005-01-01

    The central pathological cause of Alzheimer disease (AD) is hypothesized to be an excess of beta-amyloid (Abeta) which accumulates into toxic fibrillar deposits within extracellular areas of the brain. These deposits disrupt neural and synaptic function and ultimately lead to neuronal degeneration and dementia. In addition to the pathological roles attributed to Abeta, evidence from our laboratory would suggest that Abeta serves a physiological role in the modulation of CRE-directed gene expression. This commentary also highlights some of the pathological consequences of the accumulation of intracellular Abeta. Finally it discusses the impact of cortical Abeta burden on transmitter-specific synaptic numbers as well as the generation of dystrophic neurites. The fundamental thesis of my proposal is that the Abeta pathology seen in AD is a continuous process from an initial abnormal Abeta intracellular accumulation to the well-established extracellular Abeta aggregation, culminating in the formation of amyloid plaques and dystrophic neurites.

  19. Pico gauges for minimally invasive intracellular hydrostatic pressure measurements.

    Science.gov (United States)

    Knoblauch, Jan; Mullendore, Daniel L; Jensen, Kaare H; Knoblauch, Michael

    2014-11-01

    Intracellular pressure has a multitude of functions in cells surrounded by a cell wall or similar matrix in all kingdoms of life. The functions include cell growth, nastic movements, and penetration of tissue by parasites. The precise measurement of intracellular pressure in the majority of cells, however, remains difficult or impossible due to their small size and/or sensitivity to manipulation. Here, we report on a method that allows precise measurements in basically any cell type over all ranges of pressure. It is based on the compression of nanoliter and picoliter volumes of oil entrapped in the tip of microcapillaries, which we call pico gauges. The production of pico gauges can be accomplished with standard laboratory equipment, and measurements are comparably easy to conduct. Example pressure measurements are performed on cells that are difficult or impossible to measure with other methods.

  20. An Intracellular Calcium Oscillations Model Including Mitochondrial Calcium Cycling

    Institute of Scientific and Technical Information of China (English)

    SHI Xiao-Min; LIU Zeng-Rong

    2005-01-01

    @@ Calcium is a ubiquitous second messenger. Mitochondria contributes significantly to intracellular Ca2+ dynamics.The experiment of Kaftan et al. [J. Biol. Chem. 275(2000) 25465] demonstrated that inhibiting mitochondrial Ca2+ uptake can reduce the frequency of cytosolic Ca2+ concentration oscillations of gonadotropes. By considering the mitochondrial Ca2+ cycling we develop a three-variable model of intracellular Ca2+ oscillations based on the models of Atri et al. [Biophys. J. 65 (1993) 1727] and Falcke et al. [Biophys. J. 77 (1999) 37]. The model reproduces the fact that mitochondrial Ca2+ cycling increases the frequency of cytosolic Ca2+ oscillations, which accords with Kaftan's results. Moreover the model predicts that when the mitochondria overload with Ca2+, the cytosolic Ca2+ oscillations vanish, which may trigger apoptosis.

  1. Intracellular transport driven by cytoskeletal motors: General mechanisms and defects

    CERN Document Server

    Appert-Rolland, Cecile; Santen, Ludger

    2015-01-01

    Cells are strongly out-of-equilibrium systems driven by continuous energy supply. They carry out many vital functions requiring active transport of various ingredients and organelles, some being small, others being large. The cytoskeleton, composed of three types of filaments, determines the shape of the cell and plays a role in cell motion. It also serves as a road network for the so-called cytoskeletal motors. These molecules can attach to a cytoskeletal filament, perform directed motion, possibly carrying along some cargo, and then detach. It is a central issue to understand how intracellular transport driven by molecular motors is regulated, in particular because its breakdown is one of the signatures of some neuronal diseases like the Alzheimer. We give a survey of the current knowledge on microtubule based intracellular transport. We first review some biological facts obtained from experiments, and present some modeling attempts based on cellular automata. We start with background knowledge on the origi...

  2. Intracellular compartmentalization of skeletal muscle glycogen metabolism and insulin signalling

    DEFF Research Database (Denmark)

    Prats Gavalda, Clara; Gomez-Cabello, Alba; Vigelsø Hansen, Andreas

    2011-01-01

    compartmentalization in the regulation of skeletal muscle glycogen metabolism and insulin signalling. As a result, a hypothetical regulatory mechanism is proposed by which cells could direct glycogen resynthesis towards different pools of glycogen particles depending on the metabolic needs. Furthermore, we discuss......The interest in skeletal muscle metabolism and insulin signalling has increased exponentially in recent years as a consequence of their role in the development of type 2 diabetes mellitus. Despite this, the exact mechanisms involved in the regulation of skeletal muscle glycogen metabolism...... and insulin signalling transduction remain elusive. We believe that one of the reasons is that the role of intracellular compartmentalization as a regulator of metabolic pathways and signalling transduction has been rather ignored. This paper briefly reviews the literature to discuss the role of intracellular...

  3. Characterization of a Mycobacterium intracellulare variant strain by molecular techniques.

    Science.gov (United States)

    Menendez, M C; Palenque, E; Navarro, M C; Nuñez, M C; Rebollo, M J; Garcia, M J

    2001-12-01

    This paper describes a Mycobacterium intracellulare variant strain causing an unusual infection. Several isolates obtained from an immunocompromised patient were identified as members of the Mycobacterium avium complex (MAC) by the commercial AccuProbe system and biochemical standard identification. Further molecular approaches were undertaken for a more accurate characterization of the bacteria. Up to seven different genomic sequences were analyzed, ranging from conserved mycobacterial genes such as 16S ribosomal DNA to MAC-specific genes such as mig (macrophage-induced gene). The results obtained identify the isolates as a variant of M. intracellulare, an example of the internal variability described for members of the MAC, particularly within that species. The application of other molecular approaches is recommended for more accurate identification of bacteria described as MAC members.

  4. Intracellular Detection of Viral Transcription and Replication Using RNA FISH

    Science.gov (United States)

    2016-05-26

    Chapter 14. Intracellular detection of viral transcription and replication using RNA FISH i. Summary/Abstract Many hemorrhagic fever viruses...examine the mechanisms in which viruses replicate, assemble, and traffic through the cell. An additional benefit of this method is that the robust...Visualization of single RNA transcripts in situ. Science, 1998. 280(5363): p. 585-90. 4. Jambo, K.C., et al., Small alveolar macrophages are infected

  5. Novel intracellular proteins associated with cellular vitamin D action.

    Science.gov (United States)

    Angelo, Giana; Wood, Richard J; Mayer, Jean

    2002-07-01

    Work with vitamin D-resistant New World primates has revealed novel cellular proteins involved in vitamin D action. An "intracellular vitamin D-binding protein" functions to bind vitamin D metabolites in the cell and enhances vitamin D action. By contrast, a "vitamin D response element-binding protein" inhibits vitamin D receptor binding to the DNA and is responsible for vitamin D resistance in New World primates.

  6. Intracellular messengers in the generation and degeneration of hippocampal neuroarchitecture.

    Science.gov (United States)

    Mattson, M P; Guthrie, P B; Kater, S B

    1988-01-01

    The actions and interactions of the neurotransmitter glutamate and the intracellular messengers calcium, cyclic AMP, and protein kinase C (PKC) in the regulation of neurite outgrowth and cell survival were examined in hippocampal pyramidal-like neurons in isolated cell culture. Low, subtoxic levels of glutamate (10-100 microM) caused the regression of dendrites but not axons; millimolar levels caused cell death. Calcium ionophore A23187 (50-100 nM) and the PKC activator phorbol-12-myristate-13-acetate (PMA; 10-50 nM) caused the regression of both axons and dendrites, whereas the adenylate cyclase activator forskolin enhanced outgrowth rates in both axons and dendrites. The effects of glutamate, A23187, PMA, and forskolin on outgrowth were mediated locally at the growth cones; dendrites were more sensitive than axons to each of these agents. High levels of A23187 (1 microM) or PMA (100 nM) significantly reduced cell survival. Co2+ and trifluoperazine each significantly reduced glutamate-induced dendritic regression and neurotoxicity suggesting that calcium influx and/or PKC activation mediated glutamate's actions. Fura-2 measurements showed that glutamate caused a rapid rise in intracellular calcium levels; this rise was prevented by Co2+. PMA and forskolin did not alter intracellular calcium levels, nor did these agents affect glutamate-induced calcium rises. Taken together, the results indicate that parallel intracellular messenger pathways that influence neurite outgrowth and cell survival are operative in hippocampal neurons; these messengers may play roles in the formation and modification of neuronal circuitry.

  7. Mycobacterium intracellulare infection in a capybara (Hydrochoerus hydrochaeris).

    Science.gov (United States)

    Pezzone, Natalia; Eberhardt, Ayelen T; Fernández, Analia; Garbaccio, Sergio; Zumárraga, Martín; Gioffré, Andrea; Magni, Carolina; Beldomenico, Pablo M; Marini, M Rocío; Canal, Ana M

    2013-12-01

    This report describes the first case of Mycobacterium intracellulare infection with typical granulomatous lesions of mycobacteriosis in a capybara (Hydrochoerus hydrochaeris). The individual was a captive-bred young female, part of the control group of an experimental study on stress. Multiple granulomatous lesions were detected in a mesenteric lymph node of this young female. Mycobacterial infection was confirmed by bacteriologic culture and molecular identification methods. Clinical lesions were characterized by histopathology.

  8. Efficient intracellular retrotransposition of an exogenous primate retrovirus genome

    OpenAIRE

    Heinkelein, Martin; Pietschmann, Thomas; Jármy, Gergely; Dressler, Marco; Imrich, Horst; Thurow, Jana; Lindemann, Dirk,; Bock, Michael; Moebes, Astrid; Roy, Jacqueline; Herchenröder, Ottmar; Rethwilm, Axel

    2000-01-01

    The foamy virus (FV) subgroup of Retroviridae reverse transcribe their RNA (pre-)genome late in the replication cycle before leaving an infected cell. We studied whether a marker gene-transducing FV vector is able to shuttle to the nucleus and integrate into host cell genomic DNA. While a potential intracellular retrotransposition of vectors derived from other retroviruses was below the detection limit of our assay, we found that up to 5% of cells transfected with the FV vector were stably tr...

  9. Mycobacterium avium-intracellulare: a rare cause of subacromial bursitis.

    Science.gov (United States)

    Sinha, Raj; Tuckett, John; Hide, Geoff; Dildey, Petra; Karsandas, Alvin

    2015-01-01

    Septic subacromial bursitis is an uncommon disorder with only a few reported cases in the literature. The most common causative organism is Staphylococcus aureus. We report the case of a 61-year-old female with a septic subacromial bursitis where the causative organism was found to be Mycobacterium avium-intracellulare (MAI). The diagnosis was only made following a biopsy, and we use this case to highlight the importance of recognising the need to consider a biopsy and aspiration in atypical situations.

  10. Effect of mycobacteriophage to intracellular mycobateria in vitro

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ Mycobacterium tuberculosis (M. tuberculosis) is a pathogen associated with the deaths of millions of people worldwide annually. Although tuberculosis is a treatable disease, the emergence and increasing prevalence of multi-drug resistant bacterial pathogens emphasizes the need for new and innovative anti-microbial strategies. Effective therapeutic regimens, which have existed, are limited by the emergence of drug resistance and the inability of antibiotics to kill dormant or intracellular organisms.

  11. ApoHRP-based Assay to Measure Intracellular Regulatory Heme

    Science.gov (United States)

    Atamna, Hani; Brahmbhatt, Marmik; Atamna, Wafa; Shanower, Gregory A.; Dhahbi, Joseph M.

    2015-01-01

    The majority of the heme-binding proteins possess a “heme-pocket” that stably binds with heme. Usually known as housekeeping heme-proteins, they participate in a variety of metabolic reactions (e.g., catalase). Heme also binds with lower affinity to the “Heme-Regulatory Motifs” (HRM) in specific regulatory proteins. This type of heme binding is known as exchangeable or regulatory heme (RH). Heme binding to HRM proteins regulates their function (e.g., Bach1). Although there are well-established methods for assaying total cellular heme (e.g., heme-proteins plus RH), currently there is no method available for measuring RH independently from the total heme (TH). The current study describes and validates a new method to measure intracellular RH. The method is based on the reconstitution of apo-horseradish peroxidase (apoHRP) with heme to form holoHRP. The resulting holoHRP activity is then measured with a colorimetric substrate. The results show that apoHRP specifically binds RH but not with heme from housekeeping heme-proteins. The RH assay detects intracellular RH. Furthermore, using conditions that create positive (hemin) or negative (N-methyl protoporphyrin IX) controls for heme in normal human fibroblasts (IMR90), the RH assay shows that RH is dynamic and independent from TH. We also demonstrated that short-term exposure to subcytotoxic concentrations of lead (Pb), mercury (Hg), or amyloid-β(Aβ) significantly alters intracellular RH with little effect on TH. In conclusion the RH assay is an effective assay to investigate intracellular RH concentration and demonstrates that RH represents ~6% of total heme in IMR90 cells. PMID:25525887

  12. Increasing intracellular bioavailable copper selectively targets prostate cancer cells.

    Science.gov (United States)

    Cater, Michael A; Pearson, Helen B; Wolyniec, Kamil; Klaver, Paul; Bilandzic, Maree; Paterson, Brett M; Bush, Ashley I; Humbert, Patrick O; La Fontaine, Sharon; Donnelly, Paul S; Haupt, Ygal

    2013-07-19

    The therapeutic efficacy of two bis(thiosemicarbazonato) copper complexes, glyoxalbis[N4-methylthiosemicarbazonato]Cu(II) [Cu(II)(gtsm)] and diacetylbis[N4-methylthiosemicarbazonato]Cu(II) [Cu(II)(atsm)], for the treatment of prostate cancer was assessed in cell culture and animal models. Distinctively, copper dissociates intracellularly from Cu(II)(gtsm) but is retained by Cu(II)(atsm). We further demonstrated that intracellular H2gtsm [reduced Cu(II)(gtsm)] continues to redistribute copper into a bioavailable (exchangeable) pool. Both Cu(II)(gtsm) and Cu(II)(atsm) selectively kill transformed (hyperplastic and carcinoma) prostate cell lines but, importantly, do not affect the viability of primary prostate epithelial cells. Increasing extracellular copper concentrations enhanced the therapeutic capacity of both Cu(II)(gtsm) and Cu(II)(atsm), and their ligands (H2gtsm and H2atsm) were toxic only toward cancerous prostate cells when combined with copper. Treatment of the Transgenic Adenocarcinoma of Mouse Prostate (TRAMP) model with Cu(II)(gtsm) (2.5 mg/kg) significantly reduced prostate cancer burden (∼70%) and severity (grade), while treatment with Cu(II)(atsm) (30 mg/kg) was ineffective at the given dose. However, Cu(II)(gtsm) caused mild kidney toxicity in the mice, associated primarily with interstitial nephritis and luminal distention. Mechanistically, we demonstrated that Cu(II)(gtsm) inhibits proteasomal chymotrypsin-like activity, a feature further established as being common to copper-ionophores that increase intracellular bioavailable copper. We have demonstrated that increasing intracellular bioavailable copper can selectively kill cancerous prostate cells in vitro and in vivo and have revealed the potential for bis(thiosemicarbazone) copper complexes to be developed as therapeutics for prostate cancer.

  13. Molecular evolution, intracellular organization, and the quinary structure of proteins.

    OpenAIRE

    McConkey, E H

    1982-01-01

    High-resolution two-dimensional polyacrylamide gel electrophoresis shows that at least half of 370 denatured polypeptides from hamster cells and human cells are indistinguishable in terms of isoelectric points and molecular weights. Molecular evolution may have been more conservative for this set of proteins than sequence studies on soluble proteins have implied. This may be a consequence of complexities of intracellular organization and the numerous macromolecular interactions in which most ...

  14. Dual Readout BRET/FRET Sensors for Measuring Intracellular Zinc

    Science.gov (United States)

    2016-01-01

    Genetically encoded FRET-based sensor proteins have significantly contributed to our current understanding of the intracellular functions of Zn2+. However, the external excitation required for these fluorescent sensors can give rise to photobleaching and phototoxicity during long-term imaging, limits applications that suffer from autofluorescence and light scattering, and is not compatible with light-sensitive cells. For these applications, sensor proteins based on Bioluminescence Resonance Energy Transfer (BRET) would provide an attractive alternative. In this work, we used the bright and stable luciferase NanoLuc to create the first genetically encoded BRET sensors for measuring intracellular Zn2+. Using a new sensor approach, the NanoLuc domain was fused to the Cerulean donor domain of two previously developed FRET sensors, eCALWY and eZinCh-2. In addition to preserving the excellent Zn2+ affinity and specificity of their predecessors, these newly developed sensors enable both BRET- and FRET-based detection. While the dynamic range of the BRET signal for the eCALWY-based BLCALWY-1 sensor was limited by the presence of two competing BRET pathways, BRET/FRET sensors based on the eZinCh-2 scaffold (BLZinCh-1 and -2) yielded robust 25–30% changes in BRET ratio. In addition, introduction of a chromophore-silencing mutation resulted in a BRET-only sensor (BLZinCh-3) with increased BRET response (50%) and an unexpected 10-fold increase in Zn2+ affinity. The combination of robust ratiometric response, physiologically relevant Zn2+ affinities, and stable and bright luminescence signal offered by the BLZinCh sensors allowed monitoring of intracellular Zn2+ in plate-based assays as well as intracellular BRET-based imaging in single living cells in real time. PMID:27547982

  15. NAD+-Glycohydrolase Promotes Intracellular Survival of Group A Streptococcus.

    Directory of Open Access Journals (Sweden)

    Onkar Sharma

    2016-03-01

    Full Text Available A global increase in invasive infections due to group A Streptococcus (S. pyogenes or GAS has been observed since the 1980s, associated with emergence of a clonal group of strains of the M1T1 serotype. Among other virulence attributes, the M1T1 clone secretes NAD+-glycohydrolase (NADase. When GAS binds to epithelial cells in vitro, NADase is translocated into the cytosol in a process mediated by streptolysin O (SLO, and expression of these two toxins is associated with enhanced GAS intracellular survival. Because SLO is required for NADase translocation, it has been difficult to distinguish pathogenic effects of NADase from those of SLO. To resolve the effects of the two proteins, we made use of anthrax toxin as an alternative means to deliver NADase to host cells, independently of SLO. We developed a novel method for purification of enzymatically active NADase fused to an amino-terminal fragment of anthrax toxin lethal factor (LFn-NADase that exploits the avid, reversible binding of NADase to its endogenous inhibitor. LFn-NADase was translocated across a synthetic lipid bilayer in vitro in the presence of anthrax toxin protective antigen in a pH-dependent manner. Exposure of human oropharyngeal keratinocytes to LFn-NADase in the presence of protective antigen resulted in cytosolic delivery of NADase activity, inhibition of protein synthesis, and cell death, whereas a similar construct of an enzymatically inactive point mutant had no effect. Anthrax toxin-mediated delivery of NADase in an amount comparable to that observed during in vitro infection with live GAS rescued the defective intracellular survival of NADase-deficient GAS and increased the survival of SLO-deficient GAS. Confocal microscopy demonstrated that delivery of LFn-NADase prevented intracellular trafficking of NADase-deficient GAS to lysosomes. We conclude that NADase mediates cytotoxicity and promotes intracellular survival of GAS in host cells.

  16. Forced resurgence and targeting of intracellular uropathogenic Escherichia coli reservoirs.

    Directory of Open Access Journals (Sweden)

    Matthew G Blango

    Full Text Available Intracellular quiescent reservoirs of uropathogenic Escherichia coli (UPEC, which can seed the bladder mucosa during the acute phase of a urinary tract infection (UTI, are protected from antibiotic treatments and are extremely difficult to eliminate. These reservoirs are a potential source for recurrent UTIs that affect millions annually. Here, using murine infection models and the bladder cell exfoliant chitosan, we demonstrate that intracellular UPEC populations shift within the stratified layers of the urothelium during the course of a UTI. Following invasion of the terminally differentiated superficial layer of epithelial cells that line the bladder lumen, UPEC can multiply and disseminate, eventually establishing reservoirs within underlying immature host cells. If given access, UPEC can invade the superficial and immature bladder cells equally well. As infected immature host cells differentiate and migrate towards the apical surface of the bladder, UPEC can reinitiate growth and discharge into the bladder lumen. By inducing the exfoliation of the superficial layers of the urothelium, chitosan stimulates rapid regenerative processes and the reactivation and efflux of quiescent intracellular UPEC reservoirs. When combined with antibiotics, chitosan treatment significantly reduces bacterial loads within the bladder and may therefore be of therapeutic value to individuals with chronic, recurrent UTIs.

  17. Intracellular Trafficking Network of Protein Nanocapsules: Endocytosis, Exocytosis and Autophagy

    Science.gov (United States)

    Zhang, Jinxie; Zhang, Xudong; Liu, Gan; Chang, Danfeng; Liang, Xin; Zhu, Xianbing; Tao, Wei; Mei, Lin

    2016-01-01

    The inner membrane vesicle system is a complex transport system that includes endocytosis, exocytosis and autophagy. However, the details of the intracellular trafficking pathway of nanoparticles in cells have been poorly investigated. Here, we investigate in detail the intracellular trafficking pathway of protein nanocapsules using more than 30 Rab proteins as markers of multiple trafficking vesicles in endocytosis, exocytosis and autophagy. We observed that FITC-labeled protein nanoparticles were internalized by the cells mainly through Arf6-dependent endocytosis and Rab34-mediated micropinocytosis. In addition to this classic pathway: early endosome (EEs)/late endosome (LEs) to lysosome, we identified two novel transport pathways: micropinocytosis (Rab34 positive)-LEs (Rab7 positive)-lysosome pathway and EEs-liposome (Rab18 positive)-lysosome pathway. Moreover, the cells use slow endocytosis recycling pathway (Rab11 and Rab35 positive vesicles) and GLUT4 exocytosis vesicles (Rab8 and Rab10 positive) transport the protein nanocapsules out of the cells. In addition, protein nanoparticles are observed in autophagosomes, which receive protein nanocapsules through multiple endocytosis vesicles. Using autophagy inhibitor to block these transport pathways could prevent the degradation of nanoparticles through lysosomes. Using Rab proteins as vesicle markers to investigation the detail intracellular trafficking of the protein nanocapsules, will provide new targets to interfere the cellular behaver of the nanoparticles, and improve the therapeutic effect of nanomedicine. PMID:27698943

  18. Modulation of lipoprotein receptor functions by intracellular adaptor proteins.

    Science.gov (United States)

    Stolt, Peggy C; Bock, Hans H

    2006-10-01

    Members of the low density lipoprotein (LDL) receptor gene family are critically involved in a wide range of physiological processes including lipid and vitamin homeostasis, cellular migration, neurodevelopment, and synaptic plasticity, to name a few. Lipoprotein receptors exert these diverse biological functions by acting as cellular uptake receptors or by inducing intracellular signaling cascades. It was discovered that a short sequence in the intracellular region of all lipoprotein receptors, Asn-Pro-X-Tyr (NPXY) is important for mediating either endocytosis or signal transduction events, and that this motif serves as a binding site for phosphotyrosine-binding (PTB) domain containing scaffold proteins. These molecular adaptors connect the transmembrane receptors with the endocytosis machinery and regulate cellular trafficking, or function as assembly sites for dynamic multi-protein signaling complexes. Whereas the LDL receptor represents the archetype of an endocytic lipoprotein receptor, the structurally closely related apolipoprotein E receptor 2 (apoER2) and very low density lipoprotein (VLDL) receptor activate a kinase-dependent intracellular signaling cascade after binding to the neuronal signaling molecule Reelin. This review focuses on two related PTB domain containing adaptor proteins that mediate these divergent lipoprotein receptor responses, ARH (autosomal recessive hypercholesterolemia protein) and Dab1 (disabled-1), and discusses the structural and molecular basis of this different behaviour.

  19. Fatty Acid Signaling: The New Function of Intracellular Lipases

    Directory of Open Access Journals (Sweden)

    Zuzana Papackova

    2015-02-01

    Full Text Available Until recently, intracellular triacylglycerols (TAG stored in the form of cytoplasmic lipid droplets have been considered to be only passive “energy conserves”. Nevertheless, degradation of TAG gives rise to a pleiotropic spectrum of bioactive intermediates, which may function as potent co-factors of transcription factors or enzymes and contribute to the regulation of numerous cellular processes. From this point of view, the process of lipolysis not only provides energy-rich equivalents but also acquires a new regulatory function. In this review, we will concentrate on the role that fatty acids liberated from intracellular TAG stores play as signaling molecules. The first part provides an overview of the transcription factors, which are regulated by fatty acids derived from intracellular stores. The second part is devoted to the role of fatty acid signaling in different organs/tissues. The specific contribution of free fatty acids released by particular lipases, hormone-sensitive lipase, adipose triacylglycerol lipase and lysosomal lipase will also be discussed.

