WorldWideScience

Sample records for bacteriophage-mediated hemocyte agglutination

  1. Hemocyte-mediated phagocytosis differs between honey bee (Apis mellifera) worker castes.

    Science.gov (United States)

    Hystad, Eva Marit; Salmela, Heli; Amdam, Gro Vang; Münch, Daniel

    2017-01-01

    Honey bees as other insects rely on the innate immune system for protection against diseases. The innate immune system includes the circulating hemocytes (immune cells) that clear pathogens from hemolymph (blood) by phagocytosis, nodulation or encapsulation. Honey bee hemocyte numbers have been linked to hemolymph levels of vitellogenin. Vitellogenin is a multifunctional protein with immune-supportive functions identified in a range of species, including the honey bee. Hemocyte numbers can increase via mitosis, and this recruitment process can be important for immune system function and maintenance. Here, we tested if hemocyte mediated phagocytosis differs among the physiologically different honey bee worker castes (nurses, foragers and winter bees), and study possible interactions with vitellogenin and hemocyte recruitment. To this end, we adapted phagocytosis assays, which-together with confocal microscopy and flow cytometry-allow qualitative and quantitative assessment of hemocyte performance. We found that nurses are more efficient in phagocytic uptake than both foragers and winter bees. We detected vitellogenin within the hemocytes, and found that winter bees have the highest numbers of vitellogenin-positive hemocytes. Connections between phagocytosis, hemocyte-vitellogenin and mitosis were worker caste dependent. Our results demonstrate that the phagocytic performance of immune cells differs significantly between honey bee worker castes, and support increased immune competence in nurses as compared to forager bees. Our data, moreover, provides support for roles of vitellogenin in hemocyte activity.

  2. Hemocyte-mediated phagocytosis differs between honey bee (Apis mellifera worker castes.

    Directory of Open Access Journals (Sweden)

    Eva Marit Hystad

    Full Text Available Honey bees as other insects rely on the innate immune system for protection against diseases. The innate immune system includes the circulating hemocytes (immune cells that clear pathogens from hemolymph (blood by phagocytosis, nodulation or encapsulation. Honey bee hemocyte numbers have been linked to hemolymph levels of vitellogenin. Vitellogenin is a multifunctional protein with immune-supportive functions identified in a range of species, including the honey bee. Hemocyte numbers can increase via mitosis, and this recruitment process can be important for immune system function and maintenance. Here, we tested if hemocyte mediated phagocytosis differs among the physiologically different honey bee worker castes (nurses, foragers and winter bees, and study possible interactions with vitellogenin and hemocyte recruitment. To this end, we adapted phagocytosis assays, which-together with confocal microscopy and flow cytometry-allow qualitative and quantitative assessment of hemocyte performance. We found that nurses are more efficient in phagocytic uptake than both foragers and winter bees. We detected vitellogenin within the hemocytes, and found that winter bees have the highest numbers of vitellogenin-positive hemocytes. Connections between phagocytosis, hemocyte-vitellogenin and mitosis were worker caste dependent. Our results demonstrate that the phagocytic performance of immune cells differs significantly between honey bee worker castes, and support increased immune competence in nurses as compared to forager bees. Our data, moreover, provides support for roles of vitellogenin in hemocyte activity.

  3. Vibrio cholerae interactions with Mytilus galloprovincialis hemocytes mediated by serum components.

    Directory of Open Access Journals (Sweden)

    Laura eCanesi

    2013-12-01

    Full Text Available Edible bivalves (e.g., mussels, oysters can accumulate large amount of bacteria in their tissues and act as passive carriers of pathogens to humans. Bacterial persistence inside bivalves depends, at least in part, on hemolymph anti-bacterial activity that is exerted by both serum soluble factors and phagocytic cells (i.e., the hemocytes. It was previously shown that Mytilus galloprovincialis hemolymph serum contains opsonins that mediate D-mannose-sensitive interactions between hemocytes and V. cholerae O1 El Tor bacteria that carry the Mannose–Sensitive Hemagglutinin (MSHA. These opsonins enhance phagocytosis and killing of vibrios by facilitating their binding to hemocytes. Since V. cholerae strains not carrying the MSHA ligand (O1 classical, non O1/O139 are present in coastal water and can be entrapped by mussels, we studied whether in mussel serum, in addition to opsonins directed towards MSHA, other components can mediate opsonization of these bacteria. By comparing interactions of O1 classical and non O1/O139 strains with hemocytes in ASW and serum, it was found that M. galloprovincialis serum contains components that increase by at approximately two fold their adhesion to, association with and killing by hemocytes. Experiments conducted with high and low molecular mass fractions obtained by serum ultrafiltration indicated that these compounds have molecular mass higher than 5000 Da. Serum exposure to high temperature (80°C abolished its opsonizing capability suggesting that the involved serum active components are of protein nature. Further studies are needed to define the chemical properties and specificity of both the involved bacterial ligands and hemolymph opsonins. This information will be central not only to better understand V. cholerae ecology, but also to improve current bivalve depuration practices and properly protect human health.

  4. Hemocyte-mediated phagocytosis and melanization in the mosquito Armigeres subalbatus following immune challenge by bacteria.

    Science.gov (United States)

    Hillyer, Julián F; Schmidt, Shelley L; Christensen, Bruce M

    2003-07-01

    Mosquitoes are important vectors of disease. These insects respond to invading organisms with strong cellular and humoral immune responses that share many similarities with vertebrate immune systems. The strength and specificity of these responses are directly correlated to a mosquito's ability to transmit disease. In the current study, we characterized the hemocytes (blood cells) of Armigeres subalbatus by morphology (ultrastructure), lectin binding, enzyme activity, immunocytochemistry, and function. We found four hemocyte types: granulocytes, oenocytoids, adipohemocytes, and thrombocytoids. Granulocytes contained acid phosphatase activity and bound the exogenous lectins Helix pomatia agglutinin, Galanthus nivalis lectin, and wheat germ agglutinin. Following bacteria inoculation, granulocytes mounted a strong phagocytic response as early as 5 min postexposure. Bacteria also elicited a hemocyte-mediated melanization response. Phenoloxidase, the rate-limiting enzyme in the melanization pathway, was present exclusively in oenocytoids and in many of the melanotic capsules enveloping bacteria. The immune responses mounted against different bacteria were not identical; gram(-) Escherichia coli were predominantly phagocytosed and gram(+) Micrococcus luteus were melanized. These studies implicate hemocytes as the primary line of defense against bacteria.

  5. Rac1 mediates cytokine-stimulated hemocyte spreading via prostaglandin biosynthesis in the beet armyworm, Spodoptera exigua

    Science.gov (United States)

    Cell spreading is an integral component of insect hemocytic immune reactions to infections and invasions. Cell spreading is accomplished by cytoskeleton rearrangement, which is activated by three major immune mediators, biogenic monoamines, plasmatocyte-spreading peptide (PSP), and eicosanoids, part...

  6. A simple and novel modification of comet assay for determination of bacteriophage mediated bacterial cell lysis.

    Science.gov (United States)

    Khairnar, Krishna; Sanmukh, Swapnil; Chandekar, Rajshree; Paunikar, Waman

    2014-07-01

    The comet assay is the widely used method for in vitro toxicity testing which is also an alternative to the use of animal models for in vivo testing. Since, its inception in 1984 by Ostling and Johansson, it is being modified frequently for a wide range of application. In spite of its wide applicability, unfortunately there is no report of its application in bacteriophages research. In this study, a novel application of comet assay for the detection of bacteriophage mediated bacterial cell lysis was described. The conventional methods in bacteriophage research for studying bacterial lysis by bacteriophages are plaque assay method. It is time consuming, laborious and costly. The lytic activity of bacteriophage devours the bacterial cell which results in the release of bacterial genomic material that gets detected by ethidium bromide staining method by the comet assay protocol. The objective of this study was to compare efficacy of comet assay with different assay used to study phage mediated bacterial lysis. The assay was performed on culture isolates (N=80 studies), modified comet assay appear to have relatively higher sensitivity and specificity than other assay. The results of the study showed that the application of comet assay can be an economical, time saving and less laborious alternative to conventional plaque assay for the detection of bacteriophage mediated bacterial cell lysis. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. Good agreement of conventional and gel-based direct agglutination test in immune-mediated haemolytic anaemia

    NARCIS (Netherlands)

    Piek, C.J.; Teske, E.; van Leeuwen, M.W.; Day, M.J.

    2012-01-01

    Abstract Background The aim of this study was to compare a gel-based test with the traditional direct agglutination test (DAT) for the diagnosis of immune-mediated haemolytic anaemia (IMHA). Methods Canine (n = 247) and feline (n = 74) blood samples were submitted for DAT testing to two

  8. Non-Identity-Mediated CRISPR-Bacteriophage Interaction Mediated via the Csy and Cas3 Proteins ▿#

    Science.gov (United States)

    Cady, Kyle C.; O'Toole, George A.

    2011-01-01

    Studies of the Escherichia, Neisseria, Thermotoga, and Mycobacteria clustered regularly interspaced short palindromic repeat (CRISPR) subtypes have resulted in a model whereby CRISPRs function as a defense system against bacteriophage infection and conjugative plasmid transfer. In contrast, we previously showed that the Yersinia-subtype CRISPR region of Pseudomonas aeruginosa strain UCBPP-PA14 plays no detectable role in viral immunity but instead is required for bacteriophage DMS3-dependent inhibition of biofilm formation by P. aeruginosa. The goal of this study is to define the components of the Yersinia-subtype CRISPR region required to mediate this bacteriophage-host interaction. We show that the Yersinia-subtype-specific CRISPR-associated (Cas) proteins Csy4 and Csy2 are essential for small CRISPR RNA (crRNA) production in vivo, while the Csy1 and Csy3 proteins are not absolutely required for production of these small RNAs. Further, we present evidence that the core Cas protein Cas3 functions downstream of small crRNA production and that this protein requires functional HD (predicted phosphohydrolase) and DEXD/H (predicted helicase) domains to suppress biofilm formation in DMS3 lysogens. We also determined that only spacer 1, which is not identical to any region of the DMS3 genome, mediates the CRISPR-dependent loss of biofilm formation. Our evidence suggests that gene 42 of phage DMS3 (DMS3-42) is targeted by CRISPR2 spacer 1 and that this targeting tolerates multiple point mutations between the spacer and DMS3-42 target sequence. This work demonstrates how the interaction between P. aeruginosa strain UCBPP-PA14 and bacteriophage DMS3 can be used to further our understanding of the diverse roles of CRISPR system function in bacteria. PMID:21398535

  9. Impacts of papain and neuraminidase enzyme treatment on electrohydrodynamics and IgG-mediated agglutination of type A red blood cells.

    Science.gov (United States)

    Hyono, Atsushi; Gaboriaud, Fabien; Mazda, Toshio; Takata, Youichi; Ohshima, Hiroyuki; Duval, Jérôme F L

    2009-09-15

    The stability of native and enzyme-treated human red blood cells of type A (Rh D positive) against agglutination is investigated under conditions where it is mediated by immunoglobuline G (IgG) anti-D antibody binding. The propensity of cells to agglutinate is related to their interphasic (electrokinetic) properties. These properties significantly depend on the concentration of proteolytic papain enzyme and protease-free neuraminidase enzyme that the cells are exposed to. The analysis is based on the interpretation of electrophoretic data of cells by means of the numerical theory for the electrokinetics of soft (bio)particles. A significant reduction of the hydrodynamic permeability of the external soft glycoprotein layer of the cells is reported under the action of papain. This reflects a significant decrease in soft surface layer thickness and a loss in cell surface integrity/rigidity, as confirmed by nanomechanical AFM analysis. Neuraminidase action leads to an important decrease in the interphase charge density by removing sialic acids from the cell soft surface layer. This is accompanied by hydrodynamic softness modulations less significant than those observed for papain-treated cells. On the basis of these electrohydrodynamic characteristics, the overall interaction potential profiles between two native cells and two enzyme-treated cells are derived as a function of the soft surface layer thickness in the Debye-Hückel limit that is valid for cell suspensions under physiological conditions (approximately 0.16 M). The thermodynamic computation of cell suspension stability against IgG-mediated agglutination then reveals that a decrease in the cell surface layer thickness is more favorable than a decrease in interphase charge density for inducing agglutination. This is experimentally confirmed by agglutination data collected for papain- and neuraminidase-treated cells.

  10. Mediated effect of ultrasound treated Diclofenac on mussel hemocytes: First evidence for the involvement of respiratory burst enzymes in the induction of DCF-mediated unspecific mode of action.

    Science.gov (United States)

    Toufexi, Eirini; Dailianis, Stefanos; Vlastos, Dimitris; Manariotis, Ioannis D

    2016-06-01

    The present study investigates the toxic behavior of diclofenac (DCF) before and after its ultrasound (US) treatment, as well as the involvement of intracellular target molecules, such as NADPH oxidase and NO synthase, in the DCF-induced adverse effects on hemocytes of mussel Mytilus galloprovincialis. In this context, appropriate volumes (350 and 500mL) of DCF solutions (at concentrations of 2, 2.5, 5 and 10mgL(-1)) were treated under different ultrasound operating conditions (frequency at 582 and 862kHz, electric power density at 133 and 167W) for assessing US method efficiency. In parallel, DCF and US DCF-mediated cytotoxic (in terms of cell viability measured with the use of neutral red uptake/NRU method), oxidative (in terms of superoxide anions/(.)O2(-), nitric oxides such as NO2(-) and lipid peroxidation products, such as malondialdehyde/MDA content) and genotoxic (DNA damage measured by the use of Comet assay method) effects were investigated in hemocytes exposed for 1h to 5, 10 and 100ngL(-1) and 1, 10 and 20μgL(-1) of DCF. The involvement of NADPH oxidase and NO synthase to the DCF-induced toxicity was further investigated by the use of 10μΜ L-NAME, a NO synthase inhibitor and 10μΜ DPI, a NADPH oxidase inhibitor. According to the results, 350mL of 2mgL(-1) DCF showed higher degradation (>50%) under 167W electric power density and frequency at 862kHz for 120min, compared to degradation in all other cases, followed by a significant elimination of its toxicity. Specifically, US DCF-treated hemocytes showed a significant attenuation of DCF-mediated cytotoxic, oxidative and genotoxic effects, which appeared to be caused by NADPH oxidase and NO synthase activation, since their inhibition was followed by a significant elimination of (.)O2(-) and NO2(-) generation and the concomitant oxidative damage within cells. The results of the present study showed for the first time that unspecific mode of action of DCF, associated with the induction of NADPH oxidase

  11. Genesis of a novel Shigella flexneri serotype by sequential infection of serotype-converting bacteriophages SfX and SfI

    Directory of Open Access Journals (Sweden)

    Sun Qiangzheng

    2011-12-01

    Full Text Available Abstract Background Shigella flexneri is the major pathogen causing bacillary dysentery. Fifteen serotypes have been recognized up to now. The genesis of new S. flexneri serotypes is commonly mediated by serotype-converting bacteriophages. Untypeable or novel serotypes from natural infections had been reported worldwide but have not been generated in laboratory. Results A new S. flexneri serotype-serotype 1 d was generated when a S. flexneri serotype Y strain (native LPS was sequentially infected with 2 serotype-converting bacteriophages, SfX first and then SfI. The new serotype 1 d strain agglutinated with both serotype X-specific anti-7;8 grouping serum and serotype 1a-specific anti- I typing serum, and differed from subserotypes 1a, 1b and 1c. Twenty four S. flexneri clinical isolates of serotype X were all converted to serotype 1 d by infection with phage SfI. PCR and sequencing revealed that SfI and SfX were integrated in tandem into the proA-yaiC region of the host chromosome. Conclusions These findings suggest a new S. flexneri serotype could be created in nature. Such a conversion may be constrained by susceptibility of a strain to infection by a given serotype-converting bacteriophage. This finding has significant implications in the emergence of new S. flexneri serotypes in nature.

  12. Hemocyte characterization of Nasutitermes coxipoensis (Holmgren) (Isoptera: Termitidae) workers and hemocyte evaluation after parasitism by Metarhizium anisopliae

    International Nuclear Information System (INIS)

    Cunha, Franklin M.; Wanderley-Teixeira, Valeria; Albuquerque, Auristela C.; Lima, Elza A.L.A.

    2009-01-01

    We aimed to characterize the morphology and ultrastructure of hemocytes of Nasutitermes coxipoensis (Holmgren) workers and to quantify the cell types 24h, 48h and 72h after inoculation with Metarhizium anisopliae. Six hemocytes types were identified, plasmatocyte, granulocyte, spherulocyte, prohemocyte, adipohemocyte and eonocytoid Hemocytes did not present any morphological alteration at the several observation periods, but they did have a change in their abundance, as observed for spherulocytes, adipohemocytes and eonocytoids at all intervals, and for plasmatocytes and granulocytes at 48h after host inoculation. We argue on the possible reasons and implications of the observed changes. (author)

  13. On the origin of the Biomphalaria glabrata hemocytes

    Directory of Open Access Journals (Sweden)

    Samaly dos Santos Souza

    2006-10-01

    Full Text Available A histologic, morphometric and ultrastructural study performed on Biomphalaria glabrata submitted to infection with Schistosoma mansoni miracidia failed to provide significant evidences that the so-called amebocyte-producing organ (APO is really the central organ for hemocyte production. In infected snails no general reactive changes appeared in the APO, the mitoses were seen only occasionally, and the possibility of cellular hyperplasia was ruled out by morphometric measurements. Under the electron microscope the APO cells presented an essentially epithelial structure, without features indicative of transition toward hemocytes. On the other hand, the present findings pointed to a multicentric origin for the mollusck hemocytes, as earlier studies had indicated. Dense foci of hemocyte collections appeared sometimes around disintegrating sporocysts and cercariae in several organs and tissues of the infected snails, including a curious accumulation of such cells inside the ventricular cavity of the heart. In the heart and other sites, features suggestive of transformation of vascular space endothelial lining cells into hemocytes were apparent. To some extent, the postulated multicentric origin for B. glabrata hemocytes recapitulates earlier embryologic findings in vertebrates, when mesenchymal vascular spaces generate the circulating and phagocytic blood cells.

  14. Osmoregulated Chloride Currents in Hemocytes from Mytilus galloprovincialis.

    Directory of Open Access Journals (Sweden)

    Monica Bregante

    Full Text Available We investigated the biophysical properties of the transport mediated by ion channels in hemocytes from the hemolymph of the bivalve Mytilus galloprovincialis. Besides other transporters, mytilus hemocytes possess a specialized channel sensitive to the osmotic pressure with functional properties similar to those of other transport proteins present in vertebrates. As chloride fluxes may play an important role in the regulation of cell volume in case of modifications of the ionic composition of the external medium, we focused our attention on an inwardly-rectifying voltage-dependent, chloride-selective channel activated by negative membrane potentials and potentiated by the low osmolality of the external solution. The chloride channel was slightly inhibited by micromolar concentrations of zinc chloride in the bath solution, while the antifouling agent zinc pyrithione did not affect the channel conductance at all. This is the first direct electrophysiological characterization of a functional ion channel in ancestral immunocytes of mytilus, which may bring a contribution to the understanding of the response of bivalves to salt and contaminant stresses.

  15. Vibrio cholerae O1 secretes an extracellular matrix in response to antibody-mediated agglutination.

    Directory of Open Access Journals (Sweden)

    Danielle E Baranova

    Full Text Available Vibrio cholerae O1 is one of two serogroups responsible for epidemic cholera, a severe watery diarrhea that occurs after the bacterium colonizes the human small intestine and secretes a potent ADP-ribosylating toxin. Immunity to cholera is associated with intestinal anti-lipopolysaccharide (LPS antibodies, which are known to inhibit V. cholerae motility and promote bacterial cell-cell crosslinking and aggregation. Here we report that V. cholerae O1 classical and El Tor biotypes produce an extracellular matrix (ECM when forcibly immobilized and agglutinated by ZAC-3 IgG, an intestinally-derived monoclonal antibody (MAb against the core/lipid A region of LPS. ECM secretion, as demonstrated by crystal violet staining and scanning electron microscopy, occurred within 30 minutes of antibody exposure and peaked by 3 hours. Non-motile mutants of V. cholerae did not secrete ECM following ZAC-3 IgG exposure, even though they were susceptible to agglutination. The ECM was enriched in O-specific polysaccharide (OSP but not Vibrio polysaccharide (VPS. Finally, we demonstrate that ECM production by V. cholerae in response to ZAC-3 IgG was associated with bacterial resistant to a secondary complement-mediated attack. In summary, we propose that V. cholerae O1, upon encountering anti-LPS antibodies in the intestinal lumen, secretes an ECM (or O-antigen capsule possibly as a strategy to shield itself from additional host immune factors and to exit an otherwise inhospitable host environment.

  16. Effect of gamma irradiation on the hemocyte-mediated immune response of Aedes aegypti against microfilariae

    International Nuclear Information System (INIS)

    Christensen, B.M.; Huff, B.M.; Li, J.

    1990-01-01

    The effect of gamma irradiation on the melanotic encapsulation response of Aedes aegypti black eye Liverpool strain against inoculated Dirofilaria immitis microfilariae (mff) was assessed at 1, 2, 3, and 6 days postinoculation (PI). Mosquitoes received 6000 rad from a 137Cs source (Shepard Mark I irradiator) at 3 days postemergence and were inoculated with 15-20 mff 24 hr later. These mosquitoes were compared to nonirradiated controls that also were inoculated with 15-20 mff at 3 days postemergence. The immune response was significantly reduced in irradiated mosquitoes as compared with controls at all days PI. Although the response was significantly inhibited compared with controls, irradiated mosquitoes were still capable of eliciting a response against 69% of recovered mff at 6 days PI. External gamma irradiation did not significantly affect the proliferation of hemocytes associated with the melanotic encapsulation response of A. aegypti. The number of circulating hemocytes increased in irradiated mosquitoes in response to inoculated mff in a manner similar to nonirradiated, inoculated controls. Hemocyte monophenol oxidase activity, however, was significantly reduced in gamma-irradiated mosquitoes at 12 hr PI as compared with controls. The reduced immunological capacity of irradiated mosquitoes might be related to an interference with gene activity required for the synthesis or activation of enzymes that are directly or indirectly involved in the biochemical processes associated with the production of melanotic substances that sequester mff

  17. Microbead agglutination based assays

    KAUST Repository

    Kodzius, Rimantas

    2013-01-21

    We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microbeads in the presence of a specific analyte thus enabling the macroscopic observation. Such tests are most often used to explore antibody-antigen reactions. Agglutination has been used for protein assays using a biotin/streptavidin system as well as a hybridization based assay. The agglutination systems are prone to selftermination of the linking analyte, prone to active site saturation and loss of agglomeration at high analyte concentrations. We investigated the molecular target/ligand interaction, explaining the common agglutination problems related to analyte self-termination, linkage of the analyte to the same bead instead of different microbeads. We classified the agglutination process into three kinds of assays: a two- component assay, a three-component assay and a stepped three- component assay. Although we compared these three kinds of assays for recognizing DNA and protein molecules, the assay can be used for virtually any molecule, including ions and metabolites. In total, the optimized assay permits detecting analytes with high sensitivity in a short time, 5 min, at room temperature. Such a system is appropriate for POC testing.

  18. Understanding the Role of Host Hemocytes in a Squid/Vibrio Symbiosis Using Transcriptomics and Proteomics

    Directory of Open Access Journals (Sweden)

    Andrew J. Collins

    2012-05-01

    Full Text Available The symbiosis between the squid, Euprymna scolopes, and the bacterium, Vibrio fischeri, serves as a model for understanding interactions between beneficial bacteria and animal hosts. The establishment and maintenance of the association is highly specific and depends on the selection of V. fischeri and exclusion of non-symbiotic bacteria from the environment. Current evidence suggests that the host’s cellular innate immune system, in the form of macrophage-like hemocytes, helps to mediate host tolerance of V. fischeri. To begin to understand the role of hemocytes in this association, we analyzed these cells by high-throughput 454 transcriptomic and liquid chromatography/ tandem mass spectrometry (LC-MS/MS proteomic analyses. 454 high-throughput sequencing produced 650,686 reads totaling 279.9 Mb while LC-MS/MS analyses of circulating hemocytes putatively identified 702 unique proteins. Several receptors involved with the recognition of microbial associated molecular patterns (MAMPs were identified. Among these was a complete open reading frame (ORF to a putative peptidoglycan recognition protein (EsPGRP5 that has conserved residues for amidase activity. Assembly of the hemocyte transcriptome showed EsPGRP5 had high coverage, suggesting it is among the 5% most abundant transcripts in circulating hemocytes. Other transcripts and proteins identified included members of the conserved NFκB signaling pathway, putative members of the complement pathway, the carbohydrate binding protein galectin, and cephalotoxin. Quantitative PCR of complement-related genes, cephalotoxin, EsPGRP5, and a nitric oxide synthase showed differential expression in circulating hemocytes isolated from adult squid with colonized light organs compared to those for which the symbionts were removed. These data suggest that the presence of the symbiont influences gene expression of the cellular innate immune system of the host.

  19. Two Hemocyte Lineages Exist in Silkworm Larval Hematopoietic Organ

    OpenAIRE

    Nakahara, Yuichi; Kanamori, Yasushi; Kiuchi, Makoto; Kamimura, Manabu

    2010-01-01

    BACKGROUND: Insects have multiple hemocyte morphotypes with different functions as do vertebrates, however, their hematopoietic lineages are largely unexplored with the exception of Drosophila melanogaster. METHODOLOGY/PRINCIPAL FINDINGS: To study the hematopoietic lineage of the silkworm, Bombyx mori, we investigated in vivo and in vitro differentiation of hemocyte precursors in the hematopoietic organ (HPO) into the four mature hemocyte subsets, namely, plasmatocytes, granulocytes, oenocyto...

  20. Hemiuroid trematode sporocysts are undetected by hemocytes of their intermediate host, the ark cockle Anadara trapezia: potential role of surface carbohydrates in successful parasitism.

    Science.gov (United States)

    Kawasaki, Minami; Delamare-Deboutteville, Jerome; Dang, Cecile; Barnes, Andrew C

    2013-12-01

    In order to establish a successful relationship with their hosts, parasites must subvert or evade immune defences. Cockle Anadara trapezia and Sydney Rock oyster (SRO) Saccostrea glomerata live in the same location but only ark cockles are infected by sporocysts of hemiuroid trematode. This provides an opportunity to explore differing interactions between the parasite and the immune system of susceptible and refractive hosts. Rapid migration and encapsulation of sporocysts was observed by SRO hemocytes but not by cockle hemocytes. This migration/encapsulation was inhibited by N-acetylglucosamine or N-acetylgalactosamine but not by the other sugars, implicating specific surface carbohydrates in immune detection. Effector responses of hemocytes were investigated in vitro in terms of production of reactive oxygen production (ROS). Hemocytes of both species strongly reacted to Zymosan, but only SRO hemocytes responded to live sporocysts. Neither species' hemocytes produced ROS in the presence of dead/fixed sporocysts, and there was no suppression of Zymosan-induced respiratory burst by sporocysts. This suggests that immune escape is mediated by avoiding encapsulation, perhaps through molecular mimicry. Membrane-shaving with proteases indicated that sporocyst surface proteins are not a key factors in hemocytic detection. Surface carbohydrates of SRO and cockle hemocytes and of sporocysts were profiled with a panel of biotinylated lectins. This revealed substantial differences between cockle and SRO hemocytes, but greater similarity between cockle hemocytes and sporocysts. Results suggest that surface carbohydrates play an integral role in hemocyte immunorecognition and that surface carbohydrate molecular mimicry is a potential strategy for immune evasion in cockles by hemiuroid trematode sporocysts. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

  1. Bacteriophage Mediates Efficient Gene Transfer in Combination with Conventional Transfection Reagents.

    Science.gov (United States)

    Donnelly, Amanda; Yata, Teerapong; Bentayebi, Kaoutar; Suwan, Keittisak; Hajitou, Amin

    2015-12-08

    The development of commercially available transfection reagents for gene transfer applications has revolutionized the field of molecular biology and scientific research. However, the challenge remains in ensuring that they are efficient, safe, reproducible and cost effective. Bacteriophage (phage)-based viral vectors have the potential to be utilized for general gene transfer applications within research and industry. Yet, they require adaptations in order to enable them to efficiently enter cells and overcome mammalian cellular barriers, as they infect bacteria only; furthermore, limited progress has been made at increasing their efficiency. The production of a novel hybrid nanocomplex system consisting of two different nanomaterial systems, phage vectors and conventional transfection reagents, could overcome these limitations. Here we demonstrate that the combination of cationic lipids, cationic polymers or calcium phosphate with M13 bacteriophage-derived vectors, engineered to carry a mammalian transgene cassette, resulted in increased cellular attachment, entry and improved transgene expression in human cells. Moreover, addition of a targeting ligand into the nanocomplex system, through genetic engineering of the phage capsid further increased gene expression and was effective in a stable cell line generation application. Overall, this new hybrid nanocomplex system (i) provides enhanced phage-mediated gene transfer; (ii) is applicable for laboratory transfection processes and (iii) shows promise within industry for large-scale gene transfer applications.

  2. Microbead agglutination based assays

    KAUST Repository

    Kodzius, Rimantas; Castro, David; Foulds, Ian G.; Parameswaran, Ash M.; Sumanpreet, K. Chhina

    2013-01-01

    We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microbeads in the presence of a specific analyte thus enabling

  3. Ricinus communis agglutinin-mediated agglutination and fusion of glycolipid-containing phospholipid vesicles: effect of carbohydrate head group size, calcium ions, and spermine.