  20. A first step toward liposome-mediated intracellular bacteriophage therapy.

    Science.gov (United States)

    Nieth, Anita; Verseux, Cyprien; Barnert, Sabine; Süss, Regine; Römer, Winfried

    2015-01-01

    The emergence of antibiotic-resistant bacteria presents a severe challenge to medicine and public health. While bacteriophage therapy is a promising alternative to traditional antibiotics, the general inability of bacteriophages to penetrate eukaryotic cells limits their use against resistant bacteria, causing intracellular diseases like tuberculosis. Bacterial vectors show some promise in carrying therapeutic bacteriophages into cells, but also bring a number of risks like an overload of bacterial antigens or the acquisition of virulence genes from the pathogen. As a first step in the development of a non-bacterial vector for bacteriophage delivery into pathogen-infected cells, we attempted to encapsulate bacteriophages into liposomes. Here we report effective encapsulation of the model bacteriophage λeyfp and the mycobacteriophage TM4 into giant liposomes. Furthermore, we show that liposome-associated bacteriophages are taken up into eukaryotic cells more efficiently than free bacteriophages. These are important milestones in the development of an intracellular bacteriophage therapy that might be useful in the fight against multi-drug-resistant intracellular pathogens like Mycobacterium tuberculosis.

  1. Facultative Apomixis in an Endangered Dioecious Species,Woonyoungia septentrionalis (Magnoliaceae)%濒危植物焕镛木的兼性无融合生殖

    Institute of Scientific and Technical Information of China (English)

    曾庆文; 张奠湘; 高泽正; 邢福武

    2003-01-01

    germinate into seedlings. Therefore, the breeding system of W.septentrionalis belonged to facultative apomixis. The apomixis in Magnoliaceae was reported for the first time.

  2. Intracellular diffusion restrictions in isolated cardiomyocytes from rainbow trout

    Directory of Open Access Journals (Sweden)

    Birkedal Rikke

    2009-12-01

    Full Text Available Abstract Background Restriction of intracellular diffusion of adenine nucleotides has been studied intensively on adult rat cardiomyocytes. However, their cause and role in vivo is still uncertain. Intracellular membrane structures have been suggested to play a role. We therefore chose to study cardiomyocytes from rainbow trout (Oncorhynchus mykiss, which are thinner and have fewer intracellular membrane structures than adult rat cardiomyocytes. Previous studies suggest that trout permeabilized cardiac fibers also have diffusion restrictions. However, results from fibers may be affected by incomplete separation of the cells. This is avoided when studying permeabilized, isolated cardiomyocytes. The aim of this study was to verify the existence of diffusion restrictions in trout cardiomyocytes by comparing ADP-kinetics of mitochondrial respiration in permeabilized fibers, permeabilized cardiomyocytes and isolated mitochondria from rainbow trout heart. Experiments were performed at 10, 15 and 20°C in the absence and presence of creatine. Results Trout cardiomyocytes hypercontracted in the solutions used for mammalian cardiomyocytes. We developed a new solution in which they retained their shape and showed stable steady state respiration rates throughout an experiment. The apparent ADP-affinity of permeabilized cardiomyocytes was different from that of fibers. It was higher, independent of temperature and not increased by creatine. However, it was still about ten times lower than in isolated mitochondria. Conclusions The differences between fibers and cardiomyocytes suggest that results from trout heart fibers were affected by incomplete separation of the cells. However, the lower ADP-affinity of cardiomyocytes compared to isolated mitochondria indicate that intracellular diffusion restrictions are still present in trout cardiomyocytes despite their lower density of intracellular membrane structures. The lack of a creatine effect indicates that

  3. Intracellular angiotensin II elicits Ca2+ increases in A7r5 vascular smooth muscle cells

    NARCIS (Netherlands)

    Filipeanu, CM; Brailoiu, E; Kok, JW; Henning, RH; De Zeeuw, D; Nelemans, SA

    2001-01-01

    Recent studies show that angiotensin II can act within the cell, possibly via intracellular receptors pharmacologically different from typical plasma membrane angiotensin II receptors. The signal transduction of intracellular angiotensin LI is unclear. Therefore. we investigated the effects of intra

  4. DMPD: Intracellular DNA sensors in immunity. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18573338 Intracellular DNA sensors in immunity. Takeshita F, Ishii KJ. Curr Opin Im...munol. 2008 Aug;20(4):383-8. Epub 2008 Jun 23. (.png) (.svg) (.html) (.csml) Show Intracellular DNA sensors ...in immunity. PubmedID 18573338 Title Intracellular DNA sensors in immunity. Authors Takeshita F, Ishii KJ. P

  5. DMPD: Intracellular TLR signaling: a structural perspective on human disease. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 16785490 Intracellular TLR signaling: a structural perspective on human disease. La...sker MV, Nair SK. J Immunol. 2006 Jul 1;177(1):11-6. (.png) (.svg) (.html) (.csml) Show Intracellular TLR si...gnaling: a structural perspective on human disease. PubmedID 16785490 Title Intracellular TLR signaling: a s

  6. DMPD: Intracellular NOD-like receptors in host defense and disease. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17967410 Intracellular NOD-like receptors in host defense and disease. Kanneganti T...D, Lamkanfi M, Nunez G. Immunity. 2007 Oct;27(4):549-59. (.png) (.svg) (.html) (.csml) Show Intracellular NO...D-like receptors in host defense and disease. PubmedID 17967410 Title Intracellular NOD-like receptors in ho

  7. DMPD: NOD-like receptors (NLRs): bona fide intracellular microbial sensors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18585455 NOD-like receptors (NLRs): bona fide intracellular microbial sensors. Shaw...tml) (.csml) Show NOD-like receptors (NLRs): bona fide intracellular microbial sensors. PubmedID 18585455 Ti...tle NOD-like receptors (NLRs): bona fide intracellular microbial sensors. Authors

  8. Effect of roscovitine on intracellular calcium dynamics: differential enantioselective responses.

    Science.gov (United States)

    Tamma, Grazia; Ranieri, Marianna; Di Mise, Annarita; Spirlì, Alessia; Russo, Annamaria; Svelto, Maria; Valenti, Giovanna

    2013-12-02

    Cyclin-dependent kinases (CDKs) inhibitors have emerged as interesting therapeutic candidates. Of these, (S)-roscovitine has been proposed as potential neuroprotective molecule for stroke while (R)-roscovitine is currently entering phase II clinical trials against cancers and phase I clinical tests against glomerulonephritis. In addition, (R)-roscovitine has been suggested as potential antihypertensive and anti-inflammatory drug. Dysfunction of intracellular calcium balance is a common denominator of these diseases, and the two roscovitine enantiomers (S and R) are known to modulate calcium voltage channel activity differentially. Here, we provide a detailed description of short- and long-term responses of roscovitine on intracellular calcium handling in renal epithelial cells. Short-term exposure to (S)-roscovitine induced a cytosolic calcium peak, which was abolished after stores depletion with cyclopiazonic acid (CPA). Instead, (R)-roscovitine caused a calcium peak followed by a small calcium plateau. Cytosolic calcium response was prevented after stores depletion. Bafilomycin, a selective vacuolar H(+)-ATPase inhibitor, abolished the small calcium plateau. Long-term exposure to (R)-roscovitine significantly reduced the basal calcium level compared to control and (S)-roscovitine treated cells. However, both enantiomers increased calcium accumulation in the endoplasmic reticulum (ER). Consistently, cells treated with (R)-roscovitine showed a significant increase in SERCA activity, whereas (S)-roscovitine incubation resulted in a reduced PMCA expression. We also found a tonic decreased ability to release calcium from the ER, likely via IP3 signaling, under treatment with (S)- or (R)-roscovitine. Together our data revealed that (S)-roscovitine and (R)-roscovitine exert distinct enantiospecific effects on intracellular calcium signaling in renal epithelial cells. This distinct pharmacological profile can be relevant for roscovitine clinical use.

  9. Intracellular lipophilic inclusions of mycobacteria in vitro and in sputum.

    Science.gov (United States)

    Garton, Natalie J; Christensen, Henriette; Minnikin, David E; Adegbola, Richard A; Barer, Michael R

    2002-10-01

    Although most mycobacterial lipids are thought to be associated with the cell envelope, the authors previously observed substantial deposits of intracellular lipophilic material. A Nile-red-based cytological assay was used to determine factors which affect the presence and natural history of intracellular lipophilic inclusions (ILIs) in Mycobacterium smegmatis. Development of ILIs was associated with stationary-phase cultures in broth and with aged (6 days) colonies on agar. Using variants of Youmans' defined medium, the frequency and size of ILIs was observed to be minimal in carbon-poor medium. ILIs were observed to form within 15 min after provision of fatty acids to the medium and after a period of several days in nitrogen-poor medium. Analysis of the non-polar lipid extracts of ILI-rich and -poor preparations indicated that the triacylglycerols (TAGs) were a major component of the inclusions. The acyl substituents of the TAGs varied according to whether they were formed in Middlebrook 7H9 broth, in low-nitrogen Youmans' broth or rapidly after oleic acid supplementation of Youmans' broth. These studies support a storage function for TAGs in mycobacteria in addition to their previously suggested occurrence as components of the cell envelope. To assess a possible role for ILIs in Mycobacterium tuberculosis, a combined acid-fast (Auramine)/Nile red assay was applied to heavily positive sputum samples from patients with tuberculosis. Strong intracellular Nile red signals were obtained from acid-fast cells, indicating that ILI occur in M. tuberculosis in vivo. This may reflect a distinct physiological state of these cells, which it has not been possible to reproduce in vitro. These findings indicate that the uptake of long-chain fatty acids and TAG biosynthetic and degradative pathways are important aspects of mycobacterial lipid metabolism, meriting further investigation.

  10. Transient fluctuations of intracellular zinc ions in cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Li, Yuan [Division of Human Nutrition, Department of Preventive Medicine and Community Health, The University of Texas Medical Branch, Galveston, TX 77555 (United States); Maret, Wolfgang, E-mail: womaret@utmb.edu [Division of Human Nutrition, Department of Preventive Medicine and Community Health, The University of Texas Medical Branch, Galveston, TX 77555 (United States); Department of Anesthesiology, The University of Texas Medical Branch, Galveston, TX 77555 (United States)

    2009-08-15

    Zinc is essential for cell proliferation, differentiation, and viability. When zinc becomes limited for cultured cells, DNA synthesis ceases and the cell cycle is arrested. The molecular mechanisms of actions of zinc are believed to involve changes in the availability of zinc(II) ions (Zn{sup 2+}). By employing a fluorescent Zn{sup 2+} probe, FluoZin-3 acetoxymethyl ester, intracellular Zn{sup 2+} concentrations were measured in undifferentiated and in nerve growth factor (NGF)-differentiated rat pheochromocytoma (PC12) cells. Intracellular Zn{sup 2+} concentrations are pico- to nanomolar in PC12 cells and are higher in the differentiated than in the undifferentiated cells. When following cellular Zn{sup 2+} concentrations for 48 h after the removal of serum, a condition that is known to cause cell cycle arrest, Zn{sup 2+} concentrations decrease after 30 min but, remarkably, increase after 1 h, and then decrease again to about one half of the initial concentration. Cell proliferation, measured by an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, decreases after both serum starvation and zinc chelation. Two peaks of Zn{sup 2+} concentrations occur within one cell cycle: one early in the G1 phase and the other in the late G1/S phase. Thus, fluctuations of intracellular Zn{sup 2+} concentrations and established modulation of phosphorylation signaling, via an inhibition of protein tyrosine phosphatases at commensurately low Zn{sup 2+} concentrations, suggest a role for Zn{sup 2+} in the control of the cell cycle. Interventions targeted at these picomolar Zn{sup 2+} fluctuations may be a way of controlling cell growth in hyperplasia, neoplasia, and diseases associated with aberrant differentiation.

  11. Copper transporter 2 regulates intracellular copper and sensitivity to cisplatin.

    Science.gov (United States)

    Huang, Carlos P; Fofana, Mariama; Chan, Jefferson; Chang, Christopher J; Howell, Stephen B

    2014-03-01

    Mammalian cells express two copper (Cu) influx transporters, CTR1 and CTR2. CTR1 serves as an influx transporter for both Cu and cisplatin (cDDP). In mouse embryo fibroblasts, reduction of CTR1 expression renders cells resistant to cDDP whereas reduction of CTR2 makes them hypersensitive both in vitro and in vivo. To investigate the role of CTR2 on intracellular Cu and cDDP sensitivity its expression was molecularly altered in the human epithelial 2008 cancer cell model. Intracellular exchangeable Cu(+) was measured with the fluorescent probe Coppersensor-3 (CS3). The ability of CS3 to report on changes in intracellular Cu(+) was validated by showing that Cu chelators reduced its signal, and that changes in signal accompanied alterations in expression of the major Cu influx transporter CTR1 and the two Cu efflux transporters, ATP7A and ATP7B. Constitutive knock down of CTR2 mRNA by ∼50% reduced steady-state exchangeable Cu by 22-23% and increased the sensitivity of 2008 cells by a factor of 2.6-2.9 in two separate clones. Over-expression of CTR2 increased exchangeable Cu(+) by 150% and rendered the 2008 cells 2.5-fold resistant to cDDP. The results provide evidence that CS3 can quantitatively assess changes in exchangeable Cu(+), and that CTR2 regulates both the level of exchangeable Cu(+) and sensitivity to cDDP in a model of human epithelial cancer. This study introduces CS3 and related sensors as novel tools for probing and assaying Cu-dependent sensitivity to anticancer therapeutics.

  12. Intracellular Assessment of ATP Levels in Caenorhabditis elegans

    Science.gov (United States)

    Palikaras, Konstantinos; Tavernarakis, Nektarios

    2017-01-01

    Eukaryotic cells heavily depend on adenosine triphosphate (ATP) generated by oxidative phosphorylation (OXPHOS) within mitochondria. ATP is the major energy currency molecule, which fuels cell to carry out numerous processes, including growth, differentiation, transportation and cell death among others (Khakh and Burnstock, 2009). Therefore, ATP levels can serve as a metabolic gauge for cellular homeostasis and survival (Artal-Sanz and Tavernarakis, 2009; Gomes et al., 2011; Palikaras et al., 2015). In this protocol, we describe a method for the determination of intracellular ATP levels using a bioluminescence approach in the nematode Caenorhabditis elegans. PMID:28194429

  13. Effects of intracellular acidosis on endothelial function: an overview.

    Science.gov (United States)

    Crimi, Ettore; Taccone, Fabio Silvio; Infante, Teresa; Scolletta, Sabino; Crudele, Valeria; Napoli, Claudio

    2012-04-01

    The endothelium represents the largest functional organ in the human body playing an active role in vasoregulation, coagulation, inflammation, and microvascular permeability. Endothelium contributes to maintain vascular integrity, intravascular volume, and tissue oxygenation promoting inflammatory network response for local defense and repair. Acid-basis homeostasis is an important physiologic parameter that controls cell function, and changes in pH can influence vascular tone by regulating endothelium and vascular smooth muscle cells. This review presents a current perspective of the effects of intracellular acidosis on the function and the basic regulatory mechanisms of endothelial cells.

  14. Improved method for measuring intracellular Ca++ with fluo-3.

    Science.gov (United States)

    Rijkers, G T; Justement, L B; Griffioen, A W; Cambier, J C

    1990-01-01

    The accuracy of flow cytometric measurement of intracellular calcium with fluo-3 is compromised by variation in basal fluorescence intensity due to heterogeneity in dye uptake or compartmentalization. We have loaded cells simultaneously with fluo-3 and SNARF-1. When SNARF-1 fluorescence is collected at approximately 600 nm, its intensity does not change upon cell activation. Furthermore, fluo-3 and SNARF-1 fluorescence signals exhibit a linear relationship. The ratio of fluo-3 to SNARF-1 eliminates a significant proportion of variation in fluorescence intensity caused by variation in fluo-3 uptake and thus can be used as a sensitive parameter for measuring changes in [Ca2+]i.

  15. Analysis of Intracellular Calcium Signaling in Human Embryonic Stem Cells.

    Science.gov (United States)

    Péntek, Adrienn; Pászty, Katalin; Apáti, Ágota

    2016-01-01

    Measurement of changes in intracellular calcium concentration is one of the most common and useful tools for studying signal transduction pathways or cellular responses in basic research and drug screening purposes as well. Increasing number of such applications using human pluripotent stem cells and their derivatives requires development of calcium signal measurements for this special cell type. Here we describe a modified protocol for analysis of calcium signaling events in human embryonic stem cells, which can be used for other pluripotent cell types (such as iPSC) or their differentiated offspring as well.

  16. Polymeric gel nanoparticle pH sensors for intracellular measurements

    DEFF Research Database (Denmark)

    Almdal, Kristoffer; Andresen, Thomas Lars; Benjaminsen, Rikke Vicki

    Precise measurements of pH in cells and intracellular compartments are of importance to both the fundamental understanding of metabolism and to the development of drugs that are released from the endosomes-lysome pathway. We have developed polymer gel nanoparticles as carriers of covalently bound...... bonding to the gel nanoparticle. The synthesis also allows for introduction of charged groups. Particle charge influence the cell uptake heavily. The synthesis scheme is outlined below. The sensors are internalized in cells through endocytosis and the sensor allows for the accurate measurement of p...

  17. Quantitative Proteomics of Intracellular Campylobacter jejuni Reveals Metabolic Reprogramming.

    Directory of Open Access Journals (Sweden)

    Xiaoyun Liu

    Full Text Available Campylobacter jejuni is the major cause of bacterial food-borne illness in the USA and Europe. An important virulence attribute of this bacterial pathogen is its ability to enter and survive within host cells. Here we show through a quantitative proteomic analysis that upon entry into host cells, C. jejuni undergoes a significant metabolic downshift. Furthermore, our results indicate that intracellular C. jejuni reprograms its respiration, favoring the respiration of fumarate. These results explain the poor ability of C. jejuni obtained from infected cells to grow under standard laboratory conditions and provide the bases for the development of novel anti microbial strategies that would target relevant metabolic pathways.

  18. Intracellular coagulation inhibits the extraction of proteins from Prochloron

    Science.gov (United States)

    Fall, R.; Lewin, R. A.; Fall, L. R.

    1983-01-01

    Protein extraction from the prokaryotic alga Prochloron LP (isolated from the ascidian host Lissoclinum patella) was complicated by an irreversible loss of cell fragility in the isolated algae. Accompanying this phenomenon, which is termed intracellular coagulation, was a redistribution of thylakoids around the cell periphery, a loss of photosynthetic O2 production, and a drastic decrease in the extractability of cell proteins. Procedures are described for the successful preparation and transport of cell extracts yielding the enzymes glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase as well as other soluble proteins.

  19. The intracellular cyanobacteria of Paulinella chromatophora: endosymbionts or organelles?

    Science.gov (United States)

    Bodył, Andrzej; Mackiewicz, Paweł; Stiller, John W

    2007-07-01

    Endosymbiotic relationships are common across the tree of life and have had profound impacts on cellular evolution and diversity. Recent molecular investigations of the amoeba Paulinella chromatophora have raised a timely and important question: should obligatory intracellular cyanobacteria in Paulinella be considered new organelles, or do plastids and mitochondria hold a unique stature in the history of endosymbiotic events? We argue that drawing a sharp distinction between these two organelles and all other endosymbionts is not supported by accumulating data, neither is it a productive framework for investigating organelle evolution.

  20. Intracellular coagulation inhibits the extraction of proteins from Prochloron

    Science.gov (United States)

    Fall, R.; Lewin, R. A.; Fall, L. R.

    1983-01-01

    Protein extraction from the prokaryotic alga Prochloron LP (isolated from the ascidian host Lissoclinum patella) was complicated by an irreversible loss of cell fragility in the isolated algae. Accompanying this phenomenon, which is termed intracellular coagulation, was a redistribution of thylakoids around the cell periphery, a loss of photosynthetic O2 production, and a drastic decrease in the extractability of cell proteins. Procedures are described for the successful preparation and transport of cell extracts yielding the enzymes glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase as well as other soluble proteins.

  1. Effects of Time Delay on Intracellular Ca2+ Concentration Oscillations

    Institute of Scientific and Technical Information of China (English)

    YING Yang-Jun; HUANG Zu-Qia

    2001-01-01

    Based on the SS-model [Somogyi R and Stucki J W J. Biol. Chem. 266 (1991) 11 068] for the generation of intracellular Ca2+ concentration oscillations, we consider a time delay for the binding kinetics of the Ca2+ channel and find a significant phenomenon that the oscillation takes two quite different modes when a parameter of the system crosses a threshold. One is a quick oscillation mode and the other is a slow oscillation mode. The oscillation frequencies of these modes differ from each other by more than ten times. The change of oscillation form with parameters and its critical behaviour are illustrated by numerical simulation results.

  2. Study the effect of Vitamin K on intracellular NAD level in yeast by fluorescence spectrum

    Science.gov (United States)

    Yahong, Chen; Ruxiu, Cai; Ke, Zhang

    2007-05-01

    The intracellular NAD level plays a pivotal role in numerous biological processes such as rhythm, senescence, cancer and death. The study of the intracellular NAD level has been one of the "hotspots" in biomedical research. We investigated the effect of Vitamin K on intracellular NAD level in yeast by fluorescence spectrum in this paper. Plasma membrane redox system of yeast was found to be greatly promoted by the addition of Vitamin K 3 or Vitamin K 1. Ferricyanide reduction catalyzed by Vitamin K was accompanied by the decrease in intracellular NADH concentration and the increase in intracellular NAD level of yeast cells.

  3. 高盐条件下偶氮染料兼氧生物降解性能研究%Study on Facultative Anaerobic Biodegradation Performance of Azo Dyestuff under High Salt-containing Condition

    Institute of Scientific and Technical Information of China (English)

    仝攀瑞; 雷弢; 陈方方; 王晓艳; 常向东

    2012-01-01

    The facultative biodegradation performance of reactive dyestuffs is studied by using different concentrations glucose as co-substrate. Based on this, the facultative biodegradation performance of K-2BP under different salt-containing condition was investigated by static test reactor. The results indicated that facultative microorganisms had lower degradation rate when only K-2BP was used as the substrate, and the degradation rate would be increased under the existence of the glucose. The glucose concentration is 800 mg/L, while the degradation rate of K-2BP is 64. 1% at 6h. However, it is detrimental to dye' s biodegradation when the glucose concentration is up to 1000 mg/L, the degradation rate is only about 46% , that is similar to the condition without glucose. On the premise of glucose concentration 800 mg/L, when the salt concentrations are 2 g/‵ 5 g/L,10 g/L and 20 g/L respectively,the first-order kinetics constants are 0. 10578 mg/L · h,0.04947 mg/ L · h ,0.02869 mg/L · h and 0.022 75 mg/L · h respectively. And the half-lives were 6.99 h J4. 15 hN22. 55 h and 30.21h respectively. The reaction rate constants decrease gradually with the increase of the salt concentrations, and when the salt concentration is more than 2g/L, it can restrain the facultative microorganisms' degradation of K-2BP will be restrained.%研究活性染料与不同浓度葡萄糖在共基质条件下的兼氧生物降解性能,在此基础上,研究K-2BP在不同盐浓度条件下的兼氧生物降解性能.选择K-2BP作为目标污染物进行静态反应器生物降解试验,结果表明:兼氧微生物在只有K-2BP作为基质时对染料的降解率较低;葡萄糖存在时,能提高兼氧生物对染料的降解能力,葡萄糖为800 mg/L时,6h染料降解率为64.1%,而葡萄糖浓度为1 000 mg/L时,不利于染料降解,6h染料降解率为46%,与不投加葡萄糖情况的降解率接近.葡萄糖浓度为800mg/L,盐浓度分别为2,5,10和20 g/L,其一级

  4. CIRRHOSIS INDUCES APOPTOSIS IN RENAL TISSUE THROUGH INTRACELLULAR OXIDATIVE STRESS

    Directory of Open Access Journals (Sweden)

    Keli Cristina Simões da SILVEIRA

    2015-03-01

    Full Text Available Background Renal failure is a frequent and serious complication in patients with decompensated cirrhosis. Objectives We aimed to evaluate the renal oxidative stress, cell damage and impaired cell function in animal model of cirrhosis. Methods Secondary biliary cirrhosis was induced in rats by ligation of the common bile duct. We measured TBARS, ROS and mitochondrial membrane potential in kidney as markers of oxidative stress, and activities of the antioxidant enzymes. Relative cell viability was determined by trypan blue dye-exclusion assay. Annexin V-PE was used with a vital dye, 7-AAD, to distinguish apoptotic from necrotic cells and comet assay was used for determined DNA integrity in single cells. Results In bile duct ligation animals there was significant increase in the kidney lipoperoxidation and an increase of the level of intracellular ROS. There was too an increase in the activity of all antioxidant enzymes evaluated in the kidney. The percentage viability was above 90% in the control group and in bile duct ligation was 64.66% and the dominant cell death type was apoptosis. DNA damage was observed in the bile duct ligation. There was a decreased in the mitochondrial membrane potential from 71.40% ± 6.35% to 34.48% ± 11.40% in bile duct ligation. Conclusions These results indicate that intracellular increase of ROS cause damage in the DNA and apoptosis getting worse the renal function in cirrhosis.