    Science.gov (United States)

    Hoekstra, D; Düzgüneş, N

    1986-03-25

    The glycolipids galactosylcerebroside (GalCer), lactosylceramide (LacCer), and trihexosylceramide (Gb3) were inserted into phospholipid vesicles, consisting of phosphatidylethanolamine and phosphatidic acid. The extent to which their carbohydrate head groups protruded beyond the vesicle surface and their interference with membrane approach were examined by determining vesicle susceptibility toward type I Ricinus communis agglutinin (RCA1) induced agglutination and Ca2+- and spermine-induced aggregation and fusion either in the presence or in the absence of the lectin. The initial agglutination rates increased in the order GalCer much less than LacCer less than Gb3, while a reversed order was obtained for Ca2+- and spermine-induced aggregation and fusion, indicating an enhanced steric interference on close approach of bilayers with increasing head group size. The lectin-mediated agglutination rates for LacCer- and Gb3-containing vesicles increased by an order of magnitude when Ca2+ was also included in the medium, at a concentration that did not induce aggregation per se. Charge neutralization could not account for this observation as the polyvalent cation spermine did not display this synergistic effect with RCA1. Addition of Ca2+ to preagglutinated vesicles substantially reduced the threshold cation concentration for fusion (micromolar vs. millimolar). Quantitatively, this concentration decreased with decreasing carbohydrate head group size, indicating that the head group protrusion determined the interbilayer distance within the vesicle aggregate. The distinct behavior of Ca2+ vs. spermine on RCA1-induced agglutination on the one hand and fusion on the other indicated that Ca2+ regulates the steric orientation of the carbohydrate head group, which appears to be related to its ability to dehydrate the bilayer. As a result, lectin agglutinability becomes enhanced while fusion will be interrupted as the interbilayer distance increases, the threshold head group size

  4. Two hemocyte lineages exist in silkworm larval hematopoietic organ.

    Science.gov (United States)

    Nakahara, Yuichi; Kanamori, Yasushi; Kiuchi, Makoto; Kamimura, Manabu

    2010-07-28

    Insects have multiple hemocyte morphotypes with different functions as do vertebrates, however, their hematopoietic lineages are largely unexplored with the exception of Drosophila melanogaster. To study the hematopoietic lineage of the silkworm, Bombyx mori, we investigated in vivo and in vitro differentiation of hemocyte precursors in the hematopoietic organ (HPO) into the four mature hemocyte subsets, namely, plasmatocytes, granulocytes, oenocytoids, and spherulocytes. Five days after implantation of enzymatically-dispersed HPO cells from a GFP-expressing transgenic line into the hemocoel of normal larvae, differentiation into plasmatocytes, granulocytes and oenocytoids, but not spherulocytes, was observed. When the HPO cells were cultured in vitro, plasmatocytes appeared rapidly, and oenocytoids possessing prophenol oxidase activity appeared several days later. HPO cells were also able to differentiate into a small number of granulocytes, but not into spherulocytes. When functionally mature plasmatocytes were cultured in vitro, oenocytoids were observed 10 days later. These results suggest that the hemocyte precursors in HPO first differentiate into plasmatocytes, which further change into oenocytoids. From these results, we propose that B. mori hemocytes can be divided into two major lineages, a granulocyte lineage and a plasmatocyte-oenocytoid lineage. The origins of the spherulocytes could not be determined in this study. We construct a model for the hematopoietic lineages at the larval stage of B. mori.

  5. Radiation-induced alterations in binding of concanavalin A to cells and in their susceptibility to agglutination

    International Nuclear Information System (INIS)

    Takahashi, Kazuhide; Kaneko, Ichiro

    1986-01-01

    Cell susceptibility to agglutination mediated by a plant lectin, concanavalin A (Con A), and the binding capacity of Con A to cells following γ-irradiation have been examined in mouse myeloid leukaemia cells cultured in suspension. Irradiation caused an immediate decrease in the amount of Con A bound to the cell surface, whereas susceptibility of irradiated cells to agglutination by Con A was unchanged when compared to that of the unirradiated cells. Post-irradiation incubation of cells at 37 0 resulted in a temporary, more than 1.3-fold increase in cell susceptibility to agglutination 60 min after irradiation, whereas binding capacity of cells for Con A gradually-recovered following irradiation, reaching a comparable level to that of unirradiated cells 3 h after irradiation. Cell susceptibility to agglutination by Con A does not depend strongly on its binding capacity. (author)

  6. Two hemocyte lineages exist in silkworm larval hematopoietic organ.

    Directory of Open Access Journals (Sweden)

    Yuichi Nakahara

    Full Text Available BACKGROUND: Insects have multiple hemocyte morphotypes with different functions as do vertebrates, however, their hematopoietic lineages are largely unexplored with the exception of Drosophila melanogaster. METHODOLOGY/PRINCIPAL FINDINGS: To study the hematopoietic lineage of the silkworm, Bombyx mori, we investigated in vivo and in vitro differentiation of hemocyte precursors in the hematopoietic organ (HPO into the four mature hemocyte subsets, namely, plasmatocytes, granulocytes, oenocytoids, and spherulocytes. Five days after implantation of enzymatically-dispersed HPO cells from a GFP-expressing transgenic line into the hemocoel of normal larvae, differentiation into plasmatocytes, granulocytes and oenocytoids, but not spherulocytes, was observed. When the HPO cells were cultured in vitro, plasmatocytes appeared rapidly, and oenocytoids possessing prophenol oxidase activity appeared several days later. HPO cells were also able to differentiate into a small number of granulocytes, but not into spherulocytes. When functionally mature plasmatocytes were cultured in vitro, oenocytoids were observed 10 days later. These results suggest that the hemocyte precursors in HPO first differentiate into plasmatocytes, which further change into oenocytoids. CONCLUSIONS/SIGNIFICANCE: From these results, we propose that B. mori hemocytes can be divided into two major lineages, a granulocyte lineage and a plasmatocyte-oenocytoid lineage. The origins of the spherulocytes could not be determined in this study. We construct a model for the hematopoietic lineages at the larval stage of B. mori.

  7. In vitro cultivation and cryopreservation of Babesia bigemina sporokinetes in hemocytes of Rhipicephalus microplus.

    Science.gov (United States)

    de Rezende, Jania; Rangel, Charles P; McIntosh, Douglas; Silveira, Júlia A G; Cunha, Nathalie C; Ramos, Carlos A N; Fonseca, Adivaldo H

    2015-09-15

    Cultures of tick hemocytes represent alternative cell lines for the isolation and cultivation of a variety of hemoparasites. The present study reports the development and evaluation of methods for the in vitro culture and maintenance of sporokinetes of Babesia bigemina in association with hemocytes of the tick Rhipicephalus microplus. Hemolymph, from engorged females infected with B. bigemina sporokinetes, was incubated at 28 °C in L15 culture medium supplemented with 40% fetal bovine serum. Adherence of hemocytes to flask surfaces and the development of B. bigemina sporokinetes commenced on the first day of cultivation. The protozoa demonstrated clear motility and the capacity to adhere to hemocyte membranes for up to 25 days, at which time the hemocytes began to show signs of degeneration. Examination of Giemsa stained hemocyte cultures, revealed the presence of pyriformis forms, as well as mature and immature sporokinetes with dark red nuclei, centralized or near the apical extremities. Sporokinetes harvested from culture supernatants were cryopreserved in liquid nitrogen. Inoculation of parasite-free hemocyte cultures with defrosted sporokinetes, demonstrated the viability and interaction of the protozoa with the hemocytes over 21 days. Cultured hemocytes of R. microplus hold potential for development as a tool in the study of host parasite interactions and as a substrate for the in vitro maintenance of B. bigemina sporokinetes. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. Good agreement of conventional and gel-based direct agglutination test in immune-mediated haemolytic anaemia

    Directory of Open Access Journals (Sweden)

    Piek Christine J

    2012-02-01

    Full Text Available Abstract Background The aim of this study was to compare a gel-based test with the traditional direct agglutination test (DAT for the diagnosis of immune-mediated haemolytic anaemia (IMHA. Methods Canine (n = 247 and feline (n = 74 blood samples were submitted for DAT testing to two laboratories. A subset of canine samples was categorized as having idiopathic IMHA, secondary IMHA, or no IMHA. Results The kappa values for agreement between the tests were in one laboratory 0.86 for canine and 0.58 for feline samples, and in the other 0.48 for canine samples. The lower agreement in the second laboratory was caused by a high number of positive canine DATs for which the gel test was negative. This group included significantly more dogs with secondary IMHA. Conclusions The gel test might be used as a screening test for idiopathic IMHA and is less often positive in secondary IMHA than the DAT.

  9. DNA & Protein detection based on microbead agglutination

    KAUST Repository

    Kodzius, Rimantas

    2012-06-06

    We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microparticles in the presence of a specific analyte thus enabling the macroscopic observation. Agglutination-based tests are most often used to explore the antibody-antigen reactions. Agglutination has been used for mode protein assays using a biotin/streptavidin two-component system, as well as a hybridization based two-component assay; however, as our work shows, two-component systems are prone to self-termination of the linking analyte and thus have a lower sensitivity. Three component systems have also been used with DNA hybridization, as in our work; however, their assay requires 48 hours for incubation, while our assay is performed in 5 minutes making it a real candidate for POC testing. We demonstrate three assays: a two-component biotin/streptavidin assay, a three-component hybridization assay using single stranded DNA (ssDNA) molecules and a stepped three-component hybridization assay. The comparison of these three assays shows our simple stepped three-component agglutination assay to be rapid at room temperature and more sensitive than the two-component version by an order of magnitude. An agglutination assay was also performed in a PDMS microfluidic chip where agglutinated beads were trapped by filter columns for easy observation. We developed a rapid (5 minute) room temperature assay, which is based on microbead agglutination. Our three-component assay solves the linker self-termination issue allowing an order of magnitude increase in sensitivity over two–component assays. Our stepped version of the three-component assay solves the issue with probe site saturation thus enabling a wider range of detection. Detection of the agglutinated beads with the naked eye by trapping in microfluidic channels has been shown.

  10. Ocean Acidification Affects Hemocyte Physiology in the Tanner Crab (Chionoecetes bairdi)

    Science.gov (United States)

    Meseck, Shannon L.; Alix, Jennifer H.; Swiney, Katherine M.; Long, W. Christopher; Wikfors, Gary H.; Foy, Robert J.

    2016-01-01

    We used flow cytometry to determine if there would be a difference in hematology, selected immune functions, and hemocyte pH (pHi), under two different, future ocean acidification scenarios (pH = 7.50, 7.80) compared to current conditions (pH = 8.09) for Chionoecetes bairdi, Tanner crab. Hemocytes were analyzed after adult Tanner crabs were held for two years under continuous exposure to acidified ocean water. Total counts of hemocytes did not vary among control and experimental treatments; however, there were significantly greater number of dead, circulating hemocytes in crabs held at the lowest pH treatment. Phagocytosis of fluorescent microbeads by hemocytes was greatest at the lowest pH treatment. These results suggest that hemocytes were dying, likely by apoptosis, at a rate faster than upregulated phagocytosis was able to remove moribund cells from circulation at the lowest pH. Crab hemolymph pH (pHe) averaged 8.09 and did not vary among pH treatments. There was no significant difference in internal pH (pHi) within hyalinocytes among pH treatments and the mean pHi (7.26) was lower than the mean pHe. In contrast, there were significant differences among treatments in pHi of the semi-granular+granular cells. Control crabs had the highest mean semi-granular+granular pHi compared to the lowest pH treatment. As physiological hemocyte functions changed from ambient conditions, interactions with the number of eggs in the second clutch, percentage of viable eggs, and calcium concentration in the adult crab shell was observed. This suggested that the energetic costs of responding to ocean acidification and maintaining defense mechanisms in Tanner crab may divert energy from other physiological processes, such as reproduction. PMID:26859148

  11. RNA-Seq Study of Microbially Induced Hemocyte Transcripts from Larval Heliothis virescens (Lepidoptera: Noctuidae

    Directory of Open Access Journals (Sweden)

    Kent S. Shelby

    2012-08-01

    Full Text Available Larvae of the tobacco budworm are major polyphagous pests throughout the Americas. Development of effective microbial biopesticides for this and related noctuid pests has been stymied by the natural resistance mediated innate immune response. Hemocytes play an early and central role in activating and coordinating immune responses to entomopathogens. To approach this problem we completed RNA-seq expression profiling of hemocytes collected from larvae following an in vivo challenge with bacterial and fungal cell wall components to elicit an immune response. A de novo exome assembly was constructed by combination of sequence tags from all treatments. Sequence tags from each treatment were aligned separately with the assembly to measure expression. The resulting table of differential expression had > 22,000 assemblies each with a distinct combination of annotation and expression. Within these assemblies > 1,400 were upregulated and > 1,500 downregulated by immune activation with bacteria or fungi. Orthologs to innate immune components of other insects were identified including pattern recognition, signal transduction pathways, antimicrobial peptides and enzymes, melanization and coagulation. Additionally orthologs of components regulating hemocytic functions such as autophagy, apoptosis, phagocytosis and nodulation were identified. Associated cellular oxidative defenses and detoxification responses were identified providing a comprehensive snapshot of the early response to elicitation.

  12. Reactive oxygen species in unstimulated hemocytes of the pacific oyster Crassostrea gigas: a mitochondrial involvement.

    Directory of Open Access Journals (Sweden)

    Ludovic Donaghy

    Full Text Available The Pacific oyster Crassostrea gigas is a sessile bivalve mollusc whose homeostasis relies, at least partially, upon cells circulating in hemolymph and referred to as hemocytes. Oyster's hemocytes have been reported to produce reactive oxygen species (ROS, even in absence of stimulation. Although ROS production in bivalve molluscs is mostly studied for its defence involvement, ROS may also be involved in cellular and tissue homeostasis. ROS sources have not yet been described in oyster hemocytes. The objective of the present work was to characterize the ROS sources in unstimulated hemocytes. We studied the effects of chemical inhibitors on the ROS production and the mitochondrial membrane potential (Δψ(m of hemocytes. First, this work confirmed the specificity of JC-10 probe to measure Δψ(m in oyster hemocytes, without being affected by ΔpH, as reported in mammalian cells. Second, results show that ROS production in unstimulated hemocytes does not originate from cytoplasmic NADPH-oxidase, nitric oxide synthase or myeloperoxidase, but from mitochondria. In contrast to mammalian cells, incubation of hemocytes with rotenone (complex I inhibitor had no effect on ROS production. Incubation with antimycin A (complex III inhibitor resulted in a dose-dependent ROS production decrease while an over-production is usually reported in vertebrates. In hemocytes of C. gigas, the production of ROS seems similarly dependent on both Δψ(m and ΔpH. These findings point out differences between mammalian models and bivalve cells, which warrant further investigation about the fine characterization of the electron transfer chain and the respective involvement of mitochondrial complexes in ROS production in hemocytes of bivalve molluscs.

  13. Honey bee hemocyte profiling by flow cytometry.

    Science.gov (United States)

    Marringa, William J; Krueger, Michael J; Burritt, Nancy L; Burritt, James B

    2014-01-01

    Multiple stress factors in honey bees are causing loss of bee colonies worldwide. Several infectious agents of bees are believed to contribute to this problem. The mechanisms of honey bee immunity are not completely understood, in part due to limited information about the types and abundances of hemocytes that help bees resist disease. Our study utilized flow cytometry and microscopy to examine populations of hemolymph particulates in honey bees. We found bee hemolymph includes permeabilized cells, plasmatocytes, and acellular objects that resemble microparticles, listed in order of increasing abundance. The permeabilized cells and plasmatocytes showed unexpected differences with respect to properties of the plasma membrane and labeling with annexin V. Both permeabilized cells and plasmatocytes failed to show measurable mitochondrial membrane potential by flow cytometry using the JC-1 probe. Our results suggest hemolymph particulate populations are dynamic, revealing significant differences when comparing individual hive members, and when comparing colonies exposed to diverse conditions. Shifts in hemocyte populations in bees likely represent changing conditions or metabolic differences of colony members. A better understanding of hemocyte profiles may provide insight into physiological responses of honey bees to stress factors, some of which may be related to colony failure.