  5. FIB/SEM cell sectioning for intracellular metal granules characterization

    Science.gov (United States)

    Milani, Marziale; Brundu, Claudia; Santisi, Grazia; Savoia, Claudio; Tatti, Francesco

    2009-05-01

    Focused Ion Beams (FIBs) provide a cross-sectioning tool for submicron dissection of cells and subcellular structures. In combination with Scanning Electron Microscope (SEM), FIB provides complementary morphological information, that can be further completed by EDX (Energy Dispersive X-ray Spectroscopy). This study focus onto intracellular microstructures, particularly onto metal granules (typically Zn, Cu and Fe) and on the possibility of sectioning digestive gland cells of the terrestrial isopod P. scaber making the granules available for a compositional analysis with EDX. Qualitative and quantitative analysis of metal granules size, amount and distribution are performed. Information is made available of the cellular storing pattern and, indirectly, metal metabolism. The extension to human level is of utmost interest since some pathologies of relevance are metal related. Apart from the common metal-overload-diseases (hereditary hemochromatosis, Wilson's and Menkes disease) it has been demonstrated that metal in excess can influence carcinogenesis in liver, kidney and breast. Therefore protocols will be established for the observation of mammal cells to improve our knowledge about the intracellular metal amount and distribution both in healthy cells and in those affected by primary or secondary metal overload or depletion.

  6. Origins of intracellular calcium mobilization evoked by infrared laser stimulation

    Science.gov (United States)

    Olsovsky, Cory A.; Tolstykh, Gleb P.; Ibey, Bennett L.; Beier, Hope T.

    2015-03-01

    Cellular delivery of pulsed IR laser energy has been shown to stimulate action potentials in neurons. The mechanism for this stimulation is not completely understood. Certain hypotheses suggest the rise in temperature from IR exposure could activate temperature- or pressure-sensitive channels, or create pores in the cellular outer membrane. Studies using intensity-based Ca2+-responsive dyes show changes in Ca2+ levels after various IR stimulation parameters; however, determination of the origin of this signal proved difficult. An influx of larger, typically plasma-membrane-impermeant ions has been demonstrated, which suggests that Ca2+ may originate from the external solution. However, activation of intracellular signaling pathways, possibly indicating a more complex role of increasing Ca2+ concentration, has also been shown. By usingCa2+ sensitive dye Fura-2 and a high-speed ratiometric imaging system that rapidly alternates the excitation wavelengths, we have quantified the Ca2+ mobilization in terms of influx from the external solution and efflux from intracellular organelles. CHO-K1 cells, which lack voltage-gated Ca2+ channels, and NG-108 neuroblastoma cells, which do not produce action potentials in an early undifferentiated state, are used to determine the origin of the Ca2+ signals and investigate the role these mechanisms may play in IR neural stimulation.

  7. Crystallographic study of FABP5 as an intracellular endocannabinoid transporter

    Energy Technology Data Exchange (ETDEWEB)

    Sanson, Benoît; Wang, Tao [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Sun, Jing; Wang, Liqun; Kaczocha, Martin [Stony Brook University, Stony Brook, NY 11794-5213 (United States); Ojima, Iwao [Stony Brook University, Stony Brook, NY 1794-3400 (United States); Stony Brook University, Stony Brook, NY 11794-3400 (United States); Deutsch, Dale, E-mail: dale.deutsch@stonybrook.edu [Stony Brook University, Stony Brook, NY 11794-5213 (United States); Stony Brook University, Stony Brook, NY 11794-3400 (United States); Li, Huilin, E-mail: dale.deutsch@stonybrook.edu [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Stony Brook University, Stony Brook, NY 11794-5213 (United States); Stony Brook University, Stony Brook, NY 11794-3400 (United States)

    2014-02-01

    FABP5 was recently found to intracellularly transport endocannabinoid signaling lipids. The structures of FABP5 complexed with two endocannabinoids and an inhibitor were solved. Human FABP5 was found to dimerize via a domain-swapping mechanism. This work will help in the development of inhibitors to raise endocannabinoid levels. In addition to binding intracellular fatty acids, fatty-acid-binding proteins (FABPs) have recently been reported to also transport the endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG), arachidonic acid derivatives that function as neurotransmitters and mediate a diverse set of physiological and psychological processes. To understand how the endocannabinoids bind to FABPs, the crystal structures of FABP5 in complex with AEA, 2-AG and the inhibitor BMS-309403 were determined. These ligands are shown to interact primarily with the substrate-binding pocket via hydrophobic interactions as well as a common hydrogen bond to the Tyr131 residue. This work advances our understanding of FABP5–endocannabinoid interactions and may be useful for future efforts in the development of small-molecule inhibitors to raise endocannabinoid levels.

  8. Production of intracellular enzymes by enzymatic treatment of yeast

    Energy Technology Data Exchange (ETDEWEB)

    Zomer, E.; Er-El, Z.; Rokem, J.S.

    1987-01-01

    Enzymatic extraction of intracellular enzymes from various yeasts by glucanase was investigated. Favourable conditions for lysis and release of intracellular enzymes were established. The effects of yeast concentration, growth phase of yeast, storage temperature and pretreatment of yeast were studied. The yeasts investigated can be divided into two groups. The first, Kluyveromyces lactis, Saccharomyces cerevisiae, Saccharomyces oviformis, Torulopsis glabrata, Hansenula polymorpha and local bakers' yeast, lysed relatively easily (70-80% of the cells), especially when cells from the logarithmic growth phase were treated. The second, Candida utilis and Candida vini, were more susceptible to lysis (40-50%) when cells were taken from the stationary phase. Release of two enzymes, glycerol kinase from Candida utilis grown on glycerol and formate dehydrogenase from Torulopsis glabrata grown on methanol was examined. The highest specific activities were obtained by incubating the cells with glucanase for 1.5 hours at 37 degrees C. Inactivation of the released enzyme was relatively low. After 12 hours of enzymatic treatment at 28 degrees C glycerol kinase maintained about 50%, and formate dehydrogenase over 80%, of the original activities. (Refs. 12).

  9. Regulation of BMP2-induced intracellular calcium increases in osteoblasts.

    Science.gov (United States)

    Xu, Wenfeng; Liu, Bo; Liu, Xue; Chiang, Martin Y M; Li, Bo; Xu, Zichen; Liao, Xiaoling

    2016-10-01

    Although bone morphogenetic protein-2 (BMP2) is a well-characterized regulator that stimulates osteoblast differentiation, little is known about how it regulates intracellular Ca(2+) signaling. In this study, intracellular Ca(2+) concentration ([Ca(2+) ]i ) upon BMP2 application, focal adhesion kinase (FAK) and Src activities were measured in the MC3T3-E1 osteoblast cell line using fluorescence resonance energy transfer-based biosensors. Increase in [Ca(2+) ]i , FAK, and Src activities were observed during BMP2 stimulation. The removal of extracellular calcium, the application of membrane channel inhibitors streptomycin or nifedipine, the FAK inhibitor PF-573228 (PF228), and the alkaline phosphatase (ALP) siRNA all blocked the BMP2-stimulated [Ca(2+) ]i increase, while the Src inhibitor PP1 did not. In contrast, a gentle decrease of endoplasmic reticulum calcium concentration was found after BMP2 stimulation, which could be blocked by both streptomycin and PP1. Further experiments revealed that BMP2-induced FAK activation could not be inhibited by PP1, ALP siRNA or the calcium channel inhibitor nifedipine. PF228, but not PP1 or calcium channel inhibitors, suppressed ALP elevation resulting from BMP2 stimulation. Therefore, our results suggest that BMP2 can increase [Ca(2+) ]i through extracellular calcium influx regulated by FAK and ALP and can deplete ER calcium through Src signaling simultaneously. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1725-1733, 2016.

  10. Charcot–Marie–Tooth disease and intracellular traffic

    Science.gov (United States)

    Bucci, Cecilia; Bakke, Oddmund; Progida, Cinzia

    2012-01-01

    Mutations of genes whose primary function is the regulation of membrane traffic are increasingly being identified as the underlying causes of various important human disorders. Intriguingly, mutations in ubiquitously expressed membrane traffic genes often lead to cell type- or organ-specific disorders. This is particularly true for neuronal diseases, identifying the nervous system as the most sensitive tissue to alterations of membrane traffic. Charcot–Marie–Tooth (CMT) disease is one of the most common inherited peripheral neuropathies. It is also known as hereditary motor and sensory neuropathy (HMSN), which comprises a group of disorders specifically affecting peripheral nerves. This peripheral neuropathy, highly heterogeneous both clinically and genetically, is characterized by a slowly progressive degeneration of the muscle of the foot, lower leg, hand and forearm, accompanied by sensory loss in the toes, fingers and limbs. More than 30 genes have been identified as targets of mutations that cause CMT neuropathy. A number of these genes encode proteins directly or indirectly involved in the regulation of intracellular traffic. Indeed, the list of genes linked to CMT disease includes genes important for vesicle formation, phosphoinositide metabolism, lysosomal degradation, mitochondrial fission and fusion, and also genes encoding endosomal and cytoskeletal proteins. This review focuses on the link between intracellular transport and CMT disease, highlighting the molecular mechanisms that underlie the different forms of this peripheral neuropathy and discussing the pathophysiological impact of membrane transport genetic defects as well as possible future ways to counteract these defects. PMID:22465036

  11. Mechanisms of Obligatory Intracellular Infection with Anaplasma phagocytophilum

    Science.gov (United States)

    Rikihisa, Yasuko

    2011-01-01

    Summary: Anaplasma phagocytophilum persists in nature by cycling between mammals and ticks. Human infection by the bite of an infected tick leads to a potentially fatal emerging disease called human granulocytic anaplasmosis. A. phagocytophilum is an obligatory intracellular bacterium that replicates inside mammalian granulocytes and the salivary gland and midgut cells of ticks. A. phagocytophilum evolved the remarkable ability to hijack the regulatory system of host cells. A. phagocytophilum alters vesicular traffic to create an intracellular membrane-bound compartment that allows replication in seclusion from lysosomes. The bacterium downregulates or actively inhibits a number of innate immune responses of mammalian host cells, and it upregulates cellular cholesterol uptake to acquire cholesterol for survival. It also upregulates several genes critical for the infection of ticks, and it prolongs tick survival at freezing temperatures. Several host factors that exacerbate infection have been identified, including interleukin-8 (IL-8) and cholesterol. Host factors that overcome infection include IL-12 and gamma interferon (IFN-γ). Two bacterial type IV secretion effectors and several bacterial proteins that associate with inclusion membranes have been identified. An understanding of the molecular mechanisms underlying A. phagocytophilum infection will foster the development of creative ideas to prevent or treat this emerging tick-borne disease. PMID:21734244

  12. Influenza vaccine induces intracellular immune memory of human NK cells.

    Directory of Open Access Journals (Sweden)

    Yaling Dou

    Full Text Available Influenza vaccines elicit antigen-specific antibodies and immune memory to protect humans from infection with drift variants. However, what supports or limits vaccine efficacy and duration is unclear. Here, we vaccinated healthy volunteers with annual vaccine formulations and investigated the dynamics of T cell, natural killer (NK cell and antibody responses upon restimulation with heterologous or homologous influenza virus strains. Influenza vaccines induced potential memory NK cells with increased antigen-specific recall IFN-γ responses during the first 6 months. In the absence of significant changes in other NK cell markers (CD45RO, NKp44, CXCR6, CD57, NKG2C, CCR7, CD62L and CD27, influenza vaccines induced memory NK cells with the distinct feature of intracellular NKp46 expression. Indeed, surface NKp46 was internalized, and the dynamic increase in NKp46(intracellular+CD56dim NK cells positively correlated with increased IFN-γ production to influenza virus restimulation after vaccination. In addition, anti-NKp46 antibodies blocked IFN-γ responses. These findings provide insights into a novel mechanism underlying vaccine-induced immunity and NK-related diseases, which may help to design persisting and universal vaccines in the future.

  13. Influenza vaccine induces intracellular immune memory of human NK cells.

    Science.gov (United States)

    Dou, Yaling; Fu, Binqing; Sun, Rui; Li, Wenting; Hu, Wanfu; Tian, Zhigang; Wei, Haiming

    2015-01-01

    Influenza vaccines elicit antigen-specific antibodies and immune memory to protect humans from infection with drift variants. However, what supports or limits vaccine efficacy and duration is unclear. Here, we vaccinated healthy volunteers with annual vaccine formulations and investigated the dynamics of T cell, natural killer (NK) cell and antibody responses upon restimulation with heterologous or homologous influenza virus strains. Influenza vaccines induced potential memory NK cells with increased antigen-specific recall IFN-γ responses during the first 6 months. In the absence of significant changes in other NK cell markers (CD45RO, NKp44, CXCR6, CD57, NKG2C, CCR7, CD62L and CD27), influenza vaccines induced memory NK cells with the distinct feature of intracellular NKp46 expression. Indeed, surface NKp46 was internalized, and the dynamic increase in NKp46(intracellular)+CD56dim NK cells positively correlated with increased IFN-γ production to influenza virus restimulation after vaccination. In addition, anti-NKp46 antibodies blocked IFN-γ responses. These findings provide insights into a novel mechanism underlying vaccine-induced immunity and NK-related diseases, which may help to design persisting and universal vaccines in the future.

  14. Charcot-Marie-Tooth disease and intracellular traffic.

    Science.gov (United States)

    Bucci, Cecilia; Bakke, Oddmund; Progida, Cinzia

    2012-12-01

    Mutations of genes whose primary function is the regulation of membrane traffic are increasingly being identified as the underlying causes of various important human disorders. Intriguingly, mutations in ubiquitously expressed membrane traffic genes often lead to cell type- or organ-specific disorders. This is particularly true for neuronal diseases, identifying the nervous system as the most sensitive tissue to alterations of membrane traffic. Charcot-Marie-Tooth (CMT) disease is one of the most common inherited peripheral neuropathies. It is also known as hereditary motor and sensory neuropathy (HMSN), which comprises a group of disorders specifically affecting peripheral nerves. This peripheral neuropathy, highly heterogeneous both clinically and genetically, is characterized by a slowly progressive degeneration of the muscle of the foot, lower leg, hand and forearm, accompanied by sensory loss in the toes, fingers and limbs. More than 30 genes have been identified as targets of mutations that cause CMT neuropathy. A number of these genes encode proteins directly or indirectly involved in the regulation of intracellular traffic. Indeed, the list of genes linked to CMT disease includes genes important for vesicle formation, phosphoinositide metabolism, lysosomal degradation, mitochondrial fission and fusion, and also genes encoding endosomal and cytoskeletal proteins. This review focuses on the link between intracellular transport and CMT disease, highlighting the molecular mechanisms that underlie the different forms of this peripheral neuropathy and discussing the pathophysiological impact of membrane transport genetic defects as well as possible future ways to counteract these defects.

  15. Mitochondrial dynamics and their intracellular traffic in porcine oocytes.

    Science.gov (United States)

    Yamochi, T; Hashimoto, S; Amo, A; Goto, H; Yamanaka, M; Inoue, M; Nakaoka, Y; Morimoto, Y

    2016-08-01

    Meiotic maturation of oocytes requires a variety of ATP-dependent reactions, such as germinal vesicle breakdown, spindle formation, and rearrangement of plasma membrane structure, which is required for fertilization. Mitochondria are accordingly expected be localized to subcellular sites of energy utilization. Although microtubule-dependent cellular traffic for mitochondria has been studied extensively in cultured neuronal (and some other somatic) cells, the molecular mechanism of their dynamics in mammalian oocytes at different stages of maturation remains obscure. The present work describes dynamic aspects of mitochondria in porcine oocytes at the germinal vesicle stage. After incubation of oocytes with MitoTracker Orange followed by centrifugation, mitochondria-enriched ooplasm was obtained using a glass needle and transferred into a recipient oocyte. The intracellular distribution of the fluorescent mitochondria was then observed over time using a laser scanning confocal microscopy equipped with an incubator. Kinetic analysis revealed that fluorescent mitochondria moved from central to subcortical areas of oocytes and were dispersed along plasma membranes. Such movement of mitochondria was inhibited by either cytochalasin B or cytochalasin D but not by colcemid, suggesting the involvement of microfilaments. This method of visualizing mitochondrial dynamics in live cells permits study of the pathophysiology of cytoskeleton-dependent intracellular traffic of mitochondria and associated energy metabolism during meiotic maturation of oocytes.

  16. Taxonomic status of the intracellular bacterium Wolbachia pipientis.

    Science.gov (United States)

    Lo, N; Paraskevopoulos, C; Bourtzis, K; O'Neill, S L; Werren, J H; Bordenstein, S R; Bandi, C

    2007-03-01

    Wolbachia pipientis is a maternally inherited, intracellular bacterium found in more than 20 % of all insects, as well as numerous other arthropods and filarial nematodes. It has been the subject of a growing number of studies in recent decades, because of the remarkable effects it has on its arthropod hosts, its potential as a tool for biological control of arthropods of agricultural and medical importance and its use as a target for treatment of filariasis. W. pipientis was originally discovered in cells of the mosquito Culex pipiens and is the only formally described member of the genus. Molecular sequence-based studies have revealed a number of phylogenetically diverse strains of W. pipientis. Owing to uncertainty about whether W. pipientis comprises more than one species, researchers in the field now commonly refer to W. pipientis simply as Wolbachia. In this note, we briefly review higher-level phylogenetic and recombination studies of W. pipientis and propose that all the intracellular symbionts known to cluster closely with the type strain of W. pipientis, including those in the currently recognized supergroups (A-H), are officially given this name.

  17. Uptake and intracellular trafficking of superantigens in dendritic cells.

    Directory of Open Access Journals (Sweden)

    María B Ganem

    Full Text Available Bacterial superantigens (SAgs are exotoxins produced mainly by Staphylococcus aureus and Streptococcus pyogenes that can cause toxic shock syndrome (TSS. According to current paradigm, SAgs interact directly and simultaneously with T cell receptor (TCR on the T cell and MHC class II (MHC-II on the antigen-presenting cell (APC, thereby circumventing intracellular processing to trigger T cell activation. Dendritic cells (DCs are professional APCs that coat nearly all body surfaces and are the most probable candidate to interact with SAgs. We demonstrate that SAgs are taken up by mouse DCs without triggering DC maturation. SAgs were found in intracellular acidic compartment of DCs as biologically active molecules. Moreover, SAgs co-localized with EEA1, RAB-7 and LAMP-2, at different times, and were then recycled to the cell membrane. DCs loaded with SAgs are capable of triggering in vitro lymphocyte proliferation and, injected into mice, stimulate T cells bearing the proper TCR in draining lymph nodes. Transportation and trafficking of SAgs in DCs might increase the local concentration of these exotoxins where they will produce the highest effect by promoting their encounter with both MHC-II and TCR in lymph nodes, and may explain how just a few SAg molecules can induce the severe pathology associated with TSS.

  18. Efficient intracellular retrotransposition of an exogenous primate retrovirus genome

    Science.gov (United States)

    Heinkelein, Martin; Pietschmann, Thomas; Jármy, Gergely; Dressler, Marco; Imrich, Horst; Thurow, Jana; Lindemann, Dirk; Bock, Michael; Moebes, Astrid; Roy, Jacqueline; Herchenröder, Ottmar; Rethwilm, Axel

    2000-01-01

    The foamy virus (FV) subgroup of Retroviridae reverse transcribe their RNA (pre-)genome late in the replication cycle before leaving an infected cell. We studied whether a marker gene-transducing FV vector is able to shuttle to the nucleus and integrate into host cell genomic DNA. While a potential intracellular retrotransposition of vectors derived from other retroviruses was below the detection limit of our assay, we found that up to 5% of cells transfected with the FV vector were stably transduced, harboring 1 to ∼10 vector integrants. Generation of the integrants depended on expression of functional capsid, reverse transcriptase and integrase proteins, and did not involve an extracellular step. PCR analysis of the U3 region of the 5′ long terminal repeat and determination of proviral integration sites showed that a reverse transcription step had taken place to generate the integrants. Co-expression of a mutated envelope allowing particle egress and avoiding extracellular infection resulted in a significantly increased rescue of cells harboring integrants, suggesting that accumulation of proviruses via intracellular retrotransposition represents an integral part of the FV replication strategy. PMID:10880456

  19. Collective Resistance in Microbial Communities by Intracellular Antibiotic Deactivation

    Science.gov (United States)

    Sorg, Robin A.; Lin, Leo; van Doorn, G. Sander; Sorg, Moritz; Olson, Joshua; Nizet, Victor; Veening, Jan-Willem

    2016-01-01

    The structure and composition of bacterial communities can compromise antibiotic efficacy. For example, the secretion of β-lactamase by individual bacteria provides passive resistance for all residents within a polymicrobial environment. Here, we uncover that collective resistance can also develop via intracellular antibiotic deactivation. Real-time luminescence measurements and single-cell analysis demonstrate that the opportunistic human pathogen Streptococcus pneumoniae grows in medium supplemented with chloramphenicol (Cm) when resistant bacteria expressing Cm acetyltransferase (CAT) are present. We show that CAT processes Cm intracellularly but not extracellularly. In a mouse pneumonia model, more susceptible pneumococci survive Cm treatment when coinfected with a CAT-expressing strain. Mathematical modeling predicts that stable coexistence is only possible when antibiotic resistance comes at a fitness cost. Strikingly, CAT-expressing pneumococci in mouse lungs were outcompeted by susceptible cells even during Cm treatment. Our results highlight the importance of the microbial context during infectious disease as a potential complicating factor to antibiotic therapy. PMID:28027306

  20. Intracellular zinc distribution in mitochondria, ER and the Golgi apparatus.

    Science.gov (United States)

    Lu, Qiping; Haragopal, Hariprakash; Slepchenko, Kira G; Stork, Christian; Li, Yang V

    2016-01-01

    Zinc (Zn(2+)) is required for numerous cellular functions. As such, the homeostasis and distribution of intracellular zinc can influence cellular metabolism and signaling. However, the exact distribution of free zinc within live cells remains elusive. Previously we showed the release of zinc from thapsigargin/IP3-sensitive endoplasmic reticulum (ER) storage in cortical neurons. In the present study, we investigated if other cellular organelles also contain free chelatable zinc and function as organelle storage for zinc. To identify free zinc within the organelles, live cells were co-stained with Zinpyr-1, a zinc fluorescent dye, and organelle-specific fluorescent dyes (MitoFluor Red 589: mitochondria; ER Tracker Red: endoplasmic reticulum; BODIPY TR ceramide: Golgi apparatus; Syto Red 64: nucleus). We examined organelles that represent potential storing sites for intracellular zinc. We showed that zinc fluorescence staining was co-localized with MitoFluor Red 589, ER Tracker Red, and BODIPY TR ceramide respectively, suggesting the presence of free zinc in mitochondria, endoplasmic reticulum, and the Golgi apparatus. On the other hand, cytosol and nucleus had nearly no detectable zinc fluorescence. It is known that nucleus contains high amount of zinc binding proteins that have high zinc binding affinity. The absence of zinc fluorescence suggests that there is little free zinc in these two regions. It also indicates that the zinc fluorescence detected in mitochondria, ER and Golgi apparatus represents free chelatable zinc. Taken together, our results support that these organelles are potential zinc storing organelles during cellular zinc homeostasis.

  1. Multistability and dynamic transitions of intracellular Min protein patterns.

    Science.gov (United States)

    Wu, Fabai; Halatek, Jacob; Reiter, Matthias; Kingma, Enzo; Frey, Erwin; Dekker, Cees

    2016-06-08

    Cells owe their internal organization to self-organized protein patterns, which originate and adapt to growth and external stimuli via a process that is as complex as it is little understood. Here, we study the emergence, stability, and state transitions of multistable Min protein oscillation patterns in live Escherichia coli bacteria during growth up to defined large dimensions. De novo formation of patterns from homogenous starting conditions is observed and studied both experimentally and in simulations. A new theoretical approach is developed for probing pattern stability under perturbations. Quantitative experiments and simulations show that, once established, Min oscillations tolerate a large degree of intracellular heterogeneity, allowing distinctly different patterns to persist in different cells with the same geometry. Min patterns maintain their axes for hours in experiments, despite imperfections, expansion, and changes in cell shape during continuous cell growth. Transitions between multistable Min patterns are found to be rare events induced by strong intracellular perturbations. The instances of multistability studied here are the combined outcome of boundary growth and strongly nonlinear kinetics, which are characteristic of the reaction-diffusion patterns that pervade biology at many scales.

  2. Imaging the intracellular degradation of biodegradable polymer nanoparticles

    Directory of Open Access Journals (Sweden)

    Anne-Kathrin Barthel

    2014-10-01

    Full Text Available In recent years, the development of smart drug delivery systems based on biodegradable polymeric nanoparticles has become of great interest. Drug-loaded nanoparticles can be introduced into the cell interior via endocytotic processes followed by the slow release of the drug due to degradation of the nanoparticle. In this work, poly(L-lactic acid (PLLA was chosen as the biodegradable polymer. Although common degradation of PLLA has been studied in various biological environments, intracellular degradation processes have been examined only to a very limited extent. PLLA nanoparticles with an average diameter of approximately 120 nm were decorated with magnetite nanocrystals and introduced into mesenchymal stem cells (MSCs. The release of the magnetite particles from the surface of the PLLA nanoparticles during the intracellular residence was monitored by transmission electron microscopy (TEM over a period of 14 days. It was demonstrated by the release of the magnetite nanocrystals from the PLLA surface that the PLLA nanoparticles do in fact undergo degradation within the cell. Furthermore, even after 14 days of residence, the PLLA nanoparticles were found in the MSCs. Additionally, the ultrastructural TEM examinations yield insight into the long term intercellular fate of these nanoparticles. From the statistical analysis of ultrastructural details (e.g., number of detached magnetite crystals, and the number of nanoparticles in one endosome, we demonstrate the importance of TEM studies for such applications in addition to fluorescence studies (flow cytometry and confocal laser scanning microscopy.

  3. Hybrid micro-/nanogels for optical sensing and intracellular imaging

    Directory of Open Access Journals (Sweden)

    Shuiqin Zhou

    2010-12-01

    Full Text Available Hybrid micro-/nanogels are playing an increasing important part in a diverse range of applications, due to their tunable dimensions, large surface area, stable interior network structure, and a very short response time. We review recent advances and challenges in the developments of hybrid micro-/nanogels toward applications for optical sensing of pH, temperature, glucose, ions, and other species as well as for intracellular imaging. Due to their unique advantages, hybrid micro-/nanogels as optical probes are attracting substantial interests for continuous monitoring of chemical parameters in complex samples such as blood and bioreactor fluids, in chemical research and industry, and in food quality control. In particular, their intracellular probing ability enables the monitoring of the biochemistry and biophysics of live cells over time and space, thus contributing to the explanation of intricate biological processes and the development of novel diagnoses. Unlike most other probes, hybrid micro-/nanogels could also combine other multiple functions into a single probe. The rational design of hybrid micro-/nanogels will not only improve the probing applications as desirable, but also implement their applications in new arenas. With ongoing rapid advances in bionanotechnology, the well-designed hybrid micro-/nanogel probes will be able to provide simultaneous sensing, imaging diagnosis, and therapy toward clinical applications.

  4. Cyanobacteria perceive nitrogen status by sensing intracellular 2-oxoglutarate levels.

    Science.gov (United States)

    Muro-Pastor, M I; Reyes, J C; Florencio, F J

    2001-10-12

    The regulatory circuits that control nitrogen metabolism are relatively well known in several bacterial model groups. However, much less is understood about how the nitrogen status of the cell is perceived in vivo. In cyanobacteria, the transcription factor NtcA is required for regulation (activation or repression) of an extensive number of genes involved in nitrogen metabolism. In contrast, how NtcA activity is regulated is largely unknown. Assimilation of ammonium by most microorganisms occurs through the sequential action of two enzymes: glutamine synthetase (GS) and glutamate synthase. Interestingly, regulation of the expression of NtcA-dependent genes in the cyanobacterium Synechocystis sp. PCC 6803 is altered in mutants with modified levels of GS activity. Two types of mutants were analyzed: glnA null mutants that lack GS type I and gif mutants unable to inactivate GS in the presence of ammonium. Changes in the intracellular pools of 19 different amino acids and the keto acid 2-oxoglutarate were recorded in wild-type and mutant strains under different nitrogen conditions. Our data strongly indicate that the nitrogen status in cyanobacteria is perceived as changes in the intracellular 2-oxoglutarate pool.

  5. Intracellular Peptides as Natural Regulators of Cell Signaling*S⃞

    Science.gov (United States)

    Cunha, Fernanda M.; Berti, Denise A.; Ferreira, Zulma S.; Klitzke, Clécio F.; Markus, Regina P.; Ferro, Emer S.

    2008-01-01

    Protein degradation by the ubiquitin proteasome system releases large amounts of oligopeptides within cells. To investigate possible functions for these intracellularly generated oligopeptides, we fused them to a cationic transactivator peptide sequence using reversible disulfide bonds, introduced them into cells, and analyzed their effect on G protein-coupled receptor (GPCR) signal transduction. A mixture containing four of these peptides (20–80 μm) significantly inhibited the increase in the extracellular acidification response triggered by angiotensin II (ang II) in CHO-S cells transfected with the ang II type 1 receptor (AT1R-CHO-S). Subsequently, either alone or in a mixture, these peptides increased luciferase gene transcription in AT1R CHO-S cells stimulated with ang II and in HEK293 cells treated with isoproterenol. These peptides without transactivator failed to affect GPCR cellular responses. All four functional peptides were shown in vitro to competitively inhibit the degradation of a synthetic substrate by thimet oligopeptidase. Overexpression of thimet oligopeptidase in both CHO-S and HEK293 cells was sufficient to reduce luciferase activation triggered by a specific GPCR agonist. Moreover, using individual peptides as baits in affinity columns, several proteins involved in GPCR signaling were identified, including α-adaptin A and dynamin 1. These results suggest that before their complete degradation, intracellular peptides similar to those generated by proteasomes can actively affect cell signaling, probably representing additional bioactive molecules within cells. PMID:18617518

  6. Intracellular peptides as natural regulators of cell signaling.

    Science.gov (United States)

    Cunha, Fernanda M; Berti, Denise A; Ferreira, Zulma S; Klitzke, Clécio F; Markus, Regina P; Ferro, Emer S

    2008-09-05

    Protein degradation by the ubiquitin proteasome system releases large amounts of oligopeptides within cells. To investigate possible functions for these intracellularly generated oligopeptides, we fused them to a cationic transactivator peptide sequence using reversible disulfide bonds, introduced them into cells, and analyzed their effect on G protein-coupled receptor (GPCR) signal transduction. A mixture containing four of these peptides (20-80 microm) significantly inhibited the increase in the extracellular acidification response triggered by angiotensin II (ang II) in CHO-S cells transfected with the ang II type 1 receptor (AT1R-CHO-S). Subsequently, either alone or in a mixture, these peptides increased luciferase gene transcription in AT1R CHO-S cells stimulated with ang II and in HEK293 cells treated with isoproterenol. These peptides without transactivator failed to affect GPCR cellular responses. All four functional peptides were shown in vitro to competitively inhibit the degradation of a synthetic substrate by thimet oligopeptidase. Overexpression of thimet oligopeptidase in both CHO-S and HEK293 cells was sufficient to reduce luciferase activation triggered by a specific GPCR agonist. Moreover, using individual peptides as baits in affinity columns, several proteins involved in GPCR signaling were identified, including alpha-adaptin A and dynamin 1. These results suggest that before their complete degradation, intracellular peptides similar to those generated by proteasomes can actively affect cell signaling, probably representing additional bioactive molecules within cells.

  7. Intracellular speciation and transformation of inorganic mercury in marine phytoplankton.

    Science.gov (United States)

    Wu, Yun; Wang, Wen-Xiong

    2014-03-01

    Metal speciation is closely related to toxicity in aquatic organisms, but quantitative study of mercury transformation has rarely been reported. In this study, the ability of three marine phytoplankton species, including a green alga Chlorella autotrophica, a flagellate Isochrysis galbana and a diatom Thalassiosira weissflogii, to convert inorganic mercury were examined. We found that all algae tested were able to transform Hg(II) into dissolved gaseous mercury (DGM), phytochelatin (PC) complexes and metacinnabar (β-HgS). The most tolerant species, T. weissflogii, generally produced the highest level of PCs and β-HgS. Attributed to the highest DGM production ability, C. autotrophica accumulated the least Hg, but was the most sensitive due to low PC induction and β-HgS formation. Of the added Hg(II), less than 5% was reduced to DGM per day in all species. Of the intracellular Hg, <20% and 20-90% were chelated by PCs and transformed into β-HgS, respectively. These results suggest that intracellular biotransformation might be more important than bioavailability regulation in Hg(II) detoxification in marine phytoplankton.

  8. Bordetella pertussis entry into respiratory epithelial cells and intracellular survival.

    Science.gov (United States)

    Lamberti, Yanina; Gorgojo, Juan; Massillo, Cintia; Rodriguez, Maria E

    2013-12-01

    Bordetella pertussis is the causative agent of pertussis, aka whooping cough. Although generally considered an extracellular pathogen, this bacterium has been found inside respiratory epithelial cells, which might represent a survival strategy inside the host. Relatively little is known, however, about the mechanism of internalization and the fate of B. pertussis inside the epithelia. We show here that B. pertussis is able to enter those cells by a mechanism dependent on microtubule assembly, lipid raft integrity, and the activation of a tyrosine-kinase-mediated signaling. Once inside the cell, a significant proportion of the intracellular bacteria evade phagolysosomal fusion and remain viable in nonacidic lysosome-associated membrane-protein-1-negative compartments. In addition, intracellular B. pertussis was found able to repopulate the extracellular environment after complete elimination of the extracellular bacteria with polymyxin B. Taken together, these data suggest that B. pertussis is able to survive within respiratory epithelial cells and by this means potentially contribute to host immune system evasion.

  9. Detection of ubiquitinated huntingtin species in intracellular aggregates

    Directory of Open Access Journals (Sweden)

    Katrin eJuenemann

    2015-01-01

    Full Text Available Protein conformation diseases, including polyglutamine diseases, result from the accumulation and aggregation of misfolded proteins. Huntington’s disease is one of nine diseases caused by an expanded polyglutamine repeat within the affected protein and is hallmarked by intracellular inclusion bodies composed of aggregated N-terminal huntingtin fragments and other sequestered proteins. Fluorescence microscopy and filter trap assay are conventional methods to study protein aggregates, but cannot be used to analyze the presence and levels of post-translational modifications of aggregated huntingtin such as ubiquitination. Ubiquitination of proteins can be a signal for degradation and intracellular localization, but also affects protein activity and protein-protein interactions. The function of ubiquitination relies on its mono- and polymeric isoforms attached to protein substrates. Studying the ubiquitination pattern of aggregated huntingtin fragments offers an important possibility to understand huntingtin degradation and aggregation processes within the cell. For the identification of aggregated huntingtin and its ubiquitinated species, solubilization of the cellular aggregates is mandatory. Here we describe methods to identify post-translational modifications such as ubiquitination of aggregated mutant huntingtin. This approach is specifically described for use with mammalian cell culture and is suitable to study other disease-related proteins prone to aggregate.

  10. The genome sequences of Cellulomonas fimi and "Cellvibrio gilvus" reveal the cellulolytic strategies of two facultative anaerobes, transfer of "Cellvibrio gilvus" to the genus Cellulomonas, and proposal of Cellulomonas gilvus sp. nov.

    Directory of Open Access Journals (Sweden)

    Melissa R Christopherson

    Full Text Available Actinobacteria in the genus Cellulomonas are the only known and reported cellulolytic facultative anaerobes. To better understand the cellulolytic strategy employed by these bacteria, we sequenced the genome of the Cellulomonas fimi ATCC 484(T. For comparative purposes, we also sequenced the genome of the aerobic cellulolytic "Cellvibrio gilvus" ATCC 13127(T. An initial analysis of these genomes using phylogenetic and whole-genome comparison revealed that "Cellvibrio gilvus" belongs to the genus Cellulomonas. We thus propose to assign "Cellvibrio gilvus" to the genus Cellulomonas. A comparative genomics analysis between these two Cellulomonas genome sequences and the recently completed genome for Cellulomonas flavigena ATCC 482(T showed that these cellulomonads do not encode cellulosomes but appear to degrade cellulose by secreting multi-domain glycoside hydrolases. Despite the minimal number of carbohydrate-active enzymes encoded by these genomes, as compared to other known cellulolytic organisms, these bacteria were found to be proficient at degrading and utilizing a diverse set of carbohydrates, including crystalline cellulose. Moreover, they also encode for proteins required for the fermentation of hexose and xylose sugars into products such as ethanol. Finally, we found relatively few significant differences between the predicted carbohydrate-active enzymes encoded by these Cellulomonas genomes, in contrast to previous studies reporting differences in physiological approaches for carbohydrate degradation. Our sequencing and analysis of these genomes sheds light onto the mechanism through which these facultative anaerobes degrade cellulose, suggesting that the sequenced cellulomonads use secreted, multidomain enzymes to degrade cellulose in a way that is distinct from known anaerobic cellulolytic strategies.

  11. 兼氧混合菌对亚甲基蓝废水生物降解实验研究%Experimental Research on Bio-degradation of Methylene Blue Wastewater by Adopting Facultative Oxygen Mixed Bacterium

    Institute of Scientific and Technical Information of China (English)

    黄明强; 王伟艺; 叶宝水; 林丽云; 魏雪琳

    2015-01-01

    采用厦门某生物科技有限公司提供的兼氧混合菌为研究对象,利用相关设备进行中试实验,验证了该项技术的可行性。同时探究了重金属对该混合菌种生物降解能力的影响,通过中试实验研究后发现5mg/L的重铬酸钾和硫酸铜均对该混合菌种的生物降解能力有一定的影响,并且硫酸铜的影响大于重铬酸钾。此外该技术对甲基橙、刚果红废水也有一定的生物降解能力,这为兼氧混合菌用于印染废水处理提供了实验依据。%By adopting facultative oxygen mixed bacterium provided by a certain Xiamen bio-science and technology Co., Ltd as a research object, the paper uses the relevant equipment to carry out the primary production test, and validates the feasibility of the technology. At the same time, the paper probes into the impact of heavy metal on the bio-degradation capacity of the mixed bacterium genus. Through the primary production test, it shows a deifnite impact of 5mg/L potassium dichromate and bluestone on the bio-degradation capacity of the mixed bacterium genus, and also the impact of bluestone is greater than potassium dichromate. The technology has a deifnite bio-degradation capacity to the wastewater of methyl orange and Congo red. It shows that the facultative oxygen mixed bacterium used to treat the wastewaters of printing and dyeing, provides an experimental principle.

  12. Surveillance of antimicrobial susceptibility of aerobic and facultative Gram-negative bacilli isolated from patients with intra-abdominal infections in China: the 2002-2009 Study for Monitoring Antimicrobial Resistance Trends (SMART).

    Science.gov (United States)

    Yang, Qiwen; Wang, Hui; Chen, Minjun; Ni, Yuxing; Yu, Yunsong; Hu, Bijie; Sun, Ziyong; Huang, Wenxiang; Hu, Yunjian; Ye, Huifen; Badal, Robert E; Xu, Yingchun

    2010-12-01

    The objective of this study was to investigate the distribution and susceptibility of aerobic and facultative Gram-negative bacilli (GNB) isolated from patients with intra-abdominal infections (IAIs) in China. From 2002 to 2009, minimum inhibitory concentrations of 14 antibiotics for 3420 aerobic and facultative GNB from up to eight hospitals in six cities were determined by the broth microdilution method. Enterobacteriaceae comprised 82.9% (2834/3420) of the total isolates, with Escherichia coli (49.2%) being the most commonly isolated species followed by Klebsiella pneumoniae (17.0%), Enterobacter cloacae (5.8%) and Citrobacter freundii (2.3%). Amongst the antimicrobial agents tested, the three carbapenems (ertapenem, imipenem and meropenem) were the most active agents against Enterobacteriaceae, with susceptibility rates of 96.1-99.6% (2002-2009), 98.2-100% (2002-2009) and 99.6-100% (2002-2004), respectively, followed by amikacin (86.8-95.1%) and piperacillin/tazobactam (84.5-94.3%). Susceptibility rates of all tested third- and fourth-generation cephalosporins against Enterobacteriaceae declined by nearly 30%, with susceptibility rates of 40.2%, 39.1%, 56.3% and 51.8% in 2009 for ceftriaxone, cefotaxime, ceftazidime and cefepime, respectively. The occurrence of extended-spectrum β-lactamases increased rapidly, especially for E. coli (from 20.8% in 2002 to 64.9% in 2009). Susceptibility of E. coli to ciprofloxacin decreased from 57.6% in 2002 to 24.2% in 2009. The least active agent against Enterobacteriaceae was ampicillin/sulbactam (SAM) (25.3-44.3%). In conclusion, Enterobacteriaceae were the major pathogens causing IAIs, and carbapenems retained the highest susceptibility rates over the 8-year study period. Third- and fourth-generation cephalosporins, fluoroquinolones and SAM may not be ideal choices for empirical therapy of IAIs in China.

  13. 极地适冷菌Pseudoalteromonas sp.QI-1产适冷蛋白酶发酵条件的优化%Optimization of cold-active protease production by polar psychrotrophic bacterium Pseudoalteromonas sp.QI-1

    Institute of Scientific and Technical Information of China (English)

    徐国英; 林学政; 王能飞; 朱启忠

    2011-01-01

    The culture conditions of polar psychrotrophic bacterium Pseudoalteromonas sp. QI-1 were optimized for the production of cold-active protease. The optimum fermentation conditions were determined as follows: optimal temperatures for growth and enzyme production were both at 5 ℃, inoculation volume was 1% ,and liquid volume was 10% at pH 5. Culture medium for protease production of strain QI-1 was also studied. Results showed that nitrogen source and carbon source preferred by protease production were bran and sodium acetate. 2% NaC1,10 mmol/L Mg2+ ,0.5% Tween-80 and 0.75% casein were suitable for the enzyme production. The orthogonal experiment design showed that the optimum medium (per liter) consisted of 5 g bran,2.5 g yeast extract,3 g casein,3 g MgCl2 ·6H20 and 1.5 g KCl. Under the optimum condition, the protease activity was up to 166. 20 U/mL and the yield of protease increased about 56%.%对极地适冷菌Pseudoalteromonas sp.QI-1产适冷蛋白酶的发酵条件进行优化.结果表明:菌株QI-1的最适生长和产酶温度均为5℃;最佳接种量为1%;发酵培养基的最适初始pH和最佳装样量分别为5和10%;盐度为2%时对菌株的生长和产酶最为有利;麸皮和醋酸钠分别为最佳N源和C源;添加0.75%酪蛋白时菌株QI-1胞外蛋白酶的活性最高;10 mmol/L Mg2+和0.5%Tween-80有利于产酶.正交试验结果表明:菌株Pseudoalteromonas sp.QI-1产蛋白酶较佳培养基配方(g/L)为麸皮5,酵母粉2.5,酪蛋白3,MgCl2·6H2O3,KCl 1.5;发酵液比酶活为166.20 U/mL,较优化前提高了约56%.

  14. Tumour Cell Labelling by Magnetic Nanoparticles with Determination of Intracellular Iron Content and Spatial Distribution of the Intracellular Iron

    Directory of Open Access Journals (Sweden)

    Alfred Cuschieri

    2013-04-01

    Full Text Available Magnetically labelled cells are used for in vivo cell tracking by MRI, used for the clinical translation of cell-base therapies. Studies involving magnetic labelled cells may include separation of labelled cells, targeted delivery and controlled release of drugs, contrast enhanced MRI and magnetic hyperthermia for the in situ ablation of tumours. Dextran-coated super-paramagnetic iron oxide (SPIO ferumoxides are used clinically as an MR contrast agents primarily for hepatic imaging. The material is also widely used for in vitro cell labelling, as are other SPIO-based particles. Our results on the uptake by human cancer cell lines of ferumoxides indicate that electroporation in the presence of protamine sulphate (PS results in rapid high uptake of SPIO nanoparticles (SPIONs by parenchymal tumour cells without significant impairment of cell viability. Quantitative determination of cellular iron uptake performed by colorimetric assay is in agreement with data from the literature. These results on intracellular iron content together with the intracellular distribution of SPIONs by magnetic force microscopy (MFM following in vitro uptake by parenchymal tumour cells confirm the potential of this technique for clinical tumour cell detection and destruction.

  15. Detection of intracellular bacterial communities in human urinary tract infection.

    Directory of Open Access Journals (Sweden)

    David A Rosen

    2007-12-01

    Full Text Available BACKGROUND: Urinary tract infections (UTIs are one of the most common bacterial infections and are predominantly caused by uropathogenic Escherichia coli (UPEC. While UTIs are typically considered extracellular infections, it has been recently demonstrated that UPEC bind to, invade, and replicate within the murine bladder urothelium to form intracellular bacterial communities (IBCs. These IBCs dissociate and bacteria flux out of bladder facet cells, some with filamentous morphology, and ultimately establish quiescent intracellular reservoirs that can seed recurrent infection. This IBC pathogenic cycle has not yet been investigated in humans. In this study we sought to determine whether evidence of an IBC pathway could be found in urine specimens from women with acute UTI. METHODS AND FINDINGS: We collected midstream, clean-catch urine specimens from 80 young healthy women with acute uncomplicated cystitis and 20 asymptomatic women with a history of UTI. Investigators were blinded to culture results and clinical history. Samples were analyzed by light microscopy, immunofluorescence, and electron microscopy for evidence of exfoliated IBCs and filamentous bacteria. Evidence of IBCs was found in 14 of 80 (18% urines from women with UTI. Filamentous bacteria were found in 33 of 80 (41% urines from women with UTI. None of the 20 urines from the asymptomatic comparative group showed evidence of IBCs or filaments. Filamentous bacteria were present in all 14 of the urines with IBCs compared to 19 (29% of 66 samples with no evidence of IBCs (p < 0.001. Of 65 urines from patients with E. coli infections, 14 (22% had evidence of IBCs and 29 (45% had filamentous bacteria, while none of the gram-positive infections had IBCs or filamentous bacteria. CONCLUSIONS: The presence of exfoliated IBCs and filamentous bacteria in the urines of women with acute cystitis suggests that the IBC pathogenic pathway characterized in the murine model may occur in humans. The

  16. Autophagy Induced by Intracellular Infection of Propionibacterium acnes

    Science.gov (United States)

    Nakamura, Teruko; Furukawa, Asuka; Uchida, Keisuke; Ogawa, Tomohisa; Tamura, Tomoki; Sakonishi, Daisuke; Wada, Yuriko; Suzuki, Yoshimi; Ishige, Yuki; Minami, Junko; Akashi, Takumi

    2016-01-01

    Background Sarcoidosis is caused by Th1-type immune responses to unknown agents, and is linked to the infectious agent Propionibacterium acnes. Many strains of P. acnes isolated from sarcoid lesions cause intracellular infection and autophagy may contribute to the pathogenesis of sarcoidosis. We examined whether P. acnes induces autophagy. Methods Three cell lines from macrophages (Raw264.7), mesenchymal cells (MEF), and epithelial cells (HeLa) were infected by viable or heat-killed P. acnes (clinical isolate from sarcoid lymph node) at a multiplicity of infection (MOI) of 100 or 1000 for 1 h. Extracellular bacteria were killed by washing and culturing infected cells with antibiotics. Samples were examined by colony assay, electron-microscopy, and fluorescence-microscopy with anti-LC3 and anti-LAMP1 antibodies. Autophagy-deficient (Atg5-/-) MEF cells were also used. Results Small and large (≥5 μm in diameter) LC3-positive vacuoles containing few or many P. acnes cells (LC3-positive P. acnes) were frequently found in the three cell lines when infected by viable P. acnes at MOI 1000. LC3-positive large vacuoles were mostly LAMP1-positive. A few small LC3-positive/LAMP1-negative vacuoles were consistently observed in some infected cells for 24 h postinfection. The number of LC3-positive P. acnes was decreased at MOI 100 and completely abolished when heat-killed P. acnes was used. LC3-positive P. acnes was not found in autophagy-deficient Atg5-/- cells where the rate of infection was 25.3 and 17.6 times greater than that in wild-type Atg5+/+ cells at 48 h postinfection at MOI 100 and 1000, respectively. Electron-microscopic examination revealed bacterial cells surrounded mostly by a single-membrane including the large vacuoles and sometimes a double or multi-layered membrane, with occasional undigested bacterial cells in ruptured late endosomes or in the cytoplasm. Conclusion Autophagy was induced by intracellular P. acnes infection and contributed to intracellular

  17. Extraction of intracellular protein from Glaciozyma antarctica for proteomics analysis

    Science.gov (United States)

    Faizura, S. Nor; Farahayu, K.; Faizal, A. B. Mohd; Asmahani, A. A. S.; Amir, R.; Nazalan, N.; Diba, A. B. Farah; Muhammad, M. Nor; Munir, A. M. Abdul

    2013-11-01

    Two preparation methods of crude extracts of psychrophilic yeast Glaciozyma antarctica were compared in order to obtain a good recovery of intracellular proteins. Extraction with mechanical procedures using sonication was found to be more effective for obtaining good yield compare to alkaline treatment method. The procedure is simple, rapid, and produce better yield. A total of 52 proteins were identified by combining both extraction methods. Most of the proteins identified in this study involves in the metabolic process including glycolysis pathway, pentose phosphate pathway, pyruyate decarboxylation and also urea cyle. Several chaperons were identified including probable cpr1-cyclophilin (peptidylprolyl isomerase), macrolide-binding protein fkbp12 and heat shock proteins which were postulate to accelerate proper protein folding. Characteristic of the fundamental cellular processes inferred from the expressed-proteome highlight the evolutionary and functional complexity existing in this domain of life.