  14. High Fidelity, High Volume Agglutinate Manufacturing Process, Phase I

    Data.gov (United States)

    National Aeronautics and Space Administration — Up to 65% of the lunar soils are comprised of agglutinates. Although the importance of agglutinate in simulants is often debated, the fact is that agglutinates...

  15. Propagating the missing bacteriophages: a large bacteriophage in a new class

    Directory of Open Access Journals (Sweden)

    Hardies Stephen C

    2007-02-01

    Full Text Available Abstract The number of successful propagations/isolations of soil-borne bacteriophages is small in comparison to the number of bacteriophages observed by microscopy (great plaque count anomaly. As one resolution of the great plaque count anomaly, we use propagation in ultra-dilute agarose gels to isolate a Bacillus thuringiensis bacteriophage with a large head (95 nm in diameter, tail (486 × 26 nm, corkscrew-like tail fibers (187 × 10 nm and genome (221 Kb that cannot be detected by the usual procedures of microbiology. This new bacteriophage, called 0305φ8-36 (first number is month/year of isolation; remaining two numbers identify the host and bacteriophage, has a high dependence of plaque size on the concentration of a supporting agarose gel. Bacteriophage 0305φ8-36 does not propagate in the traditional gels used for bacteriophage plaque formation and also does not produce visible lysis of liquid cultures. Bacteriophage 0305φ8-36 aggregates and, during de novo isolation from the environment, is likely to be invisible to procedures of physical detection that use either filtration or centrifugal pelleting to remove bacteria. Bacteriophage 0305φ8-36 is in a new genomic class, based on genes for both structural components and DNA packaging ATPase. Thus, knowledge of environmental virus diversity is expanded with prospect of greater future expansion.

  16. Towards a high throughput droplet-based agglutination assay

    KAUST Repository

    Kodzius, Rimantas; Castro, David; Foulds, Ian G.

    2013-01-01

    This work demonstrates the detection method for a high throughput droplet based agglutination assay system. Using simple hydrodynamic forces to mix and aggregate functionalized microbeads we avoid the need to use magnetic assistance or mixing structures. The concentration of our target molecules was estimated by agglutination strength, obtained through optical image analysis. Agglutination in droplets was performed with flow rates of 150 µl/min and occurred in under a minute, with potential to perform high-throughput measurements. The lowest target concentration detected in droplet microfluidics was 0.17 nM, which is three orders of magnitude more sensitive than a conventional card based agglutination assay.

  17. Towards a high throughput droplet-based agglutination assay

    KAUST Repository

    Kodzius, Rimantas

    2013-10-22

    This work demonstrates the detection method for a high throughput droplet based agglutination assay system. Using simple hydrodynamic forces to mix and aggregate functionalized microbeads we avoid the need to use magnetic assistance or mixing structures. The concentration of our target molecules was estimated by agglutination strength, obtained through optical image analysis. Agglutination in droplets was performed with flow rates of 150 µl/min and occurred in under a minute, with potential to perform high-throughput measurements. The lowest target concentration detected in droplet microfluidics was 0.17 nM, which is three orders of magnitude more sensitive than a conventional card based agglutination assay.

  18. Recognition between symbiotic Vibrio fischeri and the hemocytes of Euprymna scolopes

    Science.gov (United States)

    Nyholm, Spencer V.; Stewart, Jennifer J.; Ruby, Edward G.; McFall-Ngai, Margaret J.

    2008-01-01

    Summary The light-organ crypts of the squid Euprymna scolopes permit colonization exclusively by the luminous bacterium Vibrio fischeri. Because the crypt interior remains in contact with seawater, the squid must not only foster the specific symbiosis but also continue to exclude other bacteria. Investigation of the role of the innate immune system in these processes revealed that macrophage-like hemocytes isolated from E. scolopes recognized and phagocytosed V. fischeri less than other closely related bacterial species common to the host’s environment. Interestingly, phagocytes isolated from hosts that had been cured of their symbionts bound five-times more V. fischeri cells than those from uncured hosts. No such change in the ability to bind other species of bacteria was observed, suggesting that the host adapts specifically to V. fischeri. Deletion of the gene encoding OmpU, the major outer membrane protein of V. fischeri, increased binding by hemocytes from uncured animals to the level observed for hemocytes from cured animals. Co-incubation with wild-type V. fischeri reduced this binding, suggesting they produce a factor that complements the mutant’s defect. Analyses of the phagocytosis of bound cells by fluorescence-activated cell sorting (FACS) indicated that, once binding to hemocytes had occurred, V. fischeri cells are phagocytosed as effectively as other bacteria. Thus, discrimination by this component of the squid immune system occurs at the level of hemocyte binding, and this response: (i) is modified by previous exposure to the symbiont and, (ii) relies on outer membrane and/or secreted components of the symbionts. These data suggest that regulation of host hemocyte binding by the symbiont may be one of many factors that contribute to specificity in this association. PMID:19196278

  19. DNA & Protein detection based on microbead agglutination

    KAUST Repository

    Kodzius, Rimantas; Castro, David; Foulds, Ian G.; Parameswaran, Ash M.; Sumanpreet, K. Chhina

    2012-01-01

    the macroscopic observation. Agglutination-based tests are most often used to explore the antibody-antigen reactions. Agglutination has been used for mode protein assays using a biotin/streptavidin two-component system, as well as a hybridization based two

  20. Exploring new biological functions of amyloids: bacteria cell agglutination mediated by host protein aggregation.

    Directory of Open Access Journals (Sweden)

    Marc Torrent

    Full Text Available Antimicrobial proteins and peptides (AMPs are important effectors of the innate immune system that play a vital role in the prevention of infections. Recent advances have highlighted the similarity between AMPs and amyloid proteins. Using the Eosinophil Cationic Protein as a model, we have rationalized the structure-activity relationships between amyloid aggregation and antimicrobial activity. Our results show how protein aggregation can induce bacteria agglutination and cell death. Using confocal and total internal reflection fluorescence microscopy we have tracked the formation in situ of protein amyloid-like aggregates at the bacteria surface and on membrane models. In both cases, fibrillar aggregates able to bind to amyloid diagnostic dyes were detected. Additionally, a single point mutation (Ile13 to Ala can suppress the protein amyloid behavior, abolishing the agglutinating activity and impairing the antimicrobial action. The mutant is also defective in triggering both leakage and lipid vesicle aggregation. We conclude that ECP aggregation at the bacterial surface is essential for its cytotoxicity. Hence, we propose here a new prospective biological function for amyloid-like aggregates with potential biological relevance.

  1. Bacteriophages and Biofilms

    Directory of Open Access Journals (Sweden)

    David R. Harper

    2014-06-01

    Full Text Available Biofilms are an extremely common adaptation, allowing bacteria to colonize hostile environments. They present unique problems for antibiotics and biocides, both due to the nature of the extracellular matrix and to the presence within the biofilm of metabolically inactive persister cells. Such chemicals can be highly effective against planktonic bacterial cells, while being essentially ineffective against biofilms. By contrast, bacteriophages seem to have a greater ability to target this common form of bacterial growth. The high numbers of bacteria present within biofilms actually facilitate the action of bacteriophages by allowing rapid and efficient infection of the host and consequent amplification of the bacteriophage. Bacteriophages also have a number of properties that make biofilms susceptible to their action. They are known to produce (or to be able to induce enzymes that degrade the extracellular matrix. They are also able to infect persister cells, remaining dormant within them, but re-activating when they become metabolically active. Some cultured biofilms also seem better able to support the replication of bacteriophages than comparable planktonic systems. It is perhaps unsurprising that bacteriophages, as the natural predators of bacteria, have the ability to target this common form of bacterial life.

  2. New insights from the oyster Crassostrea rhizophorae on bivalve circulating hemocytes.

    Directory of Open Access Journals (Sweden)

    Mauro de Freitas Rebelo

    Full Text Available Hemocytes are the first line of defense of the immune system in invertebrates, but despite their important role and enormous potential for the study of gene-environment relationships, research has been impeded by a lack of consensus on their classification. Here we used flow cytometry combined with histological procedures, histochemical reactions and transmission electron microscopy to characterize the hemocytes from the oyster Crassostrea rhizophorae. Transmission electron microscopy revealed remarkable morphological characteristics, such as the presence of membranous cisternae in all mature cells, regardless of size and granulation. Some granular cells contained many cytoplasmic granules that communicated with each other through a network of channels, a feature never previously described for hemocytes. The positive reactions for esterase and acid phosphatase also indicated the presence of mature cells of all sizes and granule contents. Flow cytometry revealed a clear separation in complexity between agranular and granular populations, which could not be differentiated by size, with cells ranging from 2.5 to 25 µm. Based on this evidence we suggest that, at least in C. rhizophorae, the different subpopulations of hemocytes may in reality be different stages of one type of cell, which accumulates granules and loses complexity (with no reduction in size as it degranulates in the event of an environmental challenge.

  3. Nano/Micro Formulations for Bacteriophage Delivery.

    Science.gov (United States)

    Cortés, Pilar; Cano-Sarabia, Mary; Colom, Joan; Otero, Jennifer; Maspoch, Daniel; Llagostera, Montserrat

    2018-01-01

    Encapsulation methodologies allow the protection of bacteriophages for overcoming critical environmental conditions. Moreover, they improve the stability and the controlled delivery of bacteriophages which is of great innovative value in bacteriophage therapy. Here, two different encapsulation methodologies of bacteriophages are described using two biocompatible materials: a lipid cationic mixture and a combination of alginate with the antacid CaCO 3 . To perform bacteriophage encapsulation, a purified lysate highly concentrated (around 10 10 -10 11  pfu/mL) is necessary, and to dispose of a specific equipment. Both methodologies have been successfully applied for encapsulating Salmonella bacteriophages with different morphologies. Also, the material employed does not modify the antibacterial action of bacteriophages. Moreover, both technologies can also be adapted to any bacteriophage and possibly to any delivery route for bacteriophage therapy.

  4. Multiple roles of genome-attached bacteriophage terminal proteins

    International Nuclear Information System (INIS)

    Redrejo-Rodríguez, Modesto; Salas, Margarita

    2014-01-01

    Protein-primed replication constitutes a generalized mechanism to initiate DNA or RNA synthesis in linear genomes, including viruses, gram-positive bacteria, linear plasmids and mobile elements. By this mechanism a specific amino acid primes replication and becomes covalently linked to the genome ends. Despite the fact that TPs lack sequence homology, they share a similar structural arrangement, with the priming residue in the C-terminal half of the protein and an accumulation of positively charged residues at the N-terminal end. In addition, various bacteriophage TPs have been shown to have DNA-binding capacity that targets TPs and their attached genomes to the host nucleoid. Furthermore, a number of bacteriophage TPs from different viral families and with diverse hosts also contain putative nuclear localization signals and localize in the eukaryotic nucleus, which could lead to the transport of the attached DNA. This suggests a possible role of bacteriophage TPs in prokaryote-to-eukaryote horizontal gene transfer. - Highlights: • Protein-primed genome replication constitutes a strategy to initiate DNA or RNA synthesis in linear genomes. • Bacteriophage terminal proteins (TPs) are covalently attached to viral genomes by their primary function priming DNA replication. • TPs are also DNA-binding proteins and target phage genomes to the host nucleoid. • TPs can also localize in the eukaryotic nucleus and may have a role in phage-mediated interkingdom gene transfer

  5. Multiple roles of genome-attached bacteriophage terminal proteins

    Energy Technology Data Exchange (ETDEWEB)

    Redrejo-Rodríguez, Modesto; Salas, Margarita, E-mail: msalas@cbm.csic.es

    2014-11-15

    Protein-primed replication constitutes a generalized mechanism to initiate DNA or RNA synthesis in linear genomes, including viruses, gram-positive bacteria, linear plasmids and mobile elements. By this mechanism a specific amino acid primes replication and becomes covalently linked to the genome ends. Despite the fact that TPs lack sequence homology, they share a similar structural arrangement, with the priming residue in the C-terminal half of the protein and an accumulation of positively charged residues at the N-terminal end. In addition, various bacteriophage TPs have been shown to have DNA-binding capacity that targets TPs and their attached genomes to the host nucleoid. Furthermore, a number of bacteriophage TPs from different viral families and with diverse hosts also contain putative nuclear localization signals and localize in the eukaryotic nucleus, which could lead to the transport of the attached DNA. This suggests a possible role of bacteriophage TPs in prokaryote-to-eukaryote horizontal gene transfer. - Highlights: • Protein-primed genome replication constitutes a strategy to initiate DNA or RNA synthesis in linear genomes. • Bacteriophage terminal proteins (TPs) are covalently attached to viral genomes by their primary function priming DNA replication. • TPs are also DNA-binding proteins and target phage genomes to the host nucleoid. • TPs can also localize in the eukaryotic nucleus and may have a role in phage-mediated interkingdom gene transfer.