  18. Variety in intracellular diffusion during the cell cycle

    DEFF Research Database (Denmark)

    Selhuber-Unkel, C.; Yde, P.; Berg-Sørensen, Kirstine;

    2009-01-01

    During the cell cycle, the organization of the cytoskeletal network undergoes dramatic changes. In order to reveal possible changes of the viscoelastic properties in the intracellular space during the cell cycle we investigated the diffusion of endogenous lipid granules within the fission yeast...... Schizosaccharomyces Pombe using optical tweezers. The cell cycle was divided into interphase and mitotic cell division, and the mitotic cell division was further subdivided in its stages. During all stages of the cell cycle, the granules predominantly underwent subdiffusive motion, characterized by an exponent...... a that is also linked to the viscoelastic moduli of the cytoplasm. The exponent a was significantly smaller during interphase than during any stage of the mitotic cell division, signifying that the cytoplasm was more elastic during interphase than during division. We found no significant differences...

  19. Semi-Discrete Systems and Intracellular Calcium Dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Pearson, J.; Dawson, S.P.; Mitkov, I.

    1998-10-24

    Intracellular calcium is sequestered in closed membranes such as the sarcoplasmic or endoplasmic reticula and released at discretely distributed protein/receptor channels. The release kinetics can result in the propagation of waves of elevated calcium concentration. The main physical processes are reactions at the release sites and diffusion between the sites. The theory of chemical wave propagation in reaction-diffusion systems is in large part devoted to the study of systems in which there are no extrinsic inhomogeneities. The discrete distribution of the release sites plays a key role in determining the nature of the propagating wave. The authors analyze some simple reaction-diffusion models in order to elucidate the role of discreteness for chemical wave propagation.

  20. Intracellular Signaling Mediators in the Circulatory and Ventilatory Systems

    CERN Document Server

    Thiriet, Marc

    2013-01-01

    The volumes in this authoritative series present a multidisciplinary approach to modeling and simulation of flows in the cardiovascular and ventilatory systems, especially multiscale modeling and coupled simulations. The cardiovascular and respiratory systems are tightly coupled, as their primary function is to supply oxygen to and remove carbon dioxide from the body's cells. Because physiological conduits have deformable and reactive walls, macroscopic flow behavior and prediction must be coupled to phenomenological models of nano- and microscopic events in a corrector scheme of regulated mechanisms when the vessel lumen caliber varies markedly. Therefore, investigation of flows of blood and air in physiological conduits requires an understanding of the biology, chemistry, and physics of these systems together with the mathematical tools to describe their functioning. Volume 4 is devoted to major sets of intracellular mediators that transmit signals upon stimulation of cell-surface receptors.  Activation of...

  1. Intracellular behaviour of samarium and europium in lactating mammary gland

    Institute of Scientific and Technical Information of China (English)

    Ayadi Ahlem; Maghraoui Samira; El Hili Ali; Galle Pierre; Tekaya Leila

    2012-01-01

    The subcellular localization of samarium and europium,two rare-earths,increasingly used in both medical and industrial fields,has been studied in several organs such as liver and kidney but never in the mammary gland despite of its importance in the biology of lactation and nutrition domains.The intracellular behaviour of samarium and europium after their intra-peritoneal administration in the lactating mammary gland cells was investigated.The results showed the presence of very electron dense deposits in the glandular epithelial cell lysosomes.These particular lysosomes were never observed in the marnrnary cell lysosomes of control rats.These intralysosomal deposits were probably composed of insoluble samarium or europium phosphates by analogy with previous studies,the transmission electron microscopy,the ion mass microscopy and the electron probe microanalysis,and other techniques allowing the identification of the chemical structure of the intralysosomal deposits.

  2. Intracellular Production of Cyclic Peptide Libraries with SICLOPPS.

    Science.gov (United States)

    Osher, Eliot L; Tavassoli, Ali

    2017-01-01

    Cyclic peptides are an important class of molecules that are increasingly viewed as an ideal scaffold for inhibition of protein-protein interactions (PPI). Here we detail an approach that enables the intracellular synthesis of cyclic peptide libraries of around 10(8) members. The method utilizes split intein mediated circular ligation of peptides and proteins (SICLOPPS), taking advantage of split intein splicing to cyclize a library of peptide sequences. SICLOPPS allows the ring size, set residues and number of random residues within a library to be predetermined by the user. SICLOPPS libraries have been combined with a variety of cell-based screens to identify cyclic peptide inhibitors of a variety of enzymes and protein-protein interactions.

  3. Intracellular pH-determination by fluorescence measurements.

    Science.gov (United States)

    Visser, J W; Jongeling, A A; Tanke, H J

    1979-01-01

    A method was developed to determine the intracellular pH (pHi) of individual cells by use of fluorescence measurements. The method is based on the observation that the fluorescence excitation spectrum of fluorescein is pH-dependent. Fluorescence excitation spectra from individual rat bone marrow cells treated with fluorescein diacetate (FDA) were compared with those of fluorescein solutions of known pH values. Cells which were suspended in media of pH between 4.0 and 8.1 with high to normal buffering capacities had pHi values equal to those of the media. Cells suspended in media with low buffering capacities maintained a pH,i of 6.7 +/- 0.2. Preliminary results indicated that the pHi of individual cells may also be determined by using flow cytometry.

  4. Measuring intracellular redox conditions using GFP-based sensors

    DEFF Research Database (Denmark)

    Björnberg, Olof; Ostergaard, Henrik; Winther, Jakob R

    2006-01-01

    Recent years have seen the development of methods for analyzing the redox conditions in specific compartments in living cells. These methods are based on genetically encoded sensors comprising variants of Green Fluorescent Protein in which vicinal cysteine residues have been introduced at solvent......-exposed positions. Several mutant forms have been identified in which formation of a disulfide bond between these cysteine residues results in changes of their fluorescence properties. The redox sensors have been characterized biochemically and found to behave differently, both spectroscopically and in terms...... of redox properties. As genetically encoded sensors they can be expressed in living cells and used for analysis of intracellular redox conditions; however, which parameters are measured depends on how the sensors interact with various cellular redox components. Results of both biochemical and cell...

  5. Giardia lamblia: intracellular localization of alpha8-giardin.

    Science.gov (United States)

    Wei, Chao Jun; Tian, Xi Feng; Adam, Rodney D; Lu, Si Qi

    2010-12-01

    Alpha8-giardin (α8-giardin) is a member of the multi-gene α-giardin family in the intestinal parasitic protozoan, Giardia lamblia. This gene family shares an ancestry with the annexin super family, whose common characteristic is calcium dependent binding to membranes that contain acidic phospholipids. In the present study, the antigenicity, hydrophilicity, flexibility, surface probability, and secondary structure of α8-giardin amino acids were predicted by bioinformatics applications. A specific anti-peptide antiserum, anti-P3, was used to determine the intracellular location of α8-giardin with confocal immunofluorescence microscopy and immunoelectron microscopy. The results indicated that α8-giardin was located on the plasma membrane and flagella, but not on the ventral disk. Reduction of α8-giardin transcript levels by ribozyme-mediated cleavage decreased trophozoite motility and growth rate, indicating the functional importance of α8-giardin to Giardia trophozoite biology.

  6. Signal Transduction and Intracellular Trafficking by the Interleukin 36 Receptor*

    Science.gov (United States)

    Saha, Siddhartha S.; Singh, Divyendu; Raymond, Ernest L.; Ganesan, Rajkumar; Caviness, Gary; Grimaldi, Christine; Woska, Joseph R.; Mennerich, Detlev; Brown, Su-Ellen; Mbow, M. Lamine; Kao, C. Cheng

    2015-01-01

    Improper signaling of the IL-36 receptor (IL-36R), a member of the IL-1 receptor family, has been associated with various inflammation-associated diseases. However, the requirements for IL-36R signal transduction remain poorly characterized. This work seeks to define the requirements for IL-36R signaling and intracellular trafficking. In the absence of cognate agonists, IL-36R was endocytosed and recycled to the plasma membrane. In the presence of IL-36, IL-36R increased accumulation in LAMP1+ lysosomes. Endocytosis predominantly used a clathrin-mediated pathway, and the accumulation of the IL-36R in lysosomes did not result in increased receptor turnover. The ubiquitin-binding Tollip protein contributed to IL-36R signaling and increased the accumulation of both subunits of the IL-36R. PMID:26269592

  7. Opposing Biological Functions of Tryptophan Catabolizing Enzymes During Intracellular Infection

    Science.gov (United States)

    Divanovic, Senad; Sawtell, Nancy M.; Trompette, Aurelien; Warning, Jamie I.; Dias, Alexandra; Cooper, Andrea M.; Yap, George S.; Arditi, Moshe; Shimada, Kenichi; DuHadaway, James B.; Prendergast, George C.; Basaraba, Randall J.; Mellor, Andrew L.; Munn, David H.; Aliberti, Julio

    2012-01-01

    Recent studies have underscored physiological and pathophysiological roles for the tryptophan-degrading enzyme indolamine 2,3-dioxygenase (IDO) in immune counterregulation. However, IDO was first recognized as an antimicrobial effector, restricting tryptophan availability to Toxoplasma gondii and other pathogens in vitro. The biological relevance of these findings came under question when infectious phenotypes were not forthcoming in IDO-deficient mice. The recent discovery of an IDO homolog, IDO-2, suggested that the issue deserved reexamination. IDO inhibition during murine toxoplasmosis led to 100% mortality, with increased parasite burdens and no evident effects on the immune response. Similar studies revealed a counterregulatory role for IDO during leishmaniasis (restraining effector immune responses and parasite clearance), and no evident role for IDO in herpes simplex virus type 1 (HSV-1) infection. Thus, IDO plays biologically important roles in the host response to diverse intracellular infections, but the dominant nature of this role—antimicrobial or immunoregulatory—is pathogen-specific. PMID:21990421

  8. Anticipatory Behavior and Intracellular Communication in Physarum polycephalum

    Science.gov (United States)

    Shirakawa, Tomohiro; Gunji, Yukio-Pegio

    2006-06-01

    We used the plasmodium of Physarum polycephalum to investigate a communicative property of a cell. We allowed the plasmodium to form a tubular network that connected three food sources, which were arranged to be located at the vertices of an equilateral triangle. The global condition of the plasmodium is changed by the connection of two networks in such a way that the local solution of each network was not affected. We used several criteria to evaluate the character of the network. The change in the global condition appeared to cause significant difference in the network properties, and two connected networks showed homologous network formation. These data imply the presence of intracellular communication and anticipatory aspect in their communication.

  9. Intracellular insulin in human tumors: examples and implications

    Directory of Open Access Journals (Sweden)

    Radulescu Razvan T

    2011-04-01

    Full Text Available Abstract Insulin is one of the major metabolic hormones regulating glucose homeostasis in the organism and a key growth factor for normal and neoplastic cells. Work conducted primarily over the past 3 decades has unravelled the presence of insulin in human breast cancer tissues and, more recently, in human non-small cell lung carcinomas (NSCLC. These findings have suggested that intracellular insulin is involved in the development of these highly prevalent human tumors. A potential mechanism for such involvement is insulin's binding and inactivation of the retinoblastoma tumor suppressor protein (RB which in turn is likely controlled by insulin-degrading enzyme (IDE. This model and its supporting data are collectively covered in this survey in order to provide further insight into insulin-driven oncogenesis and its reversal through future anticancer therapeutics.

  10. Coupling of Active Motion and Advection Shapes Intracellular Cargo Transport

    CERN Document Server

    Trong, P Khuc; Goldstein, R E; 10.1103/PhysRevLett.109.028104

    2012-01-01

    Intracellular cargo transport can arise from passive diffusion, active motor-driven transport along cytoskeletal filament networks, and passive advection by fluid flows entrained by such motor/cargo motion. Active and advective transport are thus intrinsically coupled as related, yet different representations of the same underlying network structure. A reaction-advection-diffusion system is used here to show that this coupling affects the transport and localization of a passive tracer in a confined geometry. For sufficiently low diffusion, cargo localization to a target zone is optimized either by low reaction kinetics and decoupling of bound and unbound states, or by a mostly disordered cytoskeletal network with only weak directional bias. These generic results may help to rationalize subtle features of cytoskeletal networks, for example as observed for microtubules in fly oocytes.

  11. The Growth Hormone Secretagogue Receptor: Its Intracellular Signaling and Regulation

    Directory of Open Access Journals (Sweden)

    Yue Yin

    2014-03-01

    Full Text Available The growth hormone secretagogue receptor (GHSR, also known as the ghrelin receptor, is involved in mediating a wide variety of biological effects of ghrelin, including: stimulation of growth hormone release, increase of food intake and body weight, modulation of glucose and lipid metabolism, regulation of gastrointestinal motility and secretion, protection of neuronal and cardiovascular cells, and regulation of immune function. Dependent on the tissues and cells, activation of GHSR may trigger a diversity of signaling mechanisms and subsequent distinct physiological responses. Distinct regulation of GHSR occurs at levels of transcription, receptor interaction and internalization. Here we review the current understanding on the intracellular signaling pathways of GHSR and its modulation. An overview of the molecular structure of GHSR is presented first, followed by the discussion on its signaling mechanisms. Finally, potential mechanisms regulating GHSR are reviewed.

  12. Neutrophils and intracellular pathogens: beyond phagocytosis and killing.

    Science.gov (United States)

    Appelberg, Rui

    2007-02-01

    Neutrophils are not simply scavenging phagocytes that clear extracellular spaces of rapidly proliferating microbes; they are also active in the control of infections by intracellular pathogens. Several mechanisms for nonphagocytic roles of neutrophils in protective immunity have been put forth over the years but further evidence has recently been accumulating at an increasing pace. In this review, I present the evidence that suggests neutrophils are involved in pathogen shuttling into the lymphoid tissues, in antigen presentation, and in early T cell recruitment and initiation of granuloma organization. Also, a clearer view on the antimicrobial molecules that can be acquired by macrophages to enhance their antimicrobial activity is now emerging. Finally, neutrophils can adversely affect immunity against certain parasites by causing immune deviation.

  13. Macromolecularly "Caged" Carbon Nanoparticles for Intracellular Trafficking via Switchable Photoluminescence.

    Science.gov (United States)

    Misra, Santosh K; Srivastava, Indrajit; Tripathi, Indu; Daza, Enrique; Ostadhossein, Fatemeh; Pan, Dipanjan

    2017-02-08

    Reversible switching of photoluminescence (PL) of carbon nanoparticles (CNP) can be achieved with counterionic macromolecular caging and decaging at the nanoscale. A negatively charged uncoated, "bare" CNP with high luminescence loses its PL when positively charged macromolecules are wrapped around its surface. Prepared caged carbons could regain their emission only through interaction with anionic surfactant molecules, representing anionic amphiphiles of endocytic membranes. This process could be verified by gel electrophoresis, spectroscopically and in vitro confocal imaging studies. Results indicated for the first time that luminescence switchable CNPs can be synthesized for efficient intracellular tracking. This study further supports the origin of photoluminescence in CNP as a surface phenomenon correlated a function of characteristic charged macromolecules.

  14. Cytoplasmic tail of coronavirus spike protein has intracellular targeting signals

    Indian Academy of Sciences (India)

    JIBIN SADASIVAN; MANMEET SINGH; JAYASRI DAS SARMA

    2017-06-01

    Intracellular trafficking and localization studies of spike protein from SARS and OC43 showed that SARS spikeprotein is localized in the ER or ERGIC compartment and OC43 spike protein is predominantly localized in thelysosome. Differential localization can be explained by signal sequence. The sequence alignment using Clustal Wshows that the signal sequence present at the cytoplasmic tail plays an important role in spike protein localization. Aunique GYQEL motif is identified at the cytoplasmic terminal of OC43 spike protein which helps in localization in thelysosome, and a novel KLHYT motif is identified in the cytoplasmic tail of SARS spike protein which helps in ER orERGIC localization. This study sheds some light on the role of cytoplasmic tail of spike protein in cell-to-cell fusion,coronavirus host cell fusion and subsequent pathogenicity.

  15. Diverse intracellular pathogens activate type III interferon expression from peroxisomes.

    Science.gov (United States)

    Odendall, Charlotte; Dixit, Evelyn; Stavru, Fabrizia; Bierne, Helene; Franz, Kate M; Durbin, Ann Fiegen; Boulant, Steeve; Gehrke, Lee; Cossart, Pascale; Kagan, Jonathan C

    2014-08-01

    Type I interferon responses are considered the primary means by which viral infections are controlled in mammals. Despite this view, several pathogens activate antiviral responses in the absence of type I interferons. The mechanisms controlling type I interferon-independent responses are undefined. We found that RIG-I like receptors (RLRs) induce type III interferon expression in a variety of human cell types, and identified factors that differentially regulate expression of type I and type III interferons. We identified peroxisomes as a primary site of initiation of type III interferon expression, and revealed that the process of intestinal epithelial cell differentiation upregulates peroxisome biogenesis and promotes robust type III interferon responses in human cells. These findings highlight the importance of different intracellular organelles in specific innate immune responses.

  16. Intracellular Events and Cell Fate in Filovirus Infection

    Directory of Open Access Journals (Sweden)

    Elena Ryabchikova

    2011-08-01

    Full Text Available Marburg and Ebola viruses cause a severe hemorrhagic disease in humans with high fatality rates. Early target cells of filoviruses are monocytes, macrophages, and dendritic cells. The infection spreads to the liver, spleen and later other organs by blood and lymph flow. A hallmark of filovirus infection is the depletion of non-infected lymphocytes; however, the molecular mechanisms leading to the observed bystander lymphocyte apoptosis are poorly understood. Also, there is limited knowledge about the fate of infected cells in filovirus disease. In this review we will explore what is known about the intracellular events leading to virus amplification and cell damage in filovirus infection. Furthermore, we will discuss how cellular dysfunction and cell death may correlate with disease pathogenesis.

  17. Intracellular transport mechanisms: a critique of diffusion theory.

    Science.gov (United States)

    Agutter, P S; Malone, P C; Wheatley, D N

    1995-09-21

    It is argued that Brownian motion makes a less significant contribution to the movements of molecules and particles inside cells than is commonly believed, and that the numbers of similar molecules and particles within any near-homogeneous subcompartment of the cell internum are insufficient to justify the statistical assumptions implicit in the derivation of the diffusion equation. For these reasons, it is contended that, contrary to accepted opinion, diffusion theory cannot provide an explanation for intracellular transport at the molecular level. Although attempts have been made to adapt diffusion theory to complex media, the conclusion is that none satisfactorily overcomes the problem of applying the theory to cell biology. However, the heuristic influence of the theory on cellular biophysics and physiology is noted, and possible alternative frameworks for interpreting the valuable experimental data obtained from such studies are outlined.

  18. Intracellular pH in rat pancreatic ducts

    DEFF Research Database (Denmark)

    Novak, I; Hug, M; Greger, R

    1997-01-01

    In order to study the mechanism of H+ and HCO3- transport in a HCO3- secreting epithelium, pancreatic ducts, we have measured the intracellular pH (pHi) in this tissue using the pH sensitive probe BCECF. We found that exposures of ducts to solutions containing acetate/acetic acid or NH4+/NH3...... buffers (20 mmol/l) led to pHi changes in accordance with entry of lipid-soluble forms of the buffers, followed by back-regulation of pHi by duct cells. In another type of experiment, changes in extracellular pH of solutions containing HEPES or HCO3-/CO2 buffers led to significant changes in pHi that did....... Under some conditions, these exchangers can be invoked to regulate cell pH....

  19. Bioreducible Lipid-like Nanoparticles for Intracellular Protein Delivery

    Science.gov (United States)

    Arellano, Carlos Luis

    Protein-based therapy is one of the most direct ways to manipulate cell function and treat human disease. Although protein therapeutics has made its way to clinical practice, with five of the top fifteen global pharmaceuticals being peptide or protein-based drugs, one common limitation is that the effects of protein therapy are only achieved through the targeting of cell surface receptors and intracellular domains. Due to the impermeability of the cell membrane to most foreign materials, entire classes of potentially therapeutic proteins cannot thoroughly be studied without a safe and efficient method of transporting proteins into the cytosol. We report the use of a combinatorially-designed bioreducible lipid-like material (termed "lipidoid") - based protein delivery platform for the transfection of human cancer cell lines. Lipidoid nanoparticles are synthesized through a thin film dispersion method. The degradation of the bioreducible nanoparticles was observed when exposed to glutathione, a highly reductive compound present in the cytosol. We demonstrate that the nanoparticles are capable of transfecting a dose-dependent concentration of our model protein, beta-galactosidase into HeLa cells. Furthermore, formulations of the lipidoid containing the cytotoxic proteins saporin and RNase-A are both capable of inhibiting tumor cell proliferation as observed in in vitro treatment of different human cancer cell lines. There was no observed loss in protein activity after lyophilization and long--term storage, indicating the potential of pre-clinical applications. Overall, we demonstrate an effective approach to protein formulation and intracellular delivery. We believe that our formulations will lead to the study of a whole class of previously untapped therapeutics that may generate new solutions for previously untreatable diseases.

  20. Intracellular localization of GASP/ECOP/VOPP1.

    Science.gov (United States)

    Baras, Alexander; Moskaluk, Christopher A

    2010-04-01

    Vesicular Over-expressed in cancer Prosurvival Protein 1 (VOPP1), also known as Glioblastoma Amplified and Secreted Protein and EGFR-Coamplified and Over-expressed Protein has been previously shown to be over-expressed in human glioblastoma multiforme and squamous cell carcinoma. Additionally, previous experimental work suggests that it confers a prosurvival cellular phenotype. A query of a public database of gene expression profiling data (Oncomine) shows that the VOPP1 transcript is also highly expressed in several other common human cancers, including breast carcinoma, pancreatic carcinoma, and lymphoma. Analysis of VOPP1 sequence structure shows both a signal sequence and a transmembrane domain, and examination of a public microarray dataset for endoplasmic reticulum (ER)-bound mRNA transcripts is consistent with the VOPP1 protein product being synthesized into the ER. Immunoblot analysis of cell culture and conditioned media confirms that the protein product is not secreted and is retained intracellularly. VOPP1 protein tagged with a fluorescence reporter, as well as antibody-mediated visualization of recombinant and native forms of the protein reveals an intracellular vesicular pattern of localization. Co-localization experiments reveal that VOPP1 vesicles do not co-localize with mitochondria or peroxisomes, but show partial co-localization with perinuclear lysosomes. Additionally, markers of endocytosis and autophagy show partial perinuclear co-localization, suggesting that VOPP1-containing vesicles enter final common pathways of the lysosomal system. These findings throw into doubt the hypothesis that VOPP1 interacts directly with cytoplasmic mediators of the NF kappa B pathway, and suggest that the prosurvival phenotype conferred by this gene product is mediated by other mechanisms.

  1. Intracellular Ascorbate Prevents Endothelial Barrier Permeabilization by Thrombin.

    Science.gov (United States)

    Parker, William H; Qu, Zhi-chao; May, James M

    2015-08-28

    Intracellular ascorbate (vitamin C) has previously been shown to tighten the endothelial barrier and maintain barrier integrity during acute inflammation in vitro. However, the downstream effectors of ascorbate in the regulation of endothelial permeability remain unclear. In this study, we evaluated ascorbate as a mediator of thrombin-induced barrier permeabilization in human umbilical vein endothelial cells and their immortalized hybridoma line, EA.hy926. We found that the vitamin fully prevented increased permeability to the polysaccharide inulin by thrombin in a dose-dependent manner, and it took effect both before and after subjection to thrombin. Thrombin exposure consumed intracellular ascorbate but not the endogenous antioxidant GSH. Likewise, the antioxidants dithiothreitol and tempol did not reverse permeabilization. We identified a novel role for ascorbate in preserving cAMP during thrombin stimulation, resulting in two downstream effects. First, ascorbate maintained the cortical actin cytoskeleton in a Rap1- and Rac1-dependent manner, thus preserving stable adherens junctions between adjacent cells. Second, ascorbate prevented actin polymerization and formation of stress fibers by reducing the activation of RhoA and phosphorylation of myosin light chain. Although ascorbate and thrombin both required calcium for their respective effects, ascorbate did not prevent thrombin permeabilization by obstructing calcium influx. However, preservation of cAMP by ascorbate was found to depend on both the production of nitric oxide by endothelial nitric-oxide synthase, which ascorbate is known to activate, and the subsequent generation cGMP by guanylate cyclase. Together, these data implicate ascorbate in the prevention of inflammatory endothelial barrier permeabilization and explain the underlying signaling mechanism.

  2. Carbon Dot Nanothermometry: Intracellular Photoluminescence Lifetime Thermal Sensing.

    Science.gov (United States)

    Kalytchuk, Sergii; Poláková, Kateřina; Wang, Yu; Froning, Jens P; Cepe, Klara; Rogach, Andrey L; Zbořil, Radek

    2017-02-28

    Nanoscale biocompatible photoluminescence (PL) thermometers that can be used to accurately and reliably monitor intracellular temperatures have many potential applications in biology and medicine. Ideally, such nanothermometers should be functional at physiological pH across a wide range of ionic strengths, probe concentrations, and local environments. Here, we show that water-soluble N,S-co-doped carbon dots (CDs) exhibit temperature-dependent photoluminescence lifetimes and can serve as highly sensitive and reliable intracellular nanothermometers. PL intensity measurements indicate that these CDs have many advantages over alternative semiconductor- and CD-based nanoscale temperature sensors. Importantly, their PL lifetimes remain constant over wide ranges of pH values (5-12), CD concentrations (1.5 × 10(-5) to 0.5 mg/mL), and environmental ionic strengths (up to 0.7 mol·L(-1) NaCl). Moreover, they are biocompatible and nontoxic, as demonstrated by cell viability and flow cytometry analyses using NIH/3T3 and HeLa cell lines. N,S-CD thermal sensors also exhibit good water dispersibility, superior photo- and thermostability, extraordinary environment and concentration independence, high storage stability, and reusability-their PL decay curves at temperatures between 15 and 45 °C remained unchanged over seven sequential experiments. In vitro PL lifetime-based temperature sensing performed with human cervical cancer HeLa cells demonstrated the great potential of these nanosensors in biomedicine. Overall, N,S-doped CDs exhibit excitation-independent emission with strongly temperature-dependent monoexponential decay, making them suitable for both in vitro and in vivo luminescence lifetime thermometry.

  3. Elevated Cholesterol in the Coxiella burnetii Intracellular Niche Is Bacteriolytic

    Science.gov (United States)

    Mulye, Minal; Samanta, Dhritiman; Winfree, Seth; Heinzen, Robert A.

    2017-01-01

    ABSTRACT Coxiella burnetii is an intracellular bacterial pathogen and a significant cause of culture-negative endocarditis in the United States. Upon infection, the nascent Coxiella phagosome fuses with the host endocytic pathway to form a large lysosome-like vacuole called the parasitophorous vacuole (PV). The PV membrane is rich in sterols, and drugs perturbing host cell cholesterol homeostasis inhibit PV formation and bacterial growth. Using cholesterol supplementation of a cholesterol-free cell model system, we found smaller PVs and reduced Coxiella growth as cellular cholesterol concentration increased. Further, we observed in cells with cholesterol a significant number of nonfusogenic PVs that contained degraded bacteria, a phenotype not observed in cholesterol-free cells. Cholesterol had no effect on axenic Coxiella cultures, indicating that only intracellular bacteria are sensitive to cholesterol. Live-cell microscopy revealed that both plasma membrane-derived cholesterol and the exogenous cholesterol carrier protein low-density lipoprotein (LDL) traffic to the PV. To test the possibility that increasing PV cholesterol levels affects bacterial survival, infected cells were treated with U18666A, a drug that traps cholesterol in lysosomes and PVs. U18666A treatment led to PVs containing degraded bacteria and a significant loss in bacterial viability. The PV pH was significantly more acidic in cells with cholesterol or cells treated with U18666A, and the vacuolar ATPase inhibitor bafilomycin blocked cholesterol-induced PV acidification and bacterial death. Additionally, treatment of infected HeLa cells with several FDA-approved cholesterol-altering drugs led to a loss of bacterial viability, a phenotype also rescued by bafilomycin. Collectively, these data suggest that increasing PV cholesterol further acidifies the PV, leading to Coxiella death. PMID:28246364

  4. Liposome-based Formulation for Intracellular Delivery of Functional Proteins

    Directory of Open Access Journals (Sweden)

    Benoît Chatin

    2015-01-01

    Full Text Available The intracellular delivery of biologically active protein represents an important emerging strategy for both fundamental and therapeutic applications. Here, we optimized in vitro delivery of two functional proteins, the β-galactosidase (β-gal enzyme and the anti-cytokeratin8 (K8 antibody, using liposome-based formulation. The guanidinium-cholesterol cationic lipid bis (guanidinium-tren-cholesterol (BGTC (bis (guanidinium-tren-cholesterol combined to the colipid dioleoyl phosphatidylethanolamine (DOPE (dioleoyl phosphatidylethanolamine was shown to efficiently deliver the β-gal intracellularly without compromising its activity. The lipid/protein molar ratio, protein amount, and culture medium were demonstrated to be key parameters affecting delivery efficiency. The protein itself is an essential factor requiring selection of the appropriate cationic lipid as illustrated by low K8 binding activity of the anti-K8 antibody using guanidinium-based liposome. Optimization of various lipids led to the identification of the aminoglycoside lipid dioleyl succinyl paromomycin (DOSP associated with the imidazole-based helper lipid MM27 as a potent delivery system for K8 antibody, achieving delivery in 67% of HeLa cells. Cryo-transmission electron microscopy showed that the structure of supramolecular assemblies BGTC:DOPE/β-gal and DOSP:MM27/K8 were different depending on liposome types and lipid/protein molar ratio. Finally, we observed that K8 treatment with DOSP:MM27/K8 rescues the cyclic adenosine monophosphate (cAMP-dependent chloride efflux in F508del-CFTR expressing cells, providing a new tool for the study of channelopathies.

  5. Information processing and integration with intracellular dynamics near critical point.

    Science.gov (United States)

    Kamimura, Atsushi; Kobayashi, Tetsuya J

    2012-01-01

    Recent experimental observations suggest that cells can show relatively precise and reliable responses to external signals even though substantial noise is inevitably involved in the signals. An intriguing question is the way how cells can manage to do it. One possible way to realize such response for a cell is to evolutionary develop and optimize its intracellular signaling pathways so as to extract relevant information from the noisy signal. We recently demonstrated that certain intracellular signaling reactions could actually conduct statistically optimal information processing. In this paper, we clarify that such optimal reaction operates near bifurcation point. This result suggests that critical-like phenomena in the single-cell level may be linked to efficient information processing inside a cell. In addition, improving the performance of response in the single-cell level is not the only way for cells to realize reliable response. Another possible strategy is to integrate information of individual cells by cell-to-cell interaction such as quorum sensing. Since cell-to-cell interaction is a common phenomenon, it is equally important to investigate how cells can integrate their information by cell-to-cell interaction to realize efficient information processing in the population level. In this paper, we consider roles and benefits of cell-to-cell interaction by considering integrations of obtained information of individuals with the other cells from the viewpoint of information processing. We also demonstrate that, by introducing cell movement, spatial organizations can spontaneously emerge as a result of efficient responses of the population to external signals.

  6. Superdiffusion dominates intracellular particle motion in the supercrowded space of pathogenic Acanthamoeba castellanii

    CERN Document Server

    Reverey, J F; Bao, H; Leippe, M; Metzler, R; Selhuber-Unkel, C

    2015-01-01

    Acanthamoebae are free-living protists and human pathogens, whose cellular functions and pathogenicity strongly depend on the transport of intracellular vesicles and granules through the cytosol. Using high-speed live cell imaging in combination with single-particle tracking analysis, we show here that the motion of endogenous intracellular particles in the size range from a few hundred nanometers to several micrometers in Acanthamoeba castellanii is strongly superdiffusive and influenced by cell locomotion, cytoskeletal elements, and myosin II. We demonstrate that cell locomotion significantly contributes to intracellular particle motion, but is clearly not the only origin of superdiffusivity. By analyzing the contribution of microtubules, actin, and myosin II motors we show that myosin II is a major driving force of intracellular motion in A. castellanii. The cytoplasm of A. castellanii is supercrowded with intracellular vesicles and granules, such that significant intracellular motion can only be achieved ...

  7. Superdiffusion dominates intracellular particle motion in the supercrowded cytoplasm of pathogenic Acanthamoeba castellanii

    Science.gov (United States)

    Reverey, Julia F.; Jeon, Jae-Hyung; Bao, Han; Leippe, Matthias; Metzler, Ralf; Selhuber-Unkel, Christine

    2015-06-01

    Acanthamoebae are free-living protists and human pathogens, whose cellular functions and pathogenicity strongly depend on the transport of intracellular vesicles and granules through the cytosol. Using high-speed live cell imaging in combination with single-particle tracking analysis, we show here that the motion of endogenous intracellular particles in the size range from a few hundred nanometers to several micrometers in Acanthamoeba castellanii is strongly superdiffusive and influenced by cell locomotion, cytoskeletal elements, and myosin II. We demonstrate that cell locomotion significantly contributes to intracellular particle motion, but is clearly not the only origin of superdiffusivity. By analyzing the contribution of microtubules, actin, and myosin II motors we show that myosin II is a major driving force of intracellular motion in A. castellanii. The cytoplasm of A. castellanii is supercrowded with intracellular vesicles and granules, such that significant intracellular motion can only be achieved by actively driven motion, while purely thermally driven diffusion is negligible.

  8. EFFECTS OF PDGF-BB ON INTRACELLULAR CALCIUM CONCENTRATION AND PROLIFERATION IN CULTURED GLOMERULAR MESANGIAL CELLS

    Institute of Scientific and Technical Information of China (English)

    WEN Li-ping; ZHANG Chong; BIAN Fan; ZOU Jun; JIANG Geng-ru; ZHU Han-wei

    2006-01-01

    Objective To investigate the relationship between the alteration of intracellular calcium concentration and proliferation in cultured glomerular mesangial cells. Methods Rat mesangial cells were cultured.Intracellular calcium concentrations were measured by confocal Laser Scanning Microscopy and Fura-3 fluorescence dyeing techniques. Cell growth was measured by MTT assay. Results PDGF-BB increased intracellular calcium concentrations in a dose-dependent manner, and at the same time promote the proliferation of mesangial cells. After preincubation with calcium channel blocker nifedipine or angiotensin converting enzyme inhibitor captopril, both the increase of intracellular calcium concentrations and cell proliferations induced by PDGF-BB were inhibited. Tripterigium Wilfordii Glycosides (TMG) significantly inhibited the mesangial cell proliferations, but it had no significant effect on intracellular calcium concentrations. Conclusion There was a positive relationship between the elevation of intracellular calcium concentration and cell proliferation in glomerular mesangial cells, but the increase of in- tracellular calcium concentrations wasn't the only way for proliferation.

  9. Gold nanodome-patterned microchips for intracellular surface-enhanced Raman spectroscopy.

    Science.gov (United States)

    Wuytens, Pieter C; Subramanian, Ananth Z; De Vos, Winnok H; Skirtach, Andre G; Baets, Roel

    2015-12-21

    While top-down substrates for surface-enhanced Raman spectroscopy (SERS) offer outstanding control and reproducibility of the gold nanopatterns and their related localized surface plasmon resonance, intracellular SERS experiments heavily rely on gold nanoparticles. These nanoparticles often result in varying and uncontrollable enhancement factors. Here we demonstrate the use of top-down gold-nanostructured microchips for intracellular sensing. We develop a tunable and reproducible fabrication scheme for these microchips. Furthermore we observe the intracellular uptake of these structures, and find no immediate influence on cell viability. Finally, we perform a proof-of-concept intracellular SERS experiment by the label-free detection of extraneous molecules. By bringing top-down SERS substrates to the intracellular world, we set an important step towards time-dependent and quantitative intracellular SERS.

  10. Extra- and intracellular volume monitoring by impedance during haemodialysis using Cole-Cole extrapolation.

    Science.gov (United States)

    Jaffrin, M Y; Maasrani, M; Le Gourrier, A; Boudailliez, B

    1997-05-01

    A method is presented for monitoring the relative variation of extracellular and intracellular fluid volumes using a multifrequency impedance meter and the Cole-Cole extrapolation technique. It is found that this extrapolation is necessary to obtain reliable data for the resistance of the intracellular fluid. The extracellular and intracellular resistances can be approached using frequencies of, respectively, 5 kHz and 1000 kHz, but the use of 100 kHz leads to unacceptable errors. In the conventional treatment the overall relative variation of intracellular resistance is found to be relatively small.

  11. Influence of intracellular acidosis on contractile function in the working rat heart

    Energy Technology Data Exchange (ETDEWEB)

    Jeffrey, F.M.H.; Malloy, C.R.; Radda, G.K. (Univ. of Oxford (England))

    1987-12-01

    The decrease in myocardial contractility during ischemia, hypoxia, and extracellular acidosis has been attributed to intracellular acidosis. Previous studies of the relationship between pH and contractile state have utilized respiratory or metabolic acidosis to alter intracellular pH. The authors developed a model in the working perfused rat heart to study the effects of intracellular acidosis with normal external pH and optimal O{sub 2} delivery. Intracellular pH and high-energy phosphates were monitored by {sup 31}P nuclear magnetic resonance spectroscopy. Hearts were perfused to a steady state with a medium containing 10 mM NH{sub 4}Cl. Acidosis induced a substantial decrease in aortic flow and stroke volume which was associated with little change in peak systolic pressure. It was concluded that (1) for the same intracellular acidosis the influence on tension development was more pronounced with a combined extra- and intracellular acidosis than with an isolated intracellular acidosis, and (2) stroke volume at constant preload was impaired by intracellular acidosis even though changes in developed pressure were minimal. These observations suggest that isolated intracellular acidosis has adverse effects on diastolic compliance and/or relaxation.

  12. Neutrophilic granulocytes reactive response in candida vulvovaginitis patients with intracellular microorganism persistence complications

    OpenAIRE

    YAKOVYCHUK NINA DMYTRIVNA; DJUIRIAK VALENTYNA STEPANIVNA

    2015-01-01

    Polymorphic neutrophilic granulocytes reactive response and body immune reactivity in general considerably decrease in patients suffering from candida vaginitis on the basis of intracellular microorganisms persistence.

  13. Increased intracellular free calcium and sensitivity to angiotensin II in platelets of preeclamptic women.

    Science.gov (United States)

    Haller, H; Oeney, T; Hauck, U; Distler, A; Philipp, T

    1989-04-01

    Preeclampsia is characterized by a generalized vasoconstriction and increased vascular sensitivity to angiotensin II. Intracellular free calcium, implicated in vascular smooth muscle contraction, has been found to be elevated in platelets of other hypertensive disorders. We therefore measured intracellular free calcium concentrations by using the fluorescent probe quin-2 in platelets of six patients with preeclampsia and compared them to measurements in ten normotensive pregnant women and ten age-matched nonpregnant women. Intracellular free calcium was also determined in the preeclamptic women after delivery. We found that intracellular free calcium was slightly elevated in normal pregnancy (102 +/- 13 nmol/L v 87 +/- 17 nmol/L) but was markedly increased in preeclampsia (138 +/- 13 nmol/L, P less than .05). This increase disappeared six weeks after delivery (84 + 10 nmol/L, P less than .01). To investigate whether the increased intracellular free calcium was related to angiotensin II, the platelets were exposed to thrombin and angiotensin II in vitro. Exposure to thrombin and angiotensin II caused a dose-dependent increase in intracellular free calcium. The intracellular response to thrombin was not significantly different in the three groups. However, stimulation with angiotensin II revealed an increased response in intracellular free calcium in preeclampsia (P less than .05) that disappeared after delivery. Our findings show a sustained increase in platelet intracellular free calcium in preeclampsia and suggest a functional alteration of the angiotensin II receptor in this disease.

  14. 含盐条件下偶氮染料兼厌氧性生物的降解性能%Facultative anaerobic biodegradation performance of azo dyestuff under salt-containing condition

    Institute of Scientific and Technical Information of China (English)

    雷弢; 仝攀瑞; 陈方方; 王晓艳

    2011-01-01

    The facultstive anaerobic biodegradation performance of reactive dyestuffs is studied by using different concentrations glucose. And based on this conclusion, the facultative anaerobic biodegradation performance of K-2BP under different salt-containlng condition is investigated by static test reactor. The results indicate that facultative anaerobic microorganisms has lower degradation rate when only K-2BP is used as the substrate, and the degradation rate would be increased under the existence of the glucose. The glucose concentration is 800 mg/L, while the degradation rate of K-2BP is 64. 1% in 6h. However, it is detrimental to dye's biodegradation when the glucose concentration is up to 1 000 mg/L, the degradation rate is only about 46%, that is similar to the condition of non-glucose. On the premise of glucose concentration 800 mg/L, when the salt concentrations are 2 g/L,5 g/L,l0 g/L and 20 g/L respectively,the first-order kinetics constants are 0. 10578 mg/L· h,0. 04947mg/L · h,0.028 69mg/L · h and 0.022 75mg/L · h respectively. And the half-lives are 6.99h,14. 15h,22.55h and 30.21h respectively. The reaction rate constants is daereased gradually with the salt concentrations'incresse,which is showed that high salt concentration can restrainthe facultative anaerobic microorganisms' degradation of K-2BP.%研究活性染料与不同浓度葡萄糖共基质条件下的兼厌氧性生物降解性能和K-2BP在不同盐浓度条件下的兼厌氧性生物降解性能.选择K-2BP作为目标污染物进行静态反应器生物降解实验.结果表明,兼厌氧性微生物在只有K-2BP作为基质时对染料的降解率较低,葡萄糖存在时,能提高兼厌氧性生物对染料的降解能力;葡萄糖为800mg/L时6h染料降解率为64.1%,而葡萄糖浓度为1 000mg/L时,不利于染料降解,6h染料降解率为46%,与不投加葡萄糖情况的降解率接近.葡萄糖浓度为800mg/L,盐浓度分别为2g/L,5g/L,10g/L和20g/L,其一级

  15. Study on intracellular trafficking of Mac-1 by direct visualization

    Institute of Scientific and Technical Information of China (English)

    YAN Ming; MAO Jifang; WEI Yi; ZHONG Jigen; YANG Shengsheng; XU Renbao

    2004-01-01

    Previously, we constructed DNA vectors containing cDNA of Mac-1 subunits (CD11b or CD18b) fused with fluorescence protein (FP). cDNA fragments and the DNA constructs were then transfected into CHO cells (as CHO-Mac-1-FP). The structure and function of Mac-1-FP obtained from the CHO-Mac-1-FP cells are nearly identical to that expressed in wild type leukocytes. In the present study, the intracellular trafficking of Mac-1 was visualized directly by monitoring the fluorescent intensities of YFP-CD18 and PE-conjugated monoclonal antibody against CD11b under a confocal microscope in CHO-Mac-1-FP cells. The results indicate that: (ⅰ) although Mac-1 was not detected in the cell membrane at resting state, it had been translocated and clustered into the cell membrane by 1 h and internalized 2 h after PMA stimulation, at which point the fluorescence intensity began to diminish gradually, probably due to partial degradation of Mac-1. The fluorescence of CD18 and CD11b reappeared on the cell membrane 1 h after re-treatment with PMA, suggesting the recycling of non-degraded Mac-1. (ⅱ) The adhesion rate of CHO-Mac-1-FP to magnetic beads coupled ICAM-1 increased within 4 h after their initial interaction, accompanied by the clustering of Mac-1-FP. After 8 h,the adhesion rate declined and fluorescence also decreased simultaneously. The pattern of change in fluorescence in CHO-Mac-1-FP cells elicited by ICAM-1 beads was similar to that elicited by PMA, suggesting that endocytosis and degradation of Mac-1 occurred after the interaction with ICAM-1. Thus, we conclude that the intracellular trafficking of Mac-1 after activation is associated with membrane translocation, endocytosis, degradation and recycling. These changes are in parallel with the adhesion of CHO-Mac-1-FP cells with ICAM-1, and may be involved in the adhesion and detachment of leukocytes. The detachment of leukocytes may be caused by endocytosis of Mac-1.

  16. Psychrotrophic Clostridia Causing Spoilage in Cooked Meat and Poultry Products

    National Research Council Canada - National Science Library

    Kalinowski, Robin M; Tompkin, R. Bruce

    1999-01-01

    ...°C or below, and typically given use by dates of greater than 50 days. While of rare, sporadic occurrence, an unpleasant spoilage characterized by strong H2S odor and gas production has been observed in these products...

  17. Resveratrol Interferes with Fura-2 Intracellular Calcium Measurements.

    Science.gov (United States)

    Kopp, Richard F; Leech, Colin A; Roe, Michael W

    2014-03-01

    Resveratrol, a naturally occurring polyphenol found in some fruits and especially in grapes, has been reported to provide diverse health benefits. Resveratrol's mechanism of action is the subject of many investigations, and some studies using the ratiometric calcium indicator Fura-2 suggest that it modulates cellular calcium responses. In the current study, contradictory cellular calcium responses to resveratrol applied at concentrations exceeding 10 μM were observed during in vitro imaging studies depending on the calcium indicator used, with Fura-2 indicating an increase in intracellular calcium while Fluo-4 and the calcium biosensor YC3.60 indicated no response. When cells loaded with Fura-2 were treated with 100 μM resveratrol, excitation at 340 nm resulted in a large intensity increase at 510 nm, but the expected concurrent decline with 380 nm excitation was not observed. Pre-treatment of cells with the calcium chelator BAPTA-AM did not prevent a rise in the 340/380 ratio when resveratrol was present, but it did prevent an increase in 340/380 when ATP was applied, suggesting that the resveratrol response was an artifact. Cautious data interpretation is recommended from imaging experiments using Fura-2 concurrently with resveratrol in calcium imaging experiments.

  18. The mechanical environment modulates intracellular calcium oscillation activities of myofibroblasts.

    Directory of Open Access Journals (Sweden)

    Charles Godbout

    Full Text Available Myofibroblast contraction is fundamental in the excessive tissue remodeling that is characteristic of fibrotic tissue contractures. Tissue remodeling during development of fibrosis leads to gradually increasing stiffness of the extracellular matrix. We propose that this increased stiffness positively feeds back on the contractile activities of myofibroblasts. We have previously shown that cycles of contraction directly correlate with periodic intracellular calcium oscillations in cultured myofibroblasts. We analyze cytosolic calcium dynamics using fluorescent calcium indicators to evaluate the possible impact of mechanical stress on myofibroblast contractile activity. To modulate extracellular mechanics, we seeded primary rat subcutaneous myofibroblasts on silicone substrates and into collagen gels of different elastic modulus. We modulated cell stress by cell growth on differently adhesive culture substrates, by restricting cell spreading area on micro-printed adhesive islands, and depolymerizing actin with Cytochalasin D. In general, calcium oscillation frequencies in myofibroblasts increased with increasing mechanical challenge. These results provide new insight on how changing mechanical conditions for myofibroblasts are encoded in calcium oscillations and possibly explain how reparative cells adapt their contractile behavior to the stresses occurring in normal and pathological tissue repair.

  19. Intracellular micro-rheology probed by micron-sized wires.

    Science.gov (United States)

    Chevry, Loudjy; Colin, Rémy; Abou, Bérengère; Berret, Jean-François

    2013-09-01

    In the last decade, rapid advances have been made in the field of micro-rheology of cells and tissues. Given the complexity of living systems, there is a need for the development of new types of nano- and micron-sized probes, and in particular of probes with controlled interactions with the surrounding medium. In the present paper, we evaluate the use of micron-sized wires as potential probes of the mechanical properties of cells. The wire-based micro-rheology technique is applied to living cells such as murine fibroblasts and canine kidney epithelial cells. The mean-squared angular displacement of wires associated to their rotational dynamics is obtained as a function of the time using optical microscopy and image processing. It reveals a Brownian-like diffusive regime of the form Δψ(2)(t,L) ≈ t/L(3), where L denotes the wire length. This scaling suggests that an effective viscosity of the intracellular medium can be determined, and that in the range 1-10 μm it does not depend on the length scale over which it is measured. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. Clustered Intracellular Salmonella enterica Serovar Typhimurium Blocks Host Cell Cytokinesis.