  6. Oxidative stress parameters induced by exposure to either cadmium or 17β-estradiol on Mytilus galloprovincialis hemocytes. The role of signaling molecules

    Energy Technology Data Exchange (ETDEWEB)

    Koutsogiannaki, Sophia [Laboratory of Animal Physiology, Zoology Department, School of Biology, Faculty of Science, Aristotle University of Thessaloniki, 54124 Thessaloniki (Greece); Franzellitti, Silvia [University of Bologna, Interdepartment Centre for Environmental Science Research, via S. Alberto 163, 48123 Ravenna (Italy); Fabbri, Elena [University of Bologna, Interdepartment Centre for Environmental Science Research, via S. Alberto 163, 48123 Ravenna (Italy); University of Bologna, Department of Biological, Geological, and Environmental Sciences, via Selmi 3, 40100 Bologna (Italy); Kaloyianni, Martha, E-mail: kaloyian@bio.auth.gr [Laboratory of Animal Physiology, Zoology Department, School of Biology, Faculty of Science, Aristotle University of Thessaloniki, 54124 Thessaloniki (Greece)

    2014-01-15

    Highlights: •Oxidative parameters in Mytilus galloprovincialis hemocytes were measured. •Comparison between cadmium and 17β-estradiol cytotoxicity is discussed. •NHE, PKC, PI3-K, NADPH oxidase, NO synthase, JNK involvement was observed. •Protective role of cAMP is suggested. •Signaling molecules studied could constitute novel biomarkers. -- Abstract: The aim of the present study was to determine and compare the possible effects of exposure to an estrogen, 17β-estradiol and to a metal, cadmium on oxidative parameters of Mytilus galloprovincialis hemocytes and to elucidate the signaling pathways that probably mediate the studied effects exerted by these two chemicals. In addition, it was of interest to investigate if the studied parameters could constitute biomarkers for aquatic pollution monitoring. Our results suggest that micromolar concentrations of either cadmium or 17β-estradiol affected the redox status of mussels by modulating oxidative parameters and antioxidant enzymes gene expression in mussel M. galloprovincialis hemocytes. In particular, our results showed that treatment of hemocytes with either 5 μM of cadmium chloride or with 25 nM of 17β-estradiol for 30 min caused significant increased ROS production; this led to oxidative damage exemplified by significant increased DNA damage, protein carbonylation and lipid peroxidation, as well as increased mRNA levels of the antioxidant enzymes catalase (CAT), superoxide dismoutase (SOD) and glutathione S-transferase (GST). Furthermore, our results suggest that either cadmium or 17β-estradiol signal is mediated either through one of the already known pathways initiated by photatidyl-inositol 3-kinase (PI3 K) and reaching Na{sup +}/H{sup +} exchanger (NHE) probably through protein kinase C (PKC) or a kinase-mediated signaling pathway that involves in most of the cases NHE, PKC, Ca{sup 2+}-dependent PKC isoforms, PI3-K, NADPH oxidase, nitric oxide (NO) synthase, c-Jun N-terminal kinase (JNK) and

  7. Oxidative stress parameters induced by exposure to either cadmium or 17β-estradiol on Mytilus galloprovincialis hemocytes. The role of signaling molecules

    International Nuclear Information System (INIS)

    Koutsogiannaki, Sophia; Franzellitti, Silvia; Fabbri, Elena; Kaloyianni, Martha

    2014-01-01

    Highlights: •Oxidative parameters in Mytilus galloprovincialis hemocytes were measured. •Comparison between cadmium and 17β-estradiol cytotoxicity is discussed. •NHE, PKC, PI3-K, NADPH oxidase, NO synthase, JNK involvement was observed. •Protective role of cAMP is suggested. •Signaling molecules studied could constitute novel biomarkers. -- Abstract: The aim of the present study was to determine and compare the possible effects of exposure to an estrogen, 17β-estradiol and to a metal, cadmium on oxidative parameters of Mytilus galloprovincialis hemocytes and to elucidate the signaling pathways that probably mediate the studied effects exerted by these two chemicals. In addition, it was of interest to investigate if the studied parameters could constitute biomarkers for aquatic pollution monitoring. Our results suggest that micromolar concentrations of either cadmium or 17β-estradiol affected the redox status of mussels by modulating oxidative parameters and antioxidant enzymes gene expression in mussel M. galloprovincialis hemocytes. In particular, our results showed that treatment of hemocytes with either 5 μM of cadmium chloride or with 25 nM of 17β-estradiol for 30 min caused significant increased ROS production; this led to oxidative damage exemplified by significant increased DNA damage, protein carbonylation and lipid peroxidation, as well as increased mRNA levels of the antioxidant enzymes catalase (CAT), superoxide dismoutase (SOD) and glutathione S-transferase (GST). Furthermore, our results suggest that either cadmium or 17β-estradiol signal is mediated either through one of the already known pathways initiated by photatidyl-inositol 3-kinase (PI3 K) and reaching Na + /H + exchanger (NHE) probably through protein kinase C (PKC) or a kinase-mediated signaling pathway that involves in most of the cases NHE, PKC, Ca 2+ -dependent PKC isoforms, PI3-K, NADPH oxidase, nitric oxide (NO) synthase, c-Jun N-terminal kinase (JNK) and cyclic adenosine

  8. Concholepas hemocyanin biosynthesis takes place in the hepatopancreas, with hemocytes being involved in its metabolism.

    Science.gov (United States)

    Manubens, Augusto; Salazar, Fabián; Haussmann, Denise; Figueroa, Jaime; Del Campo, Miguel; Pinto, Jonathan Martínez; Huaquín, Laura; Venegas, Alejandro; Becker, María Inés

    2010-12-01

    Hemocyanins are copper-containing glycoproteins in some molluscs and arthropods, and their best-known function is O(2) transport. We studied the site of their biosynthesis in the gastropod Concholepas concholepas by using immunological and molecular genetic approaches. We performed immunohistochemical staining of various organs, including the mantle, branchia, and hepatopancreas, and detected C. concholepas hemocyanin (CCH) molecules in circulating and tissue-associated hemocytes by electron microscopy. To characterize the hemocytes, we purified them from hemolymph. We identified three types of granular cells. The most abundant type was a phagocyte-like cell with small cytoplasmic granules. The second type contained large electron-dense granules. The third type had vacuoles containing hemocyanin molecules suggesting that synthesis or catabolism occurred inside these cells. Our failure to detect cch-mRNA in hemocytes by reverse transcription with the polymerase chain reaction (RT-PCR) led us to propose that hemocytes instead played a role in CCH metabolism. This hypothesis was supported by colloidal gold staining showing hemocyanin molecules in electron-dense granules inside hemocytes. RT-PCR analysis, complemented by in situ hybridization analyses with single-stranded antisense RNAs as specific probes, demonstrated the presence of cch-mRNA in the hepatopancreas; this was consistent with the specific hybridization signal and confirmed the hepatopancreas as the site of CCH synthesis. Finally, we investigated the possibility that CCH catabolism in hemocytes was involved in the host immune response and in the generation of secondary metabolites such as antimicrobial peptides and phenoloxidase.

  9. Cytotoxicity and cellular mechanisms involved in the toxicity of CdS quantum dots in hemocytes and gill cells of the mussel Mytilus galloprovincialis

    Energy Technology Data Exchange (ETDEWEB)

    Katsumiti, A. [CBET Research Group, Dept. Zoology and Animal Cell Biology, Faculty of Science and Technology and Research Centre for Experimental Marine Biology and Biotechnology PIE, University of the Basque Country UPV/EHU, Basque Country (Spain); Gilliland, D. [EU Commission–Joint Research Centre, Institute of Health and Consumer Protection, NSB Unit, Ispra (Italy); Arostegui, I. [Department of Applied Mathematics, Statistics and Operations Research, Faculty of Science and Technology, University of the Basque Country UPV/EHU, Leioa (Spain); Cajaraville, M.P., E-mail: mirenp.cajaraville@ehu.es [CBET Research Group, Dept. Zoology and Animal Cell Biology, Faculty of Science and Technology and Research Centre for Experimental Marine Biology and Biotechnology PIE, University of the Basque Country UPV/EHU, Basque Country (Spain)

    2014-08-15

    phagocytosis were found in hemocytes exposed to bulk CdS and to CdS QDs at concentrations equal or higher than 1.25 mg Cd/L but not in those exposed to ionic Cd, indicating a particle-specific effect on phagocytosis. In conclusion, cell-mediated immunity and gill cell function represent significant targets for CdS QDs toxicity.

  10. Toward modern inhalational bacteriophage therapy: nebulization of bacteriophages of Burkholderia cepacia complex.

    Science.gov (United States)

    Golshahi, Laleh; Seed, Kimberley D; Dennis, Jonathan J; Finlay, Warren H

    2008-12-01

    Antibiotic-resistant bacterial infections have renewed interest in finding substitute methods of treatment. The purpose of the present in vitro study was to investigate the possibility of respiratory delivery of a Burkholderia cepacia complex (BCC) bacteriophage by nebulized aerosol administration. Bacteriophages in isotonic saline were aerosolized with Pari LC star and eFlow nebulizers, at titers with mean value (standard deviation) of 2.15 x 10(8) (1.63 x 10(8)) plaque-forming unit (PFU)/mL in 2.5-mL nebulizer fills. The breathing pattern of an adult was simulated using a pulmonary waveform generator. During breath simulation, the size distributions of the nebulized aerosol were measured using phase doppler anemometry (PDA). Efficiency of nebulizer delivery was subsequently determined by collection of aerosol on low resistance filters and measurement of bacteriophage titers. These filter titers were used as input data to a mathematical lung deposition model to predict regional deposition of bacteriophages in the lung and initial bacteriophage titers in the liquid surface layer of each conducting airway generation. The results suggest that BCC bacteriophages can be nebulized successfully within a reasonable delivery time and predicted titers in the lung indicate that this method may hold potential for treatment of bacterial lung infections common among cystic fibrosis patients.

  11. Endogenous growth factor stimulation of hemocyte proliferation induces resistance to Schistosoma mansoni challenge in the snail host.

    Science.gov (United States)

    Pila, Emmanuel A; Gordy, Michelle A; Phillips, Valerie K; Kabore, Alethe L; Rudko, Sydney P; Hanington, Patrick C

    2016-05-10

    Digenean trematodes are a large, complex group of parasitic flatworms that infect an incredible diversity of organisms, including humans. Larval development of most digeneans takes place within a snail (Gastropoda). Compatibility between snails and digeneans is often very specific, such that suitable snail hosts define the geographical ranges of diseases caused by these worms. The immune cells (hemocytes) of a snail are sentinels that act as a crucial barrier to infection by larval digeneans. Hemocytes coordinate a robust and specific immunological response, participating directly in parasite killing by encapsulating and clearing the infection. Hemocyte proliferation and differentiation are influenced by unknown digenean-specific exogenous factors. However, we know nothing about the endogenous control of hemocyte development in any gastropod model. Here, we identify and functionally characterize a progranulin [Biomphalaria glabrata granulin (BgGRN)] from the snail B. glabrata, a natural host for the human blood fluke Schistosoma mansoni Granulins are growth factors that drive proliferation of immune cells in organisms, spanning the animal kingdom. We demonstrate that BgGRN induces proliferation of B. glabrata hemocytes, and specifically drives the production of an adherent hemocyte subset that participates centrally in the anti-digenean defense response. Additionally, we demonstrate that susceptible B. glabrata snails can be made resistant to infection with S. mansoni by first inducing hemocyte proliferation with BgGRN. This marks the functional characterization of an endogenous growth factor of a gastropod mollusc, and provides direct evidence of gain of resistance in a snail-digenean infection model using a defined factor to induce snail resistance to infection.

  12. Surgical management of vulvovaginal agglutination due to lichen planus.

    Science.gov (United States)

    Fairchild, Pamela S; Haefner, Hope K

    2016-02-01

    Lichen planus is a rare dermatological disorder that is often associated with painful and disfiguring vulvovaginal effects. At the University of Michigan Center for Vulvar Diseases, we see many women with vulvovaginal lichen planus each year, with marked scarring and vulvovaginal agglutination that precludes vaginal intercourse and causes difficulty with urination. Through our experience, we developed a protocol for the operative management and postoperative care for severe vulvovaginal agglutination. Our objective is to share this protocol with a wider audience so that providers who see patients with these devastating effects of lichen planus can benefit from our experience to better serve this patient population. The figure represents a case of erosive lichen planus with early vaginal agglutination. The video reviews the pathophysiology and presentation of lichen planus. We then present a case of scarring and agglutination in a young woman, including our surgical management and postoperative care recommendations. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Nitric oxide mediates insect cellular immunity via phospholipase A2 activation

    Science.gov (United States)

    After infection or invasion is recognized, biochemical mediators act in signaling insect immune functions. These include biogenic amines, insect cytokines, eicosanoids and nitric oxide (NO). Treating insects or isolated hemocyte populations with different mediators often leads to similar results. Se...

  14. Morphologic, cytometric and functional characterization of the common octopus (Octopus vulgaris) hemocytes.

    Science.gov (United States)

    Castellanos-Martínez, S; Prado-Alvarez, M; Lobo-da-Cunha, A; Azevedo, C; Gestal, C

    2014-05-01

    The hemocytes of Octopus vulgaris were morphologically and functionally characterized. Light and electron microscopy (TEM and SEM), and flow cytometry analyses revealed the existence of two hemocyte populations. Large granulocytes showed U-shaped nucleus, a mean of 11.6 μm±1.2 in diameter with basophilic granules, polysaccharide and lysosomic deposits in the cytoplasm. Small granulocytes measured a mean of 8.1 μm±0.7 in diameter, and have a round nucleus occupying almost the entire cell and few or not granules in the cytoplasm. Flow cytometry analysis showed that large granulocytes are the principal cells that develop phagocytosis of latex beads (rising up to 56%) and ROS after zymosan stimulation. Zymosan induced the highest production of both ROS and NO. This study is the first tread towards understanding the O. vulgaris immune system by applying new tools to provide a most comprehensive morpho-functional study of their hemocytes. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. Hemocyte characterization of Nasutitermes coxipoensis (Holmgren) (Isoptera: Termitidae) workers and hemocyte evaluation after parasitism by Metarhizium anisopliae; Caracterizacao dos hemocitos de operarios de Nasutitermes coxipoensis (Holmgren) (Isoptera: Termitidae) e avaliacao hemocitaria apos parasitismo por Metarhizium anisopliae

    Energy Technology Data Exchange (ETDEWEB)

    Cunha, Franklin M.; Wanderley-Teixeira, Valeria; Albuquerque, Auristela C. [Universidade Federal Rural de Pernambuco (UFRPE), Recife, PE (Brazil). Programa de Pos-Graduacao em Entomologia Agricola], e-mail: ukento@yahoo.com.br; Teixeira, Alvaro A.C. [Universidade Federal Rural de Pernambuco (UFRPE), Recife, PE (Brazil). Dept. de Morfologia e Fisiologia Animal], e-mail: valeria@dmfa.ufrpe.br, e-mail: auritermes@yahoo.com.br; Alves, Luiz C. [Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil). Lab. de Imunopatologia Keizo Asami (LIKA); Lima, Elza A.L.A. [Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil). Dept. de Micologia. Lab. de Controle Biologico

    2009-03-15

    We aimed to characterize the morphology and ultrastructure of hemocytes of Nasutitermes coxipoensis (Holmgren) workers and to quantify the cell types 24h, 48h and 72h after inoculation with Metarhizium anisopliae. Six hemocytes types were identified, plasmatocyte, granulocyte, spherulocyte, prohemocyte, adipohemocyte and eonocytoid Hemocytes did not present any morphological alteration at the several observation periods, but they did have a change in their abundance, as observed for spherulocytes, adipohemocytes and eonocytoids at all intervals, and for plasmatocytes and granulocytes at 48h after host inoculation. We argue on the possible reasons and implications of the observed changes. (author)

  16. Ultrastructural and functional characterization of circulating hemocytes from Galleria mellonella larva: Cell types and their role in the innate immunity.

    Science.gov (United States)

    Wu, Gongqing; Liu, Yi; Ding, Ying; Yi, Yunhong

    2016-08-01

    Galleria mellonella larvae have been widely used as a model to study the virulence of various human pathogens. Hemocytes play important roles in the innate immune response of G. mellonella. In this study, the hemocytes of G. mellonella larvae were analyzed by transmission electron microscope, light microscope, and cytochemistry. The cytological and morphological analyses revealed four types of hemocytes; Plasmatocytes, granular cells, spherule cells and oenocytoids. Differential hemocyte counts showed that under our conditions plasmatocytes and granular cells were the most abundant circulating cell types in the hemolymph. We also investigated the role of different types of hemocytes in the cellular and humoral immune defenses. The in-vivo experiment showed that plasmatocytes, granular cells and oenocytoids phagocytized FITC-labelled Escherichia coli bacteria in larvae of G. mellonella, whereas the granular cells exhibited the strongest phagocytic ability against these microbial cells. After incubation with L-DOPA, plasmatocytes, granular cells and oenocytoids are stained brown, indicating the presence of phenoloxidase activity. These results shed new light on our understanding of the immune function of G. mellonella hemocytes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Synthetic Biology to Engineer Bacteriophage Genomes.

    Science.gov (United States)

    Rita Costa, Ana; Milho, Catarina; Azeredo, Joana; Pires, Diana Priscila

    2018-01-01

    Recent advances in the synthetic biology field have enabled the development of new molecular biology techniques used to build specialized bacteriophages with new functionalities. Bacteriophages have been engineered towards a wide range of applications including pathogen control and detection, targeted drug delivery, or even assembly of new materials.In this chapter, two strategies that have been successfully used to genetically engineer bacteriophage genomes are addressed: a yeast-based platform and bacteriophage recombineering of electroporated DNA.