    Science.gov (United States)

    Santos, António J M; Durkin, Charlotte H; Helaine, Sophie; Boucrot, Emmanuel; Holden, David W

    2016-07-01

    Several bacterial pathogens and viruses interfere with the cell cycle of their host cells to enhance virulence. This is especially apparent in bacteria that colonize the gut epithelium, where inhibition of the cell cycle of infected cells enhances the intestinal colonization. We found that intracellular Salmonella enterica serovar Typhimurium induced the binucleation of a large proportion of epithelial cells by 14 h postinvasion and that the effect was dependent on an intact Salmonella pathogenicity island 2 (SPI-2) type 3 secretion system. The SPI-2 effectors SseF and SseG were required to induce binucleation. SseF and SseG are known to maintain microcolonies of Salmonella-containing vacuoles close to the microtubule organizing center of infected epithelial cells. During host cell division, these clustered microcolonies prevented the correct localization of members of the chromosomal passenger complex and mitotic kinesin-like protein 1 and consequently prevented cytokinesis. Tetraploidy, arising from a cytokinesis defect, is known to have a deleterious effect on subsequent cell divisions, resulting in either chromosomal instabilities or cell cycle arrest. In infected mice, proliferation of small intestinal epithelial cells was compromised in an SseF/SseG-dependent manner, suggesting that cytokinesis failure caused by S Typhimurium delays epithelial cell turnover in the intestine.

  1. Polyarginine nanocapsules: a new platform for intracellular drug delivery

    Energy Technology Data Exchange (ETDEWEB)

    Lozano, M. V.; Lollo, G. [University of Santiago de Compostela, Department of Pharmaceutical Technology, Faculty of Pharmacy (Spain); Alonso-Nocelo, M. [Complexo Hospitalario, Universitario de Santiago de Compostela/SERGAS, Translational Laboratory, Medical Oncology Department (Spain); Brea, J. [University of Santiago de Compostela, Department of Pharmacology, Faculty of Pharmacy (Spain); Vidal, A. [University of Santiago de Compostela, Department of Physiology (Spain); Torres, D.; Alonso, M. J., E-mail: mariaj.alonso@usc.es [University of Santiago de Compostela, Department of Pharmaceutical Technology, Faculty of Pharmacy (Spain)

    2013-03-15

    This report describes the development of a new nanocarrier, named as polyarginine (PArg) nanocapsules, specifically designed for overcoming cellular barriers. These nanocapsules are composed of an oily core and a PArg corona. The attachment of the PArg corona was mediated by its interaction with the oily core, which was conveniently stabilized with phosphatidylcholine. Hybrid PArg/PEG nanocapsules could also be obtained by introducing PEG-stearate in the nanocapsules formation process. The nanocapsules had an average size in the range of 120-160 nm, and a positive surface charge, which varied between +56 and +28 mV for PArg and PArg/PEG nanocapsules, respectively. They could accommodate significant amounts of lipophilic drugs, i.e., docetaxel, in their core, and also polar negatively charged molecules, i.e., plasmid DNA, on their coating. As a preliminary proof-of-principle, we explored the ability of these nanocarriers to enter cancer cells and to inhibit proliferation in the non-small cell lung cancer NCI-H460 cell line, using flow cytometry and confocal microscopy analysis. The results indicated that PArg nanocapsules are rapidly and massively accumulated into the NCI-H460 cells and that the PArg shell plays a critical role in the internalization process. Moreover, the incubation with docetaxel-loaded nanocapsules with NCI-H460 cells led to an enhanced inhibition of their proliferation, as compared to the free drug. Overall, this is the first report of the potential of PArg nanocapsules as intracellular drug delivery vehicles.

  2. Initial intracellular proteome profile of Aspergillus niger biofilms

    Directory of Open Access Journals (Sweden)

    Gretty K. Villena

    2011-07-01

    Full Text Available An initial profiling of the intracellular proteome of Aspergillus niger ATCC 10864 biofilm cultures developed on polyester cloth was carried out by using 2D-PAGE and MS-TOF analysis and it was compared to the proteome of conventionally grown free-living submerged cultures. A number of 2D-PAGE protein spots from both types of cultures were subjected to MS-TOF analysis and data interrogation of the NCBI nr database available for this species. Proteomic maps showed different expression patterns in both culture systems with differentially expressed proteins in each case. In biofilm cultures, 19% and 32% of the selected protein spots were over- expressed and differentially expressed, respectively. On the contrary, in free-living cultures, 44% and 7% of the selected protein spots were over-expressed and differentially expressed, respectively. Although preliminary, results presented in this paper show that there are significant differences between the proteomes of A. niger biofilm and free-living mycelia. It seems that cell adhesion is the most important stimulus responsible for biofilm development which is the basis of Surface Adhesion Fermentation.

  3. Iron in intracellular infection: to provide or to deprive?

    Directory of Open Access Journals (Sweden)

    Sandro eSilva-Gomes

    2013-12-01

    Full Text Available Due to their chemical versatility, transition metals were incorporated as cofactors for several basic metabolic pathways in living organisms. This same characteristic makes them potentially harmful, since they can be engaged in deleterious reactions like Fenton chemistry. As such, organisms have evolved highly specialized mechanisms to supply their own metal needs while keeping their toxic potential in check.This dual character comes into play in host-pathogen interactions, given that the host can either deprive the pathogen of these key nutrients or exploit them to induce toxicity towards the invading agent. Iron stands as the prototypic example of how a metal can be used to limit the growth of pathogens by nutrient deprivation, a mechanism widely studied in Mycobacterium infections. However, the host can also take advantage of iron-induced toxicity to control pathogen proliferation, as observed in infections caused by Leishmania. Whether we may harness either of the two pathways for therapeutical purposes is still ill-defined.In this review, we discuss how modulation of the host iron availability impacts the course of infections, focusing on those caused by two relevant intracellular pathogens, Mycobacterium and Leishmania.

  4. Hydrodynamic regulation of monocyte inflammatory response to an intracellular pathogen.

    Directory of Open Access Journals (Sweden)

    Shankar J Evani

    Full Text Available Systemic bacterial infections elicit inflammatory response that promotes acute or chronic complications such as sepsis, arthritis or atherosclerosis. Of interest, cells in circulation experience hydrodynamic shear forces, which have been shown to be a potent regulator of cellular function in the vasculature and play an important role in maintaining tissue homeostasis. In this study, we have examined the effect of shear forces due to blood flow in modulating the inflammatory response of cells to infection. Using an in vitro model, we analyzed the effects of physiological levels of shear stress on the inflammatory response of monocytes infected with chlamydia, an intracellular pathogen which causes bronchitis and is implicated in the development of atherosclerosis. We found that chlamydial infection alters the morphology of monocytes and trigger the release of pro-inflammatory cytokines TNF-α, IL-8, IL-1β and IL-6. We also found that the exposure of chlamydia-infected monocytes to short durations of arterial shear stress significantly enhances the secretion of cytokines in a time-dependent manner and the expression of surface adhesion molecule ICAM-1. As a functional consequence, infection and shear stress increased monocyte adhesion to endothelial cells under flow and in the activation and aggregation of platelets. Overall, our study demonstrates that shear stress enhances the inflammatory response of monocytes to infection, suggesting that mechanical forces may contribute to disease pathophysiology. These results provide a novel perspective on our understanding of systemic infection and inflammation.

  5. Immunity to intracellular Salmonella depends on surface-associated antigens.

    Directory of Open Access Journals (Sweden)

    Somedutta Barat

    Full Text Available Invasive Salmonella infection is an important health problem that is worsening because of rising antimicrobial resistance and changing Salmonella serovar spectrum. Novel vaccines with broad serovar coverage are needed, but suitable protective antigens remain largely unknown. Here, we tested 37 broadly conserved Salmonella antigens in a mouse typhoid fever model, and identified antigen candidates that conferred partial protection against lethal disease. Antigen properties such as high in vivo abundance or immunodominance in convalescent individuals were not required for protectivity, but all promising antigen candidates were associated with the Salmonella surface. Surprisingly, this was not due to superior immunogenicity of surface antigens compared to internal antigens as had been suggested by previous studies and novel findings for CD4 T cell responses to model antigens. Confocal microscopy of infected tissues revealed that many live Salmonella resided alone in infected host macrophages with no damaged Salmonella releasing internal antigens in their vicinity. In the absence of accessible internal antigens, detection of these infected cells might require CD4 T cell recognition of Salmonella surface-associated antigens that could be processed and presented even from intact Salmonella. In conclusion, our findings might pave the way for development of an efficacious Salmonella vaccine with broad serovar coverage, and suggest a similar crucial role of surface antigens for immunity to both extracellular and intracellular pathogens.

  6. Real-Time monitoring of intracellular wax ester metabolism

    Directory of Open Access Journals (Sweden)

    Karp Matti

    2011-09-01

    Full Text Available Abstract Background Wax esters are industrially relevant molecules exploited in several applications of oleochemistry and food industry. At the moment, the production processes mostly rely on chemical synthesis from rather expensive starting materials, and therefore solutions are sought from biotechnology. Bacterial wax esters are attractive alternatives, and especially the wax ester metabolism of Acinetobacter sp. has been extensively studied. However, the lack of suitable tools for rapid and simple monitoring of wax ester metabolism in vivo has partly restricted the screening and analyses of potential hosts and optimal conditions. Results Based on sensitive and specific detection of intracellular long-chain aldehydes, specific intermediates of wax ester synthesis, bacterial luciferase (LuxAB was exploited in studying the wax ester metabolism in Acinetobacter baylyi ADP1. Luminescence was detected in the cultivation of the strain producing wax esters, and the changes in signal levels could be linked to corresponding cell growth and wax ester synthesis phases. Conclusions The monitoring system showed correlation between wax ester synthesis pattern and luminescent signal. The system shows potential for real-time screening purposes and studies on bacterial wax esters, revealing new aspects to dynamics and role of wax ester metabolism in bacteria.

  7. Nondestructive nanostraw intracellular sampling for longitudinal cell monitoring

    Science.gov (United States)

    Cao, Yuhong; Chen, Haodong; Birey, Fikri; Leal-Ortiz, Sergio A.; Han, Crystal M.; Santiago, Juan G.; Paşca, Sergiu P.; Wu, Joseph C.; Melosh, Nicholas A.

    2017-01-01

    Here, we report a method for time-resolved, longitudinal extraction and quantitative measurement of intracellular proteins and mRNA from a variety of cell types. Cytosolic contents were repeatedly sampled from the same cell or population of cells for more than 5 d through a cell-culture substrate, incorporating hollow 150-nm-diameter nanostraws (NS) within a defined sampling region. Once extracted, the cellular contents were analyzed with conventional methods, including fluorescence, enzymatic assays (ELISA), and quantitative real-time PCR. This process was nondestructive with >95% cell viability after sampling, enabling long-term analysis. It is important to note that the measured quantities from the cell extract were found to constitute a statistically significant representation of the actual contents within the cells. Of 48 mRNA sequences analyzed from a population of cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs), 41 were accurately quantified. The NS platform samples from a select subpopulation of cells within a larger culture, allowing native cell-to-cell contact and communication even during vigorous activity such as cardiomyocyte beating. This platform was applied both to cell lines and to primary cells, including CHO cells, hiPSC-CMs, and human astrocytes derived in 3D cortical spheroids. By tracking the same cell or group of cells over time, this method offers an avenue to understand dynamic cell behavior, including processes such as induced pluripotency and differentiation. PMID:28223521

  8. Sigma-1 receptor: The novel intracellular target of neuropsychotherapeutic drugs

    Directory of Open Access Journals (Sweden)

    Teruo Hayashi

    2015-01-01

    Full Text Available Sigma-1 receptor ligands have been long expected to serve as drugs for treatment of human diseases such as neurodegenerative disorders, depression, idiopathic pain, drug abuse, and cancer. Recent research exploring the molecular function of the sigma-1 receptor started unveiling underlying mechanisms of the therapeutic activity of those ligands. Via the molecular chaperone activity, the sigma-1 receptor regulates protein folding/degradation, ER/oxidative stress, and cell survival. The chaperone activity is activated or inhibited by synthetic sigma-1 receptor ligands in an agonist-antagonist manner. Sigma-1 receptors are localized at the endoplasmic reticulum (ER membranes that are physically associated with the mitochondria (MAM: mitochondria-associated ER membrane. In specific types of neurons (e.g., those at the spinal cord, sigma-1 receptors are also clustered at ER membranes that juxtapose postsynaptic plasma membranes. Recent studies indicate that sigma-1 receptors, partly in sake of its unique subcellular localization, regulate the mitochondria function that involves bioenergetics and free radical generation. The sigma-1 receptor may thus provide an intracellular drug target that enables controlling ER stress and free radical generation under pathological conditions.

  9. Repurposing bacterial toxins for intracellular delivery of therapeutic proteins.

    Science.gov (United States)

    Beilhartz, Greg L; Sugiman-Marangos, Seiji N; Melnyk, Roman A

    2017-10-15

    Despite enormous efforts, achieving efficacious levels of proteins inside mammalian cells remains one of the greatest challenges in biologics-based drug discovery and development. The inability of proteins to readily cross biological membranes precludes access to the wealth of intracellular targets and applications that lie within mammalian cells. Existing methods of delivery commonly suffer from an inability to target specific cells and tissues, poor endosomal escape, and limited in vivo efficacy. The aim of the present commentary is to highlight the potential of certain classes of bacterial toxins, which naturally deliver a large protein into the cytosolic compartment of target cells after binding a host cell-surface receptor with high affinity, as robust protein delivery platforms. We review the progress made in recent years toward demonstrating the utility of these systems at delivering a wide variety of protein cargo, with special attention paid to three distinct toxin-based platforms. We contend that with recent advances in protein deimmunization strategies, bacterial toxins are poised to introduce biologics into the inner sanctum of cells and treat a wealth of heretofore untreatable diseases with a new generation of therapeutics. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Intracellular magnetophoresis of amyloplasts and induction of root curvature

    Science.gov (United States)

    Kuznetsov, O. A.; Hasenstein, K. H.

    1996-01-01

    High-gradient magnetic fields (HGMFs) were used to induce intracellular magnetophoresis of amyloplasts. The HGMFs were generated by placing a small ferromagnetic wedge into a uniform magnetic field or at the gap edge between two permanent magnets. In the vicinity of the tip of the wedge the dynamic factor of the magnetic field, delta(H2/2), was about 10(9) Oe2.cm-1, which subjected the amyloplasts to a force comparable to that of gravity. When roots of 2-d-old seedlings of flax (Linum usitatissimum L.) were positioned vertically and exposed to an HGMF, curvature away from the wedge was transient and lasted approximately 1 h. Average curvature obtained after placing magnets, wedge and seedlings on a 1-rpm clinostat for 2 h was 33 +/- 5 degrees. Roots of horizontally placed control seedlings without rotation curved about 47 +/- 4 degrees. The time course of curvature and changes in growth rate were similar for gravicurvature and for root curvature induced by HGMFs. Microscopy showed displacement of amyloplasts in vitro and in vivo. Studies with Arabidopsis thaliana (L.) Heynh. showed that the wild type responded to HGMFs but the starchless mutant TC7 did not. The data indicate that a magnetic force can be used to study the gravisensing and response system of roots.

  11. Targeting of a chlamydial protease impedes intracellular bacterial growth.

    Directory of Open Access Journals (Sweden)

    Jan G Christian

    2011-09-01

    Full Text Available Chlamydiae are obligate intracellular bacteria that propagate in a cytosolic vacuole. Recent work has shown that growth of Chlamydia induces the fragmentation of the Golgi apparatus (GA into ministacks, which facilitates the acquisition of host lipids into the growing inclusion. GA fragmentation results from infection-associated cleavage of the integral GA protein, golgin-84. Golgin-84-cleavage, GA fragmentation and growth of Chlamydia trachomatis can be blocked by the peptide inhibitor WEHD-fmk. Here we identify the bacterial protease chlamydial protease-like activity factor (CPAF as the factor mediating cleavage of golgin-84 and as the target of WEHD-fmk-inhibition. WEHD-fmk blocked cleavage of golgin-84 as well as cleavage of known CPAF targets during infection with C. trachomatis and C. pneumoniae. The same effect was seen when active CPAF was expressed in non-infected cells and in a cell-free system. Ectopic expression of active CPAF in non-infected cells was sufficient for GA fragmentation. GA fragmentation required the small GTPases Rab6 and Rab11 downstream of CPAF-activity. These results define CPAF as the first protein that is essential for replication of Chlamydia. We suggest that this role makes CPAF a potential anti-infective therapeutic target.

  12. Regulation of intracellular heme trafficking revealed by subcellular reporters.

    Science.gov (United States)

    Yuan, Xiaojing; Rietzschel, Nicole; Kwon, Hanna; Walter Nuno, Ana Beatriz; Hanna, David A; Phillips, John D; Raven, Emma L; Reddi, Amit R; Hamza, Iqbal

    2016-08-30

    Heme is an essential prosthetic group in proteins that reside in virtually every subcellular compartment performing diverse biological functions. Irrespective of whether heme is synthesized in the mitochondria or imported from the environment, this hydrophobic and potentially toxic metalloporphyrin has to be trafficked across membrane barriers, a concept heretofore poorly understood. Here we show, using subcellular-targeted, genetically encoded hemoprotein peroxidase reporters, that both extracellular and endogenous heme contribute to cellular labile heme and that extracellular heme can be transported and used in toto by hemoproteins in all six subcellular compartments examined. The reporters are robust, show large signal-to-background ratio, and provide sufficient range to detect changes in intracellular labile heme. Restoration of reporter activity by heme is organelle-specific, with the Golgi and endoplasmic reticulum being important sites for both exogenous and endogenous heme trafficking. Expression of peroxidase reporters in Caenorhabditis elegans shows that environmental heme influences labile heme in a tissue-dependent manner; reporter activity in the intestine shows a linear increase compared with muscle or hypodermis, with the lowest heme threshold in neurons. Our results demonstrate that the trafficking pathways for exogenous and endogenous heme are distinct, with intrinsic preference for specific subcellular compartments. We anticipate our results will serve as a heuristic paradigm for more sophisticated studies on heme trafficking in cellular and whole-animal models.

  13. Ornithine decarboxylase antizyme inhibitor 2 regulates intracellular vesicle trafficking

    Energy Technology Data Exchange (ETDEWEB)

    Kanerva, Kristiina; Maekitie, Laura T. [Department of Pathology, Haartman Institute, University of Helsinki, Helsinki (Finland); Baeck, Nils [Department of Anatomy, Institute of Biomedicine, University of Helsinki, Helsinki (Finland); Andersson, Leif C., E-mail: leif.andersson@helsinki.fi [Department of Pathology, Haartman Institute, University of Helsinki, Helsinki (Finland); HUSLAB, Helsinki (Finland); Department of Oncology and Pathology, Karolinska Institutet, Stockholm (Sweden)

    2010-07-01

    Antizyme inhibitor 1 (AZIN1) and 2 (AZIN2) are proteins that activate ornithine decarboxylase (ODC), the key enzyme of polyamine biosynthesis. Both AZINs release ODC from its inactive complex with antizyme (AZ), leading to formation of the catalytically active ODC. The ubiquitously expressed AZIN1 is involved in cell proliferation and transformation whereas the role of the recently found AZIN2 in cellular functions is unknown. Here we report the intracellular localization of AZIN2 and present novel evidence indicating that it acts as a regulator of vesicle trafficking. We used immunostaining to demonstrate that both endogenous and FLAG-tagged AZIN2 localize to post-Golgi vesicles of the secretory pathway. Immuno-electron microscopy revealed that the vesicles associate mainly with the trans-Golgi network (TGN). RNAi-mediated knockdown of AZIN2 or depletion of cellular polyamines caused selective fragmentation of the TGN and retarded the exocytotic release of vesicular stomatitis virus glycoprotein. Exogenous addition of polyamines normalized the morphological changes and reversed the inhibition of protein secretion. Our findings demonstrate that AZIN2 regulates the transport of secretory vesicles by locally activating ODC and polyamine biosynthesis.

  14. [Carboxyl nanodiamond as intracellular transporters of anticancer drug--podophyllotoxin].

    Science.gov (United States)

    Sun, Tao-Li; Wang, Bin; Peng, Yan; Ni, Jing-Man

    2013-01-01

    The purpose of this study is to investigate the intracellular transporters effect and the cytotoxicity of carboxyl nanodiamond (CND) - podophyllotoxin (PPT). Nanodiamond (ND) was treated with mixed carboxylic acid and finally got 64 nm CND by centrifugation, and then it was reacted with PPT to form CND-PPT. UV spectrophotometry was used to calculate the content of PPT in CND-PPT, the particle size distribution and zeta potential were measured by Dynamic laser scattering instrument. CND, PPT, CND-PPT and CND + PPT (physical mixture of CND and PPT) were characterized by Fourier transform infrared spectroscopy, at the same time, thermal analysis and element analysis were used to estimate the content of the PPT in CND-PPT. The affect of CND, PPT, CND-PPT on HeLa cell was measured with MTT assay. The results showed that content of PPT combined with CND accounted for about 10%. MTT assay showed that CND has low cytotoxicity and CND-PPT can increase the water soluble of PPT. As a conclusion, CND as a hydrophilic pharmaceutical carrier combined with PPT is able to increase the water solubility of PPT, at low concentration, CND-PPT can enhance the antitumor activity in comparison with PPT, so CND can be used as a potential anticancer drug carrier.

  15. Global intracellular slow-wave dynamics of the thalamocortical system.

    Science.gov (United States)

    Sheroziya, Maxim; Timofeev, Igor

    2014-06-25

    It is widely accepted that corticothalamic neurons recruit the thalamus in slow oscillation, but global slow-wave thalamocortical dynamics have never been experimentally shown. We analyzed intracellular activities of neurons either from different cortical areas or from a variety of specific and nonspecific thalamic nuclei in relation to the phase of global EEG signal in ketamine-xylazine anesthetized mice. We found that, on average, slow-wave active states started off within frontal cortical areas as well as higher-order and intralaminar thalamus (posterior and parafascicular nuclei) simultaneously. Then, the leading edge of active states propagated in the anteroposterior/lateral direction over the cortex at ∼40 mm/s. The latest structure we recorded within the slow-wave cycle was the anterior thalamus, which followed active states of the retrosplenial cortex. Active states from different cortical areas tended to terminate simultaneously. Sensory thalamic ventral posterior medial and lateral geniculate nuclei followed cortical active states with major inhibitory and weak tonic-like "modulator" EPSPs. In these nuclei, sharp-rising, large-amplitude EPSPs ("drivers") were not modulated by cortical slow waves, suggesting their origin in ascending pathways. The thalamic active states in other investigated nuclei were composed of depolarization: some revealing "driver"- and "modulator"-like EPSPs, others showing "modulator"-like EPSPs only. We conclude that sensory thalamic nuclei follow the propagating cortical waves, whereas neurons from higher-order thalamic nuclei display "hub dynamics" and thus may contribute to the generation of cortical slow waves.

  16. Enlightening intracellular complexity of living cells with quantitative phase microscopy

    Science.gov (United States)

    Martinez Torres, C.; Laperrousaz, B.; Berguiga, L.; Boyer Provera, E.; Elezgaray, J.; Nicolini, F. E.; Maguer-Satta, V.; Arneodo, A.; Argoul, F.

    2016-03-01

    The internal distribution of refractive indices (RIs) of a living cell is much more complex than usually admitted in multi-shell models. The reconstruction of RI maps from single phase images has rarely been achieved for several reasons: (i) we still have very little knowledge of the impact of internal macromolecular complexes on the local RI and (ii) phase changes produced by light propagation through the sample are mixed with diffraction effects by internal cell bodies. We propose the implementation a 2D wavelet-based contour chain detection method to distinguish internal boundaries thanks to their greatest optical path difference gradients. These contour chains correspond to the highest image phase contrast and follow the local RI inhomogeneities linked to the intracellular structural intricacy. Their statistics and spatial distribution are morphological indicators for distinguishing cells of different origins and to follow their transformation in pathologic situations. We use this method to compare non adherent blood cells from primary and laboratory culture origins, in healthy and pathological situations (chronic myelogenous leukaemia). In a second part of this presentation, we concentrate on the temporal dynamics of the phase contour chains and we discuss the spectral decomposition of their dynamics in both health and disease.

  17. Morphological and intracellular alterations induced by Serratia marcescens cytotoxin.

    Science.gov (United States)

    Carbonell, Gleize Villela; Falcón, Rosabel; Yamada, Aureo T; da Fonseca, Benedito Antonio Lopes; Yano, Tomomasa

    2004-01-01

    In the present work, in vitro assays were used to investigate the toxicity of Serratia marcescens cytotoxin in cultured Chinese hamster ovary (CHO) cells. The time necessary to detect cellular alterations such as the onset of apoptosis, the perturbation of mitochondrial function, and cytoskeletal changes was assessed. The internalization of the cytotoxin by CHO cells was also examined. Within 10-15 min of exposure to cytotoxin, CHO cells became round, the nucleus shrank, the chromatin became more compact, and cytoplasmic blebs appeared on the cell surface. TUNEL (TdT-mediated dUTP nick end labeling) and propidium iodide staining identified some nuclei with fragmented DNA, and electrophoresis of CHO cell DNA obtained after 30-min exposure to S. marcescens toxin showed a pattern of DNA fragments typically associated with apoptosis. The cells also lost their characteristic actin organization within 10 min of exposure to cytotoxin. Lactate dehydrogenase leakage was detected after 20-min exposure to the cytotoxin and increased with time thereafter. Concomitantly, there was a time-dependent reduction in mitochondrial activity. Fluorescein-labeled S. marcescens cytotoxin was detected only on the surface of CHO cells, even after 30-min exposure to the toxin. These results show that there was no internalization of the toxin by CHO cells, and that, once bound to the cell surface, the toxin was able to induce changes in intracellular metabolism and to trigger cell death by apoptosis.

  18. IQGAP1: a regulator of intracellular spacetime relativity.

    Science.gov (United States)

    Malarkannan, Subramaniam; Awasthi, Aradhana; Rajasekaran, Kamalakannan; Kumar, Pawan; Schuldt, Kristina M; Bartoszek, Allison; Manoharan, Niranjan; Goldner, Nicholas K; Umhoefer, Colleen M; Thakar, Monica S

    2012-03-01

    Activating and inhibiting receptors of lymphocytes collect valuable information about their mikròs kósmos. This information is essential to initiate or to turn off complex signaling pathways. Irrespective of these advances, our knowledge on how these intracellular activation cascades are coordinated in a spatiotemporal manner is far from complete. Among multiple explanations, the scaffolding proteins have emerged as a critical piece of this evolutionary tangram. Among many, IQGAP1 is one of the essential scaffolding proteins that coordinate multiple signaling pathways. IQGAP1 possesses multiple protein interaction motifs to achieve its scaffolding functions. Using these domains, IQGAP1 has been shown to regulate a number of essential cellular events. This includes actin polymerization, tubulin multimerization, microtubule organizing center formation, calcium/calmodulin signaling, Pak/Raf/Mek1/2-mediated Erk1/2 activation, formation of maestrosome, E-cadherin, and CD44-mediated signaling and glycogen synthase kinase-3/adenomatous polyposis coli-mediated β-catenin activation. In this review, we summarize the recent developments and exciting new findings of cellular functions of IQGAP1.

  19. Subcellular site and nature of intracellular cadmium in plants

    Energy Technology Data Exchange (ETDEWEB)

    Wagner, George J.

    1979-01-01

    The mechanisms underlying heavy metal accumulation, toxicity and tolerance in higher plants are poorly understood. Since subcellular processes are undoubtedly involved in all these phenomena, it is of interest to study the extent of, subcellular site of and nature of intracellularly accumulated cadmium in higher plants. Whole plants supplied /sup 109/CdCl/sub 2/ or /sup 112/CdSO/sub 4/ accumulated Cd into roots and aerial tissues. Preparation of protoplasts from aerial tissue followed by subcellular fractionation of the protoplasts to obtain intact vacuoles, chloroplasts and cytosol revealed the presence of Cd in the cytosol but not in vacuoles or chloroplasts. Particulate materials containing other cell components were also labeled. Of the /sup 109/Cd supplied to plants, 2 to 10% was recovered in both cytosol preparations and in particulate materials. Cytosol contained proteinaceous--Cd complexes, free metal and low molecular weight Cd complexes. Labeling of protoplasts gave similar results. No evidence was obtained for the production of volatile Cd complexes in tobacco.

  20. Intracellular protein mass spectroscopy using mid-infrared laser ionization

    Science.gov (United States)

    Awazu, K.; Suzuki, S.

    2007-07-01

    Large-scale analysis of proteins, which can be regarded as functional biomolecule, assumes an important role in the life science. A MALDI using an ultraviolet laser (UV-MALDI) is one of ionization methods without fragmentation and has achieved conformation analysis of proteins. Recently, protein analysis has shifted from conformation analysis to functional and direct one that reserves posttranslational modifications such as the sugar chain addition and phosphorylation. We have proposed a MALDI using a mid-infrared tunable laser (IR-MALDI) as a new ionization method. IR-MALDI is promising because most biomolecules have a specific absorption in mid-infrared range, and IR-MALDI is expected to offer; (1) use of various matrices, (2) use of biomolecules such as water and lipid as the matrix, and (3) super-soft ionization. First, we evaluated the wavelength dependence of ionization of different matrices using a difference frequency generation (DFG) laser, which can tune the wavelength within a range from 5.5 to 10.0 μm. As results, ionization was specifically occurred at 5.8 μm which the C=O vibration stretching bond in matrix material and mass spectrum was observed. Next, protein mass spectrum was observed in the culture cells, MIN6, which secrete insulin, without the conventional cell-preparation processes. We demonstrate that the IR-MALDI has an advantage over the conventional method (UV-MALDI) in direct analysis of intracellular proteins.

  1. Intracellular Adenosine Triphosphate Deprivation through Lanthanide-Doped Nanoparticles.

    Science.gov (United States)

    Tian, Jing; Zeng, Xiao; Xie, Xiaoji; Han, Sanyang; Liew, Oi-Wah; Chen, Yei-Tsung; Wang, Lianhui; Liu, Xiaogang

    2015-05-27

    Growing interest in lanthanide-doped nanoparticles for biological and medical uses has brought particular attention to their safety concerns. However, the intrinsic toxicity of this new class of optical nanomaterials in biological systems has not been fully evaluated. In this work, we systematically evaluate the long-term cytotoxicity of lanthanide-doped nanoparticles (NaGdF4 and NaYF4) to HeLa cells by monitoring cell viability (mitochondrial activity), adenosine triphosphate (ATP) level, and cell membrane integrity (lactate dehydrogenase release), respectively. Importantly, we find that ligand-free lanthanide-doped nanoparticles induce intracellular ATP deprivation of HeLa cells, resulting in a significant decrease in cell viability after exposure for 7 days. We attribute the particle-induced cell death to two distinct cell death pathways, autophagy and apoptosis, which are primarily mediated via the interaction between the nanoparticle and the phosphate group of cellular ATP. The understanding gained from the investigation of cytotoxicity associated with lanthanide-doped nanoparticles provides keen insights into the safe use of these nanoparticles in biological systems.

  2. Comparison of the intracellular trafficking itinerary of ctla-4 orthologues.

    Directory of Open Access Journals (Sweden)

    Satdip Kaur

    Full Text Available CTLA-4 is an essential inhibitor of T cell immune responses. At steady state, most CTLA-4 resides in intracellular compartments due to constitutive internalisation mediated via a tyrosine based endocytic motif (YVKM within the cytoplasmic domain. This domain is highly conserved in mammals suggesting strong selective pressure. In contrast, the C-terminal domain varies considerably in non-mammals such as fish, xenopus and birds. We compared the ability of the C-terminus of these species to direct the trafficking of CTLA-4 with human CTLA-4. Using a chimeric approach, endocytosis was found to be conserved between human, xenopus and chicken CTLA-4 but was reduced substantially in trout CTLA-4, which lacks the conserved YXXM motif. Nevertheless, we identified an alternative YXXF motif in trout CTLA-4 that permitted limited endocytosis. Post-internalisation, CTLA-4 was either recycled or targeted for degradation. Human and chicken CTLA-4, which contain a YVKM motif, showed efficient recycling compared to xenopus CTLA-4 which contains a less efficient YEKM motif. Specific mutation of this motif in human CTLA-4 reduced receptor recycling. These findings suggest evolutionary development in the endocytic and recycling potential of CTLA-4, which may facilitate more refined functions of CTLA-4 within the mammalian immune system.

  3. Backscattered light confocal imaging of intracellular MTT-formazan crystals.

    Science.gov (United States)

    Bernas, Tytus; Dobrucki, Jurek W

    2004-06-01

    Metabolically active animal and plant cells reduce MTT tetrazolium salt to a corresponding nonfluorescent formazan. Reduction of MTT by viable cells is exploited in a number of tests widely used in biological research. The aim of this study was to optimize a microscopy method of detecting small crystals of MTT-formazan formed in intact cells maintained in in vitro cultures. We examined scattering properties of small intracellular crystals of MTT formazan and found that the efficiency of light scattering was dependent on wavelength. Small (formazan, formed in viable cells, scattered red, but not blue, light. Large crystals, which are formed later at a stage when cells begin to lose viability, scattered both red and blue light. We conclude that optimal detection of early stages of crystallization of MTT-formazan in living cells is possible using confocal microscopy of red, but not blue, scattered light. High contrast and resolution of images can be achieved by filtering out interference effects in the frequency domain.

  4. The Chlamydia psittaci genome: a comparative analysis of intracellular pathogens.

    Directory of Open Access Journals (Sweden)

    Anja Voigt

    Full Text Available BACKGROUND: Chlamydiaceae are a family of obligate intracellular pathogens causing a wide range of diseases in animals and humans, and facing unique evolutionary constraints not encountered by free-living prokaryotes. To investigate genomic aspects of infection, virulence and host preference we have sequenced Chlamydia psittaci, the pathogenic agent of ornithosis. RESULTS: A comparison of the genome of the avian Chlamydia psittaci isolate 6BC with the genomes of other chlamydial species, C. trachomatis, C. muridarum, C. pneumoniae, C. abortus, C. felis and C. caviae, revealed a high level of sequence conservation and synteny across taxa, with the major exception of the human pathogen C. trachomatis. Important differences manifest in the polymorphic membrane protein family specific for the Chlamydiae and in the highly variable chlamydial plasticity zone. We identified a number of psittaci-specific polymorphic membrane proteins of the G family that may be related to differences in host-range and/or virulence as compared to closely related Chlamydiaceae. We calculated non-synonymous to synonymous substitution rate ratios for pairs of orthologous genes to identify putative targets of adaptive evolution and predicted type III secreted effector proteins. CONCLUSIONS: This study is the first detailed analysis of the Chlamydia psittaci genome sequence. It provides insights in the genome architecture of C. psittaci and proposes a number of novel candidate genes mostly of yet unknown function that may be important for pathogen-host interactions.

  5. Biochemistry and pathophysiology of intravascular and intracellular lipolysis.

    Science.gov (United States)

    Young, Stephen G; Zechner, Rudolf

    2013-03-01

    All organisms use fatty acids (FAs) for energy substrates and as precursors for membrane and signaling lipids. The most efficient way to transport and store FAs is in the form of triglycerides (TGs); however, TGs are not capable of traversing biological membranes and therefore need to be cleaved by TG hydrolases ("lipases") before moving in or out of cells. This biochemical process is generally called "lipolysis." Intravascular lipolysis degrades lipoprotein-associated TGs to FAs for their subsequent uptake by parenchymal cells, whereas intracellular lipolysis generates FAs and glycerol for their release (in the case of white adipose tissue) or use by cells (in the case of other tissues). Although the importance of lipolysis has been recognized for decades, many of the key proteins involved in lipolysis have been uncovered only recently. Important new developments include the discovery of glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 (GPIHBP1), the molecule that moves lipoprotein lipase from the interstitial spaces to the capillary lumen, and the discovery of adipose triglyceride lipase (ATGL) and comparative gene identification-58 (CGI-58) as crucial molecules in the hydrolysis of TGs within cells. This review summarizes current views of lipolysis and highlights the relevance of this process to human disease.

  6. Enhancing Endosomal Escape for Intracellular Delivery of Macromolecular Biologic Therapeutics.

    Science.gov (United States)

    Lönn, Peter; Kacsinta, Apollo D; Cui, Xian-Shu; Hamil, Alexander S; Kaulich, Manuel; Gogoi, Khirud; Dowdy, Steven F

    2016-09-08

    Bioactive macromolecular peptides and oligonucleotides have significant therapeutic potential. However, due to their size, they have no ability to enter the cytoplasm of cells. Peptide/Protein transduction domains (PTDs), also called cell-penetrating peptides (CPPs), can promote uptake of macromolecules via endocytosis. However, overcoming the rate-limiting step of endosomal escape into the cytoplasm remains a major challenge. Hydrophobic amino acid R groups are known to play a vital role in viral escape from endosomes. Here we utilize a real-time, quantitative live cell split-GFP fluorescence complementation phenotypic assay to systematically analyze and optimize a series of synthetic endosomal escape domains (EEDs). By conjugating EEDs to a TAT-PTD/CPP spilt-GFP peptide complementation assay, we were able to quantitatively measure endosomal escape into the cytoplasm of live cells via restoration of GFP fluorescence by intracellular molecular complementation. We found that EEDs containing two aromatic indole rings or one indole ring and two aromatic phenyl groups at a fixed distance of six polyethylene glycol (PEG) units from the TAT-PTD-cargo significantly enhanced cytoplasmic delivery in the absence of cytotoxicity. EEDs address the critical rate-limiting step of endosomal escape in delivery of macromolecular biologic peptide, protein and siRNA therapeutics into cells.

  7. Cell-targeting aptamers act as intracellular delivery vehicles.

    Science.gov (United States)

    Gopinath, Subash C B; Lakshmipriya, Thangavel; Chen, Yeng; Arshad, M K Md; Kerishnan, Jesinda P; Ruslinda, A R; Al-Douri, Yarub; Voon, C H; Hashim, Uda

    2016-08-01

    Aptamers are single-stranded nucleic acids or peptides identified from a randomized combinatorial library through specific interaction with the target of interest. Targets can be of any size, from small molecules to whole cells, attesting to the versatility of aptamers for binding a wide range of targets. Aptamers show drug properties that are analogous to antibodies, with high specificity and affinity to their target molecules. Aptamers can penetrate disease-causing microbial and mammalian cells. Generated aptamers that target surface biomarkers act as cell-targeting agents and intracellular delivery vehicles. Within this context, the "cell-internalizing aptamers" are widely investigated via the process of cell uptake with selective binding during in vivo systematic evolution of ligands by exponential enrichment (SELEX) or by cell-internalization SELEX, which targets cell surface antigens to be receptors. These internalizing aptamers are highly preferable for the localization and functional analyses of multiple targets. In this overview, we discuss the ways by which internalizing aptamers are generated and their successful applications. Furthermore, theranostic approaches featuring cell-internalized aptamers are discussed with the purpose of analyzing and diagnosing disease-causing pathogens.

  8. How do antimalarial drugs reach their intracellular targets?

    Directory of Open Access Journals (Sweden)

    Katherine eBasore

    2015-05-01

    Full Text Available Drugs represent the primary treatment available for human malaria, as caused by Plasmodium spp. Currently approved drugs and antimalarial drug leads generally work against parasite enzymes or activities within infected erythrocytes. To reach their specific targets, these chemicals must cross at least three membranes beginning with the host cell membrane. Uptake at each membrane may involve partitioning and diffusion through the lipid bilayer or facilitated transport through channels or carriers. Here, we review the features of available antimalarials and examine whether transporters may be required for their uptake. Our computational analysis suggests that most antimalarials have high intrinsic membrane permeability, obviating the need for uptake via transporters; a subset of compounds appear to require facilitated uptake. We also review parasite and host transporters that may contribute to drug uptake. Broad permeability channels at the erythrocyte and parasitophorous vacuolar membranes of infected cells relax permeability constraints on antimalarial drug design; however, this uptake mechanism is prone to acquired resistance as the parasite may alter channel activity to reduce drug uptake. A better understanding of how antimalarial drugs reach their intracellular targets is critical to prioritizing drug leads for antimalarial development and may reveal new targets for therapeutic intervention.

  9. Intracellular pH gradients in migrating cells

    DEFF Research Database (Denmark)

    Martin, Christine; Pedersen, Stine Helene Falsig; Schwab, Albrecht

    2011-01-01

    might function as such unevenly distributed regulators as they modulate the interaction of focal adhesion proteins and components of the cytoskeleton in vitro. However, an intracellular pH (pH(i)) gradient reflecting a spatial asymmetry of protons has not been shown so far. One major regulator of pH......(i), the Na(+)/H(+) exchanger NHE1, is essential for cell migration and accumulates at the cell front. Here, we test the hypothesis that the uneven distribution of NHE1 activity creates a pH(i) gradient in migrating cells. Using the pH-sensitive fluorescent dye BCECF, pH(i) was measured in five cell lines (MV......3, B16V, NIH3T3, MDCK-F1, EA.hy926) along the axis of movement. Differences in pH(i) between the front and the rear end (¿pH(i) front-rear) were present in all cell lines, and inhibition of NHE1 either with HOE642 or by absence of extracellular Na(+) caused the pH(i) gradient to flatten or disappear...

  10. Transepithelial Na+ transport and the intracellular fluids: a computer study.

    Science.gov (United States)

    Civan, M M; Bookman, R J

    1982-01-01

    Computer simulations of tight epithelia under three experimental conditions have been carried out, using the rheogenic nonlinear model of Lew, Ferreira and Moura (Proc. Roy. Soc. London. B 206:53-83, 1979) based largely on the formulation of Koefoed-Johnsen and Ussing (Acta Physiol. Scand. 42: 298-308. 1958). First, analysis of the transition between the short-circuited and open-circuited states has indicated that (i) apical Cl- permeability is a critical parameter requiring experimental definition in order to analyze cell volume regulation, and (ii) contrary to certain experimental reports, intracellular Na+ concentration (ccNa) is expected to be a strong function of transepithelial clamping voltage. Second, analysis of the effects of lowering serosal K+ concentration (csK) indicates that the basic model cannot simulate several well-documented observations; these defects can be overcome, at least qualitatively, by modifying the model to take account of the negative feedback interaction likely to exist between the apical Na+ permeability and ccNa. Third, analysis of the strongly supports the concept that osmotically induced permeability changes in the apical intercellular junctions play a physiological role in conserving the body's stores of NaCl. The analyses also demonstrate that the importance of Na+ entry across the basolateral membrane is strongly dependent upon transepithelial potential, cmNa and csK; under certain conditions, net Na+ entry could be appreciably greater across the basolateral than across the apical membrane.

  11. Isolate and genetic heterogeneity of psychrotrophic lactic acid bacteria from the intestinal tract of cold-water fishes from the Eerqisi river, Xinjang%新疆额尔齐斯河流域冷水鱼肠道耐低温乳酸菌的分离筛选及其遗传差异

    Institute of Scientific and Technical Information of China (English)

    高阳; 王俊钢; 顾燕玲; 邓梅; 周红; 倪永清

    2013-01-01

    [Objective] The purpose of this study was to select cold-adapted lactic acid bacteria ( LAB) from the intestinal tract of cold-water fishes from the Eerqisi river Alatai. Xinjiang. [ Methods] By using culture medium MRS and Elliker, isolation of lactic acid bacteria ( LAB) from 9 intestinal canal of cold-water fishes was carried out. Taxonomic identity and genetic diversity of strains isolated were determined by partial 16S rRNA gene sequences and rep-PCR with three different kinds of primers named BOX, (GTG),, ERIC. [Results] A total of 78 psychrotrophic isolates were obtained. Among them, 24 isolates had characteristics of LAB and showed the optimal growth temperature ranging from 15 to 24℃. The phylogenesis result showed that 24 strains belonged to 6 Cenuses, Carnobacterium (3 strains) , Lactococcus (9 strains), Enterococcus (7 strains), Brochothrix (1 strain), Weissella (2 strains), Streptococcus (2 strains). Rep-PCR clustering analysis showed that Lactococcus and Enterococcus strains were from different Species. Lactococcus were belonged to 4 species while Enterococcus assigned to 2 species. [ Conclusion ] The phylogenetic diversity of cold-adapted LAB from the intestinal tract of cold-water Fishes from the Eerqisi river in the Alatai Xinjiang was relatively abundant.%[目的]从新疆阿勒泰地区额尔齐斯河流域冷水鱼肠道中分离耐低温的乳酸菌,挖掘乳酸菌的物种和遗传多样性,为低温乳酸菌生物技术研发提供优良菌种.[方法]利用MRS、Elliker两种不同培养基从9种冷水鱼肠道中,分离筛选出耐低温菌,测定细菌最适生长温度并进行常规生理生化实验.根据16S rRNA基因序列初步确定耐低温菌的系统进化地位,并采用BOX、(GTG)5、ERIC三种不同的引物进行rep-PCR,对16S rRNA基因高度同源性的菌株进一步区分.[结果]分离得到78株耐冷菌,最终确定有24株为耐低温乳酸菌,菌株的最适生长温度在15-24℃之间.

  12. Photoactivatable Drug-Caged Fluorophore Conjugate Allows Direct Quantification of Intracellular Drug Transport

    Science.gov (United States)

    Kohler, Rainer H.; Weissleder, Ralph

    2013-01-01

    We report here a method that utilizes photoactivatable drug-caged fluorophore conjugate to quantify intracellular drug trafficking processes at single cell resolution. Photoactivation is performed in labeled cellular compartments to visualize intracellular drug exchange at physiologic conditions, without the need for washing, facilitating its translation to in vivo cancer models. PMID:24135896

  13. Differentiating Intracellular from Extracellular Alkaline Phosphatase Activity in Soil by Sonication

    NARCIS (Netherlands)

    Qin, S.P.; Hu, C.S.; Oenema, O.

    2013-01-01

    Differentiating intracellular from extracellular enzyme activity is important in soil enzymology, but not easy. Here, we report on an adjusted sonication method for the separation of intracellular from extracellular phosphatase activity in soil. Under optimal sonication conditions [soil:water ratio

  14. Boron nitride nanotubes functionalized with mesoporous silica for intracellular delivery of chemotherapy drugs.

    Science.gov (United States)

    Li, Xia; Zhi, Chunyi; Hanagata, Nobutaka; Yamaguchi, Maho; Bando, Yoshio; Golberg, Dmitri

    2013-08-25

    Boron nitride nanotube (BNNT)@mesoporous silica hybrids with controllable surface zeta potential were fabricated for intracellular delivery of doxorubicin. The materials showed higher suspension ability, doxorubicin intracellular endocytosis efficiency, and LNcap prostate cancer cell killing ability compared with naked BNNTs.

  15. Thioredoxin 80-Activated-Monocytes (TAMs) Inhibit the Replication of Intracellular Pathogens

    DEFF Research Database (Denmark)

    Cortes-Bratti, Ximena; Brasseres, Eugenie; Herrera-Rodriquez, Fabiola

    2011-01-01

    for a role of TAMs in the control of intracellular bacterial infections. As model pathogens we have chosen Listeria monocytogenes and Brucella abortus which replicate in the cytosol and the endoplasmic reticulum respectively. Our data indicate that TAMs efficiently inhibit intracellular growth of both L...

  16. Microscopy tools for the investigation of intracellular lipid storage and dynamics

    Directory of Open Access Journals (Sweden)

    Sabine Daemen

    2016-03-01

    Major conclusions: The continuous development of lipid probes in combination with the rapid development of microscopic techniques can provide new insights in the role and dynamics of intracellular lipids. Moreover, in situ identification of intracellular lipids is now possible and promises to add a new dimensionality to analysis of lipid biochemistry, and its relation to (pathophysiology.

  17. Intracellular plant microbe associations: secretory pathways and the formation of perimicrobial compartments

    NARCIS (Netherlands)

    Ivanov, S.E.; Fedorova, E.; Bisseling, T.

    2010-01-01

    Plants can establish intracellular interactions with symbiotic as well as pathogenic microbes. Such intracellular accommodation of microbes always involves the formation of a host membrane compartment - the interface between the cytoplasm of the host and the microbe. These are the so-called perimicr

  18. Intracellular signal transduction by the extracellular calcium-sensing receptor of Xenopus melanotrope cells.

    NARCIS (Netherlands)

    Hurk, MJ van den; Cruijsen, P.M.; Schoeber, J.P.H.; Scheenen, W.J.J.M.; Roubos, E.W.; Jenks, B.G.

    2008-01-01

    The extracellular calcium-sensing receptor (CaR) i