  18. The virus-like particles of a braconid endoparasitoid wasp, Meteorus pulchricornis, inhibit hemocyte spreading in its noctuid host, Pseudaletia separata.

    Science.gov (United States)

    Suzuki, M; Miura, K; Tanaka, T

    2008-06-01

    We previously reported that the virus-like particles of Meteorus pulchricornis (MpVLPs) are capable of inducing apoptosis by around 6h in the hemocytes of the host, Pseudaletia separata [Suzuki, M., Tanaka, T., 2006. Virus-like particles in venom of Meteorus pulchricornis induce host hemocyte apoptosis. Journal of Insect Physiology 52, 602-611], thereby protecting the oviposited egg. In the present study, we focused on analyses of the earlier events caused by the MpVLPs upon the host immune response, namely their effects on hemocyte spreading. After recognition and attachment on foreign substance, the granulocytes and plasmatocytes assemble focal complexes and focal adhesions and spread by protruding filopodia/lamellipodia. The well-spread, cultured hemocytes were subjected to MpVLPs exposure, and the morphological changes were observed. The granulocytes lost the focal complexes/adhesions visualized as phosphotyrosine clusters and retracted the filopodia/lamellipodia within 30min after exposure, while the plasmatocytes exhibited similar but distinct responses. The two hemocyte species prepared from either parasitized or MpVLP-injected hosts lost the ability to form both filopodia/lamellipodia and phosphotyrosine clusters. A caspase inhibitor, Z-VAD-FMK, did not affect these MpVLP-induced morphological changes, indicating that these earlier changes found in the hemocytes precede apoptosis. The present study together with our previous data has established that the attenuation of host immune defense by the MpVLPs comprises at least two temporally distinguishable phases: immediate and early inhibition of hemocyte spreading and the eventual induction of hemocyte apoptosis.

  19. Sperm Impairment by Sperm Agglutinating Factor Isolated from Escherichia coli: Receptor Specific Interactions

    Directory of Open Access Journals (Sweden)

    Kiranjeet Kaur

    2013-01-01

    Full Text Available In an earlier work done in our laboratory, we have been able to isolate a sperm agglutinating strain of Escherichia coli from the semen sample of a male attending infertility clinic. Further, factor responsible for sperm agglutination (SAF was isolated and purified, and, using SAF as a tool, corresponding SAF binding receptor from human spermatozoa has been purified. Characterization of SAF and SAF binding receptor using MALDI-TOF showed homology to glutamate decarboxylase and MHC class I molecule, respectively. Coincubation of SAF with spermatozoa not only resulted in spermagglutination but could also compromise other sperm parameters, namely, Mg2+ dependent ATPase activity and apoptosis. Intravaginal administration of SAF could lead to infertility in Balb/c mice. SAF induced impairment of sperm parameters, and infertility was observed to be due to interaction of SAF with sperm surface receptor component as, when purified receptor was introduced, receptor completely inhibited all the detrimental effects induced by SAF. From these results, it could be concluded that interaction of SAF with spermatozoa is receptor mediated.

  20. Incorporation of T4 bacteriophage in electrospun fibres.

    Science.gov (United States)

    Korehei, R; Kadla, J

    2013-05-01

    Antibacterial food packaging materials, such as bacteriophage-activated electrospun fibrous mats, may address concerns triggered by waves of bacterial food contamination. To address this, we investigated several efficient methods for incorporating T4 bacteriophage into electrospun fibrous mats. The incorporation of T4 bacteriophage using simple suspension electrospinning led to more than five orders of magnitude decrease in bacteriophage activity. To better maintain bacteriophage viability, emulsion electrospinning was developed where the T4 bacteriophage was pre-encapsulated in an alginate reservoir via an emulsification process and subsequently electrospun into fibres. This resulted in an increase in bacteriophage viability, but there was still two orders of magnitude drop in activity. Using a coaxial electrospinning process, full bacteriophage activity could be maintained. In this process, a core/shell fibre structure was formed with the T4 bacteriophage being directly incorporated into the fibre core. The core/shell fibre encapsulated bacteriophage exhibited full bacteriophage viability after storing for several weeks at +4°C. Coaxial electrospinning was shown to be capable of encapsulating bacteriophages with high loading capacity, high viability and long storage time. These results are significant in the context of controlling and preventing bacterial infections in perishable foods during storage. © 2013 The Society for Applied Microbiology.

  1. Rapid latex agglutination test for the serodiagnosis of human brucellosis

    NARCIS (Netherlands)

    Abdoel, Theresia H.; Smits, Henk L.

    2007-01-01

    We developed and evaluated a user-friendly latex agglutination assay for the serodiagnosis of human brucellosis. The assay was obtained by coating colored latex beads with Brucella lipopolysaccharides and drying of the activated beads onto white agglutination cards. Individual cards were sealed in a

  2. Bacteriophage Assembly

    Directory of Open Access Journals (Sweden)

    Anastasia A. Aksyuk

    2011-02-01

    Full Text Available Bacteriophages have been a model system to study assembly processes for over half a century. Formation of infectious phage particles involves specific protein-protein and protein-nucleic acid interactions, as well as large conformational changes of assembly precursors. The sequence and molecular mechanisms of phage assembly have been elucidated by a variety of methods. Differences and similarities of assembly processes in several different groups of bacteriophages are discussed in this review. The general principles of phage assembly are applicable to many macromolecular complexes.

  3. Application of direct agglutination test (DAT) and fast agglutination screening test (FAST) for sero-diagnosis of visceral leishmaniasis in endemic area of Minas Gerais, Brazil

    NARCIS (Netherlands)

    Silva, Eduardo S.; Schoone, Gerard J.; Gontijo, Celia M. F.; Brazil, Reginaldo P.; Pacheco, Raquel S.; Schallig, Henk D. F. H.

    2005-01-01

    The direct agglutination test (DAT) has proved to be a very important sero-diagnostic tool combining high levels of intrinsic validity and ease of performance. Otherwise, fast agglutination screening test (FAST) utilises only one serum dilution making the test very suitable for the screening of

  4. The ascidian Styela plicata hemocytes as a potential biomarker of marine pollution: In vitro effects of seawater and organic mercury.

    Science.gov (United States)

    Parrinello, D; Bellante, A; Parisi, M G; Sanfratello, M A; Indelicato, S; Piazzese, D; Cammarata, M

    2017-02-01

    Toxic metals, such as mercury, contribute substantially to anthropogenic pollution in many estuarine environments. Animals living in those environments, particularly invertebrate filter feeders like tunicates, can be used as bioindicators. In an attempt to identify cellular markers for revealing pollution, this study examined in vitro the effects of different concentrations of methyl mercury on Styela plicata hemocytes. The harvested hemocytes from S. plicata that were exposed to the metal had a significant mortality, cellular count and morphometric alterations. These findings provided evidence of MeHg immunotoxic effects on S. plicata, resulting in hemocyte death and morphological changes induced by cytoskeleton alterations. Thus, a morphometric cellular parameter, such as spreading ability, was used as a complementary method for differentiation between hemocytes treated with a marine solution (as a negative control) and hemocytes incubated with methylmercury and/or Sicilian seawater samples. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Evolution of Shock Melt Compositions in Lunar Agglutinates

    Science.gov (United States)

    Vance, A. M.; Christoffersen, R.; Keller, L. P.

    2015-01-01

    Lunar agglutinates are aggregates of regolith grains fused together in a glassy matrix of shock melt produced during smaller-scale (mostly micrometeorite) impacts. Agglutinate formation is a key space weathering process under which the optically-active component of nanophase metallic Fe (npFe(sup 0)) is added to the lunar regolith. Here we have used energy-dispersive X-ray (EDX) compositional spectrum imaging in the SEM to quantify the chemical homogeneity of agglutinitic glass, correlate its homogeneity to its parent soil maturity, and identify the principle chemical components contributing to the shock melt compositional variations.

  6. Pyrovanadolysis: a Pyrophosphorolysis-like Reaction Mediated by Pyrovanadate MN2plus and DNA Polymerase of Bacteriophage T7

    Energy Technology Data Exchange (ETDEWEB)

    B Akabayov; A Kulczyk; S Akabayov; C Thiele; L McLaughlin; B Beauchamp; C Richardson

    2011-12-31

    DNA polymerases catalyze the 3'-5'-pyrophosphorolysis of a DNA primer annealed to a DNA template in the presence of pyrophosphate (PP{sub i}). In this reversal of the polymerization reaction, deoxynucleotides in DNA are converted to deoxynucleoside 5'-triphosphates. Based on the charge, size, and geometry of the oxygen connecting the two phosphorus atoms of PP{sub i}, a variety of compounds was examined for their ability to carry out a reaction similar to pyrophosphorolysis. We describe a manganese-mediated pyrophosphorolysis-like activity using pyrovanadate (VV) catalyzed by the DNA polymerase of bacteriophage T7. We designate this reaction pyrovanadolysis. X-ray absorption spectroscopy reveals a shorter Mn-V distance of the polymerase-VV complex than the Mn-P distance of the polymerase-PP{sub i} complex. This structural arrangement at the active site accounts for the enzymatic activation by Mn-VV. We propose that the Mn{sup 2+}, larger than Mg{sup 2+}, fits the polymerase active site to mediate binding of VV into the active site of the polymerase. Our results may be the first documentation that vanadium can substitute for phosphorus in biological processes.

  7. Use of Biomphalaria glabrata hemocytes as a biomarker of exposure to ionizing radiation

    Energy Technology Data Exchange (ETDEWEB)

    Silva, Hianna A.M.F.; Lima, Maira V.; Sa, Jose L.F.; Siqueira, Williams N.; Luna Filho, Ricardo L.C.; Melo, Larissa S.A.; Morais, Vinicius H.T.; Melo, Ana M.M.A., E-mail: hiannaamfs@gmail.com, E-mail: mairavasconceloslima@gmail.com, E-mail: luismuma6@gmail.com, E-mail: williams.wns@gmail.com, E-mail: ricardolclf@hotmail.com, E-mail: larissamelo.pe@gmail.com, E-mail: viniciushtmorais@hotmail.com, E-mail: amdemelo@hotmail.com [Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil). Departamento de Biofisica e Radiobiologia

    2017-11-01

    The increase of the applications of ionizing radiation in several areas and sectors of modern society has given rise to a greater probability of occurrence of accidents. These accidental occurrences have revealed the need for methods that provide quantitative data on the radiation doses absorbed by biological systems. The mollusk Biomphalaria glabrata presents as a good bioindicator in several works referenced in the literature. In this way, the objective of this work was to evaluate the morphological and quantitative alterations of hemocytes of the Biomphalaria glabrata exposed to ionizing radiation. For the experiments, adult mollusks of the species B. glabrata pigmented were used. The selected mollusks were divided into six groups: five submitted to doses of 10, 20, 30, 40, 50 Gy of {sup 60}Co gamma radiation and the control group. After 48 h, the slides were prepared and then read in a microscope. Quantitative analysis showed a decrease in the total number of hemocytes after irradiation. In the cell classification, a higher number of hyalinocytes were observed in relation to the number of granulocytes, except for the animals exposed to a dose of 30 Gy. The presence of micronuclei and binucleations were observed at all doses used. Apoptosis was observed at doses starting at 30 Gy. Therefore, it is possible to conclude that the morphological and quantitative analysis of B. glabrata hemocytes provided significant data for the identification of biological damage caused by ionizing radiation, allowing the beginning of standardization of the morphological alteration counting technique in B. glabrata hemocytes as An environmental biomarker for the action of physical agents. (author)

  8. Use of Biomphalaria glabrata hemocytes as a biomarker of exposure to ionizing radiation

    International Nuclear Information System (INIS)

    Silva, Hianna A.M.F.; Lima, Maira V.; Sa, Jose L.F.; Siqueira, Williams N.; Luna Filho, Ricardo L.C.; Melo, Larissa S.A.; Morais, Vinicius H.T.; Melo, Ana M.M.A.

    2017-01-01

    The increase of the applications of ionizing radiation in several areas and sectors of modern society has given rise to a greater probability of occurrence of accidents. These accidental occurrences have revealed the need for methods that provide quantitative data on the radiation doses absorbed by biological systems. The mollusk Biomphalaria glabrata presents as a good bioindicator in several works referenced in the literature. In this way, the objective of this work was to evaluate the morphological and quantitative alterations of hemocytes of the Biomphalaria glabrata exposed to ionizing radiation. For the experiments, adult mollusks of the species B. glabrata pigmented were used. The selected mollusks were divided into six groups: five submitted to doses of 10, 20, 30, 40, 50 Gy of "6"0Co gamma radiation and the control group. After 48 h, the slides were prepared and then read in a microscope. Quantitative analysis showed a decrease in the total number of hemocytes after irradiation. In the cell classification, a higher number of hyalinocytes were observed in relation to the number of granulocytes, except for the animals exposed to a dose of 30 Gy. The presence of micronuclei and binucleations were observed at all doses used. Apoptosis was observed at doses starting at 30 Gy. Therefore, it is possible to conclude that the morphological and quantitative analysis of B. glabrata hemocytes provided significant data for the identification of biological damage caused by ionizing radiation, allowing the beginning of standardization of the morphological alteration counting technique in B. glabrata hemocytes as An environmental biomarker for the action of physical agents. (author)

  9. Immune responses and ultrastructural changes of hemocytes in freshwater crab Sinopotamon henanense exposed to elevated cadmium

    Energy Technology Data Exchange (ETDEWEB)

    Qin Qin [College of Life Science, Shanxi University, Taiyuan, Shanxi, 030006 (China); Central Laboratory, Shanxi Provincial People' s Hospital, Taiyuan, Shanxi, 030012 (China); Qin Shengjuan [College of Life Science, Shanxi University, Taiyuan, Shanxi, 030006 (China); Wang Lan, E-mail: lanwang@sxu.edu.cn [College of Life Science, Shanxi University, Taiyuan, Shanxi, 030006 (China); Lei Wenwen [College of Life Science, Shanxi University, Taiyuan, Shanxi, 030006 (China)

    2012-01-15

    Cadmium (Cd) is one of the most toxic heavy metals that can impact immunological parameters in aquatic animals. To investigate the immunotoxicity and ultrastructural changes of hemocytes, specimens of Sinopotamon henanense were exposed to different concentrations of cadmium and the differences in immunologic parameters between Cd exposure groups and control groups were investigated. Total hemocyte count (THC) in Cd-exposure groups were decreased significantly when compared with the control groups, especially in the groups treated with higher Cd concentrations and longer exposure time, while no significant differences were observed in the proportions of the three types of hemocytes. Phenoloxidase (PO) activities were significantly higher in Cd-exposure groups than the control groups. Superoxide dismutase (SOD) activities gradually increased in 7.25 and 14.5 mg L{sup -1} Cd groups, but in other higher Cd groups, they showed first increase and following decrease with the exposure time prolonged. Acid phosphatase (ACP) activities were induced at 48 h, and then decreased, while alkaline phosphatase (AKP) activities increased gradually until 96 h. Electron microscopic results showed that nucleus, mitochondria and rough endoplasm recutulum (rER) of three types of hemocytes were sensitive to acute Cd toxicity. In Cd-exposed groups, chromatin condensation, nucleus deformation and nucleus envelope rupture were noted. Additionally, mitochondrial dilation and rER degranulation were observed in Cd-treated crabs. These results suggested that immune response and organelles of hemocyte of S. henanense were impacted by Cd exposure, and the changes of these immunologic parameters reflect changes in crab immune response capability consequent to Cd exposure.

  10. Immune responses and ultrastructural changes of hemocytes in freshwater crab Sinopotamon henanense exposed to elevated cadmium

    International Nuclear Information System (INIS)

    Qin Qin; Qin Shengjuan; Wang Lan; Lei Wenwen

    2012-01-01

    Cadmium (Cd) is one of the most toxic heavy metals that can impact immunological parameters in aquatic animals. To investigate the immunotoxicity and ultrastructural changes of hemocytes, specimens of Sinopotamon henanense were exposed to different concentrations of cadmium and the differences in immunologic parameters between Cd exposure groups and control groups were investigated. Total hemocyte count (THC) in Cd-exposure groups were decreased significantly when compared with the control groups, especially in the groups treated with higher Cd concentrations and longer exposure time, while no significant differences were observed in the proportions of the three types of hemocytes. Phenoloxidase (PO) activities were significantly higher in Cd-exposure groups than the control groups. Superoxide dismutase (SOD) activities gradually increased in 7.25 and 14.5 mg L −1 Cd groups, but in other higher Cd groups, they showed first increase and following decrease with the exposure time prolonged. Acid phosphatase (ACP) activities were induced at 48 h, and then decreased, while alkaline phosphatase (AKP) activities increased gradually until 96 h. Electron microscopic results showed that nucleus, mitochondria and rough endoplasm recutulum (rER) of three types of hemocytes were sensitive to acute Cd toxicity. In Cd-exposed groups, chromatin condensation, nucleus deformation and nucleus envelope rupture were noted. Additionally, mitochondrial dilation and rER degranulation were observed in Cd-treated crabs. These results suggested that immune response and organelles of hemocyte of S. henanense were impacted by Cd exposure, and the changes of these immunologic parameters reflect changes in crab immune response capability consequent to Cd exposure.

  11. Phagocytic activities of hemocytes from the deep-sea symbiotic mussels Bathymodiolus japonicus, B. platifrons, and B. septemdierum.

    Science.gov (United States)

    Tame, Akihiro; Yoshida, Takao; Ohishi, Kazue; Maruyama, Tadashi

    2015-07-01

    Deep-sea mytilid mussels harbor symbiotic bacteria in their gill epithelial cells that are horizontally or environmentally transmitted to the next generation of hosts. To understand the immune defense system in deep-sea symbiotic mussels, we examined the hemocyte populations of the symbiotic Bathymodiolus mussel species Bathymodiolus japonicus, Bathymodiolus platifrons, and Bathymodiolus septemdierum, and characterized three types of hemocytes: agranulocytes (AGs), basophilic granulocytes (BGs), and eosinophilic granulocytes (EGs). Of these, the EG cells were the largest (diameter, 8.4-10.0 μm) and had eosinophilic cytoplasm with numerous eosinophilic granules (diameter, 0.8-1.2 μm). Meanwhile, the BGs were of medium size (diameter, 6.7-8.0 μm) and contained small basophilic granules (diameter, 0.3-0.4 μm) in basophilic cytoplasm, and the AGs, the smallest of the hemocytes (diameter, 4.8-6.0 μm), had basophilic cytoplasm lacking granules. A lectin binding assay revealed that concanavalin A bound to all three hemocyte types, while wheat germ agglutinin bound exclusively to EGs and BGs. The total hemocyte population densities within the hemolymph of all three Bathymodiolus mussel species were similar (8.4-13.3 × 10(5) cells/mL), and the percentages of circulating AGs, BGs, and EGs in the hemolymph of these organisms were 44.7-48.5%, 14.3-17.6%, and 34.3-41.0%, respectively. To analyze the functional differences between these hemocytes, the phagocytic activity and post-phagocytic phagosome-lysosome fusion events were analyzed in each cell type using a fluorescent Alexa Fluor(®) 488-conjugated Escherichia coli bioparticle and a LysoTracker(®) lysosomal marker, respectively. While the AGs exhibited no phagocytic activity, both types of granulocytes were phagocytic. Of the three hemocyte types, the EGs exhibited the highest level of phagocytic activity as well as rapid phagosome-lysosome fusion, which occurred within 2 h of incubation. Meanwhile, the BGs showed

  12. Immune Defenses of the Invasive Apple Snail Pomacea canaliculata (Caenogastropoda, Ampullariidae: Phagocytic Hemocytes in the Circulation and the Kidney.

    Directory of Open Access Journals (Sweden)

    Juan A Cueto

    Full Text Available Hemocytes in the circulation and kidney islets, as well as their phagocytic responses to microorganisms and fluorescent beads, have been studied in Pomacea canaliculata, using flow cytometry, light microscopy (including confocal laser scanning microscopy and transmission electron microscopy (TEM. Three circulating hemocyte types (hyalinocytes, agranulocytes and granulocytes were distinguished by phase contrast microscopy of living cells and after light and electron microscopy of fixed material. Also, three different populations of circulating hemocytes were separated by flow cytometry, which corresponded to the three hemocyte types. Hyalinocytes showed a low nucleus/cytoplasm ratio, and no apparent granules in stained material, but showed granules of moderate electron density under TEM (L granules and at least some L granules appear acidic when labeled with LysoTracker Red. Both phagocytic and non-phagocytic hyalinocytes lose most (if not all L granules when exposed to microorganisms in vitro. The phagosomes formed differed whether hyalinocytes were exposed to yeasts or to Gram positive or Gram negative bacteria. Agranulocytes showed a large nucleus/cytoplasm ratio and few or no granules. Granulocytes showed a low nucleus/cytoplasm ratio and numerous eosinophilic granules after staining. These granules are electron dense and rod-shaped under TEM (R granules. Granulocytes may show merging of R granules into gigantic ones, particularly when exposed to microorganisms. Fluorescent bead exposure of sorted hemocytes showed phagocytic activity in hyalinocytes, agranulocytes and granulocytes, but the phagocytic index was significantly higher in hyalinocytes. Extensive hemocyte aggregates ('islets' occupy most renal hemocoelic spaces and hyalinocyte-like cells are the most frequent component in them. Presumptive glycogen deposits were observed in most hyalinocytes in renal islets (they also occur in the circulation but less frequently and may mean that

  13. Effects of cadmium on aneuploidy and hemocyte parameters in the Pacific oyster, Crassostrea gigas

    International Nuclear Information System (INIS)

    Bouilly, Karine; Gagnaire, Beatrice; Bonnard, Marc; Thomas-Guyon, Helene; Renault, Tristan; Miramand, Pierre; Lapegue, Sylvie

    2006-01-01

    Pacific oysters, Crassostrea gigas, are commonly reared in estuaries where they are exposed to anthropogenic pollution. Much research has been made on the toxicity of cadmium to aquatic organisms because the compound recurrently contaminates their environment. Our study examined the influence of cadmium on aneuploidy level (lowered chromosome number in a percentage of somatic cells) and hemocyte parameters in C. gigas at different stages of life. Adults and juveniles were exposed to two different concentrations of cadmium. The first concentration applied was equivalent to a peak value found in Marennes-Oleron bay (Charente-Maritime, France; 50 ng L -1 ) and the second was 10 times higher (500 ng L -1 ). Exposure to 50 ng L -1 cadmium caused a significant decrease in the survival time of C. gigas, but exposure to 500 ng L -1 surprisingly affected the survival time positively. Significant differences in aneuploidy level were observed between the cadmium treatments and the control in adults but not in juveniles or the offspring of the adult groups. The effects of cadmium on hemocyte parameters were analyzed by flow cytometry. Several hemocyte parameters increased significantly after 21 days of cadmium exposure and subsequently decreased. Phenoloxidase-like activity, evaluated by spectrophotometry, varied over the time of the experiment and increased after 66 days of contact with 500 ng L -1 cadmium. Taken together, cadmium at environmentally relevant concentrations seems to have only moderate effects on aneuploidy and hemocyte parameters

  14. Imaging based agglutination measurement of magnetic micro-particles on a lab-on-a-disk platform

    DEFF Research Database (Denmark)

    Wantiya, P.; Burger, Robert; Alstrøm, Tommy Sonne

    2014-01-01

    In this work we present a magnetic micro beads based agglutination assay on a centrifugal microfluidic platform. An imaging based method is used to quantify bead agglutination and measure the concentration of antibodies or C-reactive protein in solution.......In this work we present a magnetic micro beads based agglutination assay on a centrifugal microfluidic platform. An imaging based method is used to quantify bead agglutination and measure the concentration of antibodies or C-reactive protein in solution....

  15. Cationic antimicrobial peptides inactivate Shiga toxin-encoding bacteriophages

    Science.gov (United States)

    Del Cogliano, Manuel E.; Hollmann, Axel; Martinez, Melina; Semorile, Liliana; Ghiringhelli, Pablo D.; Maffía, Paulo C.; Bentancor, Leticia V.

    2017-12-01

    Shiga toxin (Stx) is the principal virulence factor during Shiga toxin-producing Escherichia coli (STEC) infections. We have previously reported the inactivation of bacteriophage encoding Stx after treatment with chitosan, a linear polysaccharide polymer with cationic properties. Cationic antimicrobial peptides (cAMPs) are short linear aminoacidic sequences, with a positive net charge, which display bactericidal or bacteriostatic activity against a wide range of bacterial species. They are promising novel antibiotics since they have shown bactericidal effects against multiresistant bacteria. To evaluate whether cationic properties are responsible for bacteriophage inactivation, we tested seven cationic peptides with proven antimicrobial activity as anti-bacteriophage agents, and one random sequence cationic peptide with no antimicrobial activity as a control. We observed bacteriophage inactivation after incubation with five cAMPs, but no inactivating activity was observed with the random sequence cationic peptide or with the non alpha helical cAMP Omiganan. Finally, to confirm peptide-bacteriophage interaction, zeta potential was analyzed by following changes on bacteriophage surface charges after peptide incubation. According to our results we could propose that: 1) direct interaction of peptides with phage is a necessary step for bacteriophage inactivation, 2) cationic properties are necessary but not sufficient for bacteriophage inactivation, and 3) inactivation by cationic peptides could be sequence (or structure) specific. Overall our data suggest that these peptides could be considered a new family of molecules potentially useful to decrease bacteriophage replication and Stx expression.

  16. Cationic Antimicrobial Peptides Inactivate Shiga Toxin-Encoding Bacteriophages

    Directory of Open Access Journals (Sweden)

    Manuel E. Del Cogliano

    2017-12-01

    Full Text Available Shiga toxin (Stx is the principal virulence factor during Shiga toxin-producing Escherichia coli (STEC infections. We have previously reported the inactivation of bacteriophage encoding Stx after treatment with chitosan, a linear polysaccharide polymer with cationic properties. Cationic antimicrobial peptides (cAMPs are short linear aminoacidic sequences, with a positive net charge, which display bactericidal or bacteriostatic activity against a wide range of bacterial species. They are promising novel antibiotics since they have shown bactericidal effects against multiresistant bacteria. To evaluate whether cationic properties are responsible for bacteriophage inactivation, we tested seven cationic peptides with proven antimicrobial activity as anti-bacteriophage agents, and one random sequence cationic peptide with no antimicrobial activity as a control. We observed bacteriophage inactivation after incubation with five cAMPs, but no inactivating activity was observed with the random sequence cationic peptide or with the non-alpha helical cAMP Omiganan. Finally, to confirm peptide-bacteriophage interaction, zeta potential was analyzed by following changes on bacteriophage surface charges after peptide incubation. According to our results we could propose that: (1 direct interaction of peptides with phage is a necessary step for bacteriophage inactivation, (2 cationic properties are necessary but not sufficient for bacteriophage inactivation, and (3 inactivation by cationic peptides could be sequence (or structure specific. Overall our data suggest that these peptides could be considered a new family of molecules potentially useful to decrease bacteriophage replication and Stx expression.

  17. The immune response of hemocytes of the insect Oncopeltus fasciatus against the flagellate Phytomonas serpens.

    Directory of Open Access Journals (Sweden)

    Thiago L Alves e Silva

    Full Text Available The genus Phytomonas includes parasites that are etiological agents of important plant diseases, especially in Central and South America. These parasites are transmitted to plants via the bite of an infected phytophagous hemipteran. Despite the economic impact of these parasites, many basic questions regarding the genus Phytomonas remain unanswered, such as the mechanism by which the parasites cope with the immune response of the insect vector. In this report, using a model of systemic infection, we describe the function of Oncopeltus fasciatus hemocytes in the immune response towards the tomato parasite Phytomonas serpens. Hemocytes respond to infection by trapping parasites in nodular structures and phagocytizing the parasites. In electron microscopy of hemocytes, parasites were located inside vacuoles, which appear fused with lysosomes. The parasites reached the O. fasciatus salivary glands at least six hours post-infection. After 72 hours post-infection, many parasites were attached to the salivary gland outer surface. Thus, the cellular responses did not kill all the parasites.

  18. Two-Phase Microfluidic Systems for High Throughput Quantification of Agglutination Assays

    KAUST Repository

    Castro, David

    2018-04-01

    Lab-on-Chip, the miniaturization of the chemical and analytical lab, is an endeavor that seems to come out of science fiction yet is slowly becoming a reality. It is a multidisciplinary field that combines different areas of science and engineering. Within these areas, microfluidics is a specialized field that deals with the behavior, control and manipulation of small volumes of fluids. Agglutination assays are rapid, single-step, low-cost immunoassays that use microspheres to detect a wide variety molecules and pathogens by using a specific antigen-antibody interaction. Agglutination assays are particularly suitable for the miniaturization and automation that two-phase microfluidics can offer, a combination that can help tackle the ever pressing need of high-throughput screening for blood banks, epidemiology, food banks diagnosis of infectious diseases. In this thesis, we present a two-phase microfluidic system capable of incubating and quantifying agglutination assays. The microfluidic channel is a simple fabrication solution, using laboratory tubing. These assays are incubated by highly efficient passive mixing with a sample-to-answer time of 2.5 min, a 5-10 fold improvement over traditional agglutination assays. It has a user-friendly interface that that does not require droplet generators, in which a pipette is used to continuously insert assays on-demand, with no down-time in between experiments at 360 assays/h. System parameters are explored, using the streptavidin-biotin interaction as a model assay, with a minimum detection limit of 50 ng/mL using optical image analysis. We compare optical image analysis and light scattering as quantification methods, and demonstrate the first light scattering quantification of agglutination assays in a two-phase ow format. The application can be potentially applied to other biomarkers, which we demonstrate using C-reactive protein (CRP) assays. Using our system, we can take a commercially available CRP qualitative slide

  19. Morphological and functional characterization of hemocytes from two deep-sea vesicomyid clams Phreagena okutanii and Abyssogena phaseoliformis.

    Science.gov (United States)

    Tame, Akihiro; Ozawa, Genki; Maruyama, Tadashi; Yoshida, Takao

    2018-03-01

    Deep-sea vesicomyid clams harboring intracellular symbiotic sulfur-oxidizing bacteria are often dominant in chemosynthetic animal communities. Although they are known to have erythrocytes, little is known about other hemocytes. To investigate the types and roles of various hemocytes in vesicomyid clams, we performed morphological, histochemical and functional characterization of the hemocytes in two species, Phreagena okutanii, collected from 873 to 978 m depth, and Abyssogena phaseoliformis, from 5199 to 5355 m. Both were found to have three types of hemocytes: erythrocytes (ERCs), eosinophilic granulocytes (EGs), and basophilic granulocytes (BGs). The ERCs contain hemoglobin in the cytoplasm, with basophilic vacuoles containing acid polysaccharide, neutral lipids, and peroxidase. The EGs were found to contain acid polysaccharides and eosinophilic granules containing lysosomal enzymes, acid and alkaline phosphatases, chloroacetate esterase, and peroxidase. Although BGs had some basophilic granules with alkaline phosphatase, they lacked acid phosphatase and acid polysaccharides. The EGs and BGs were shown to have phagocytic ability, while the ERCs exhibited no phagocytosis. The EGs showed higher phagocytic activity as well as a higher phagosome-lysosome fusion rate than BGs. The hemocytes of the two vesicomyid species differed in the intracellular structures. In A. phaseoliformis, ERCs additionally contained neutral polysaccharides in vacuoles and had vesicles with acinus-like acidic mucus in the cytoplasm, neither of which were observed in P. okutanii. The eosinophilic granules in the EGs had heteromorphically-elongated shapes containing homogeneously electron-dense material in P. okutanii, but were more spherical and composed of fibrous structures in A. phaseoliformis. The difference in hemocytes between the two clams seems to be reflective of phylogenetically differentiated lineages adapting to differing conditions in their respective deep-sea environments

  20. INFLUENCE OF SEASONAL FACTORS ON OYSTER HEMOCYTE KILLING OF VIBRIO PARAHEMOLYTICUS

    Science.gov (United States)

    Seasonal variation of cellular defenses of oyster Crassostrea virginica against Vibrio parahaemolyticus were examined from June 1997 to December 1998 using a recently developed bactericidal assay that utilizes a tetrazolium dye. Mean hemocyte numbers, plasma lysozyme, and P. mari...

  1. Campylobacter jejuni acquire new host-derived CRISPR spacers when in association with bacteriophages harbouring a CRISPR-like Cas4 protein

    Directory of Open Access Journals (Sweden)

    Ian F. Connerton

    2015-01-01

    Full Text Available Campylobacter jejuni is a worldwide cause of human diarrhoeal disease. Clustered Repetitively Interspaced Palindromic Repeats (CRISPRs and associated proteins allow Bacteria and Archaea to evade bacteriophage and plasmid infection. Type II CRISPR systems are found in association with combinations of genes encoding the CRISPR-associated Cas1, Cas2, Cas4 or Csn2, and Cas9 proteins. C. jejuni possesses a minimal subtype II-C CRISPR system containing cas1, cas2, and cas9 genes whilst cas4 is notably absent. Cas4 proteins possess 5ʹ-3ʹ exonuclease activity to create recombinogenic-ends for spacer acquisition. Here we report a conserved Cas4-like protein in Campylobacter bacteriophages that creates a novel split arrangement between the bacteriophage and host that represents a new twist in the bacteriophage/host co-evolutionary arms race. The continuous association of bacteriophage and host in the carrier state life cycle of C. jejuni provided an opportunity to study spacer acquisition in this species. Remarkably all the spacer sequences observed were of host origin. We hypothesise that Campylobacter bacteriophages can use Cas4-like protein to activate spacer acquisition to use host DNA as an effective decoy to bacteriophage DNA. Bacteria that acquire self-spacers and escape phage infection must overcome CRISPR-mediated autoimmunity either by loss of the interference functions leaving them susceptible to foreign DNA incursion or tolerate changes in gene regulation.

  2. Bacteriophages

    International Nuclear Information System (INIS)

    Klieve, A.V.

    2005-01-01

    Bacteriophages or phages are bacterial viruses and are present in the rumen in large numbers. They are obligate pathogens of bacteria and are ubiquitous to the rumen ecosystem. Bacteriophages are capable of lysing their bacterial hosts within the rumen and are therefore regarded as contributing to protein recycling within the rumen, a process identified as reducing the efficiency of feed utilization. However, their presence may not be entirely detrimental to the ecosystem, and it has been argued that phages may also be involved in the maintenance of a balanced ecosystem and may play a role in recycling limiting nutrients within the rumen. Furthermore, phage therapy is enjoying a renaissance and the use of phages to control or eliminate detrimental or unwanted microbes from the gastro-intestinal tract, such as Shiga-toxin producing E. coli (food-borne disease), Streptococcus bovis (acidosis in grain-fed cattle) and methanogens (produce the greenhouse gas methane), is the focus of current investigation. In order to be able to study the interaction between individual bacteriophages and their bacterial hosts, it is necessary to: (a) isolate the phage of interest from other viruses in the source material; (b) to derive stock cultures of known phage concentration; (c) store the isolated phages; and (d) determine basic physical characteristics, such as morphology. These procedures are achieved using classical microbiological procedures and this will be the methodology described in this chapter. It is also necessary to determine nucleic acid characteristics of the phage genome and to fingerprint the phage population in the rumen using molecular biological techniques. These will be described and discussed in Chapter 4.2

  3. A study of the incubation of microbead agglutination assays in a microfluidic system

    KAUST Repository

    Castro, David

    2016-12-19

    This work reports on a quantitative study of the incubation of a microbead-based agglutination assay inside a microfluidic system. In this system, a droplet (1.25µL) consisting of a mixture of functionalized microbeads and analyte is flowed through a 0.51mm internal diameter silicone tube. Hydrodynamic forces alone produce a very efficient mixing of the beads within the droplet. We tested the agglutination at different speeds and show a robust response at the higher range of speeds (150 – 200µL/min), while also reaching a completion in the agglutination process. At these velocities, a length of 180cm is shown to be sufficient to confidently measure the agglutination assay, which takes between 2.5 – 3 minutes. This high throughput quantification method has the potential of accelerating the measurements of various types of biomarkers, which can greatly benefit the fields of biology and medicine.

  4. IN VITRO KILLING OF PERKINSUS MARINUS BY HEMOCYTES OF OYSTERS CRASSOSTREA VIRGINICA

    Science.gov (United States)

    A colorimetric microbicidal assay was adapted, optimized and applied in experiments to characterize the in vitro capacity of eastern oyster (Crassostrea virginica) hemocytes to kill cultured isolates of Perkinsus marinus, a protozoan parasite causing a highly destructive disease...

  5. Red Blood Cell Agglutination for Blood Typing Within Passive Microfluidic Biochips.

    Science.gov (United States)

    Huet, Maxime; Cubizolles, Myriam; Buhot, Arnaud

    2018-04-19

    Pre-transfusion bedside compatibility test is mandatory to check that the donor and the recipient present compatible groups before any transfusion is performed. Although blood typing devices are present on the market, they still suffer from various drawbacks, like results that are based on naked-eye observation or difficulties in blood handling and process automation. In this study, we addressed the development of a red blood cells (RBC) agglutination assay for point-of-care blood typing. An injection molded microfluidic chip that is designed to enhance capillary flow contained anti-A or anti-B dried reagents inside its microchannel. The only blood handling step in the assay protocol consisted in the deposit of a blood drop at the tip of the biochip, and imaging was then achieved. The embedded reagents were able to trigger RBC agglutination in situ, allowing for us to monitor in real time the whole process. An image processing algorithm was developed on diluted bloods to compute real-time agglutination indicator and was further validated on undiluted blood. Through this proof of concept, we achieved efficient, automated, real time, and quantitative measurement of agglutination inside a passive biochip for blood typing which could be further generalized to blood biomarker detection and quantification.

  6. Immunomodulation by different types of N-oxides in the hemocytes of the marine bivalve Mytilus galloprovincialis.

    Directory of Open Access Journals (Sweden)

    Caterina Ciacci

    Full Text Available The potential toxicity of engineered nanoparticles (NPs for humans and the environment represents an emerging issue. Since the aquatic environment represents the ultimate sink for NP deposition, the development of suitable assays is needed to evaluate the potential impact of NPs on aquatic biota. The immune system is a sensitive target for NPs, and conservation of innate immunity represents an useful basis for studying common biological responses to NPs. Suspension-feeding invertebrates, such as bivalves, are particularly at risk to NP exposure, since they have extremely developed systems for uptake of nano and microscale particles integral to intracellular digestion and cellular immunity. Evaluation of the effects of NPs on functional parameters of bivalve immunocytes, the hemocytes, may help understanding the major toxic mechanisms and modes of actions that could be relevant for different NP types in aquatic organisms.In this work, a battery of assays was applied to the hemocytes of the marine bivalve Mytilus galloprovincialis to compare the in vitro effects of different n-oxides (n-TiO(2, n-SiO(2, n-ZnO, n-CeO(2 chosen on the basis of their commercial and environmental relevance. Physico-chemical characterization of both primary particles and NP suspensions in artificial sea water-ASW was performed. Hemocyte lysosomal and mitochondrial parameters, oxyradical and nitric oxide production, phagocytic activity, as well as NP uptake, were evaluated. The results show that different n-oxides rapidly elicited differential responses hemocytes in relation to their chemical properties, concentration, behavior in sea water, and interactions with subcellular compartments. These represent the most extensive data so far available on the effects of NPs in the cells of aquatic organisms. The results indicate that Mytilus hemocytes can be utilized as a suitable model for screening the potential effects of NPs in the cells of aquatic invertebrates, and may

  7. Chlamydial plasmids and bacteriophages.

    Science.gov (United States)

    Pawlikowska-Warych, Małgorzata; Śliwa-Dominiak, Joanna; Deptuła, Wiesław

    2015-01-01

    Chlamydia are absolute pathogens of humans and animals; despite being rather well recognised, they are still open for discovery. One such discovery is the occurrence of extrachromosomal carriers of genetic information. In prokaryotes, such carriers include plasmids and bacteriophages, which are present only among some Chlamydia species. Plasmids were found exclusively in Chlamydia (C.) trachomatis, C. psittaci, C. pneumoniae, C. suis, C. felis, C. muridarum and C. caviae. In prokaryotic organisms, plasmids usually code for genes that facilitate survival of the bacteria in the environment (although they are not essential). In chlamydia, their role has not been definitely recognised, apart from the fact that they participate in the synthesis of glycogen and encode proteins responsible for their virulence. Furthermore, in C. suis it was evidenced that the plasmid is integrated in a genomic island and contains the tetracycline-resistance gene. Bacteriophages specific for chlamydia (chlamydiaphages) were detected only in six species: C. psittaci, C. abortus, C. felis, C. caviae C. pecorum and C. pneumoniae. These chlamydiaphages cause inhibition of the developmental cycle, and delay transformation of reticulate bodies (RBs) into elementary bodies (EBs), thus reducing the possibility of infecting other cells in time. Plasmids and bacteriophages can be used in the diagnostics of chlamydioses; although especially in the case of plasmids, they are already used for detection of chlamydial infections. In addition, bacteriophages could be used as therapeutic agents to replace antibiotics, potentially addressing the problem of increasing antibiotic-resistance among chlamydia.

  8. Pacific oyster (Crassostrea gigas) hemocyte are not affected by a mixture of pesticides in short-term in vitro assays.

    Science.gov (United States)

    Moreau, Pierrick; Burgeot, Thierry; Renault, Tristan

    2014-04-01

    Pesticides are frequently detected in estuaries among the pollutants found in estuarine and coastal areas and may have major ecological consequences. They could endanger organism growth, reproduction, or survival. In the context of high-mortality outbreaks affecting Pacific oysters, Crassostrea gigas, in France since 2008, it appears of importance to determine the putative effects of pesticides on oyster susceptibility to infectious agents. Massive mortality outbreaks reported in this species, mainly in spring and summer, may suggest an important role played by the seasonal use of pesticides and freshwater input in estuarine areas where oyster farms are frequently located. To understand the impact of some pesticides detected in French waters, their effects on Pacific oyster hemocytes were studied through short-term in vitro experiments. Bivalve immunity is mainly supported by hemocytes eliminating pathogens by phagocytosis and producing compounds including lysosomal enzymes and antimicrobial molecules. In this study, oyster hemocytes were incubated with a mixture of 14 pesticides and metaldehyde alone, a molecule used to eliminate land mollusks. Hemocyte parameters including dead/alive cells, nonspecific esterase activities, intracytoplasmic calcium, lysosome number and activity, and phagocytosis were monitored by flow cytometry. No significant effect of pesticides tested at different concentrations was reported on oyster hemocytes maintained in vitro for short-term periods in the present study. It could be assumed that these oyster cells were resistant to pesticide exposure in tested conditions and developing in vivo assays appears as necessary to better understand the effects of pollutants on immune system in mollusks.

  9. Hemocyte quantitative changes in Anticarsia gemmatalis (Lepidoptera: Noctuidae larvae infected by AgMNPV

    Directory of Open Access Journals (Sweden)

    Fábio Goulart de Andrade

    2010-04-01

    Full Text Available The initial effects of the infection by AgMNPV in the total and differential counts of the hemocytes in Anticarsia gemmatalis (Lepidoptera: Noctuidae larvae were studied. The total number of the hemocytes did not decrease in infected larvae, as it occurred in non infected larvae. In infected larvae, the hemocyte types showed the following frequencies: plasmatocytes - 47.8%, esferulocytes - 25.9%, granulocytes - 15.8%, oenocytoids - 7.2%, prohemocytes - 2.8%, vermicytes - 0,5%. Only the percentage of the granulocytes was different among infected and non infected larvae, indicating that these cells responded quickly to the initial viral infection. These results showed the effective role of the hemocytes in the response of the A. gemmatalis to the infection by AgMNPV. The comprehension of the immunological mechanisms of this insect is an important tool to understand its biological control.Os efeitos iniciais da infecção por AgMNPV nas contagens total e diferencial dos hemócitos em Anticarsia gemmatalis (Lepidoptera: Noctuidae foram estudados. O número total de hemócitos não diminuiu nas larvas infectadas, como ocorreu nas larvas não infectadas. Nas larvas infectadas, os tipos de hemócitos apresentaram as seguintes freqüências: plasmatócitos - 47,8%, esferulócitos - 25,9%, granulócitos - 15,8%, oenocitóides - 7,2%, prohemócitos - 2,8%, vermiformes - 0,5%. Apenas a porcentagem de granulócitos foi diferente entre larvas infectadas e não infectadas, indicando que estas células responderam rapidamente à infecção viral inicial. Estes resultados mostraram o papel efetivo que dos hemócitos na resposta de A. gemmatalis à infecção por AgMNPV. A compreensão dos mecanismos imunológicos deste inseto é uma ferramenta importante para compreender seu controle biológico.

  10. Polymer-based delivery systems for support and delivery of bacteriophages

    Science.gov (United States)

    Brown, Alyssa Marie

    One of the most urgent problems in the fields of medicine and agriculture is the decreasing effectiveness of antibiotics. Once a miracle drug, antibiotics have recently become associated with the creation of antibiotic-resistant bacteria. The main limitations of these treatments include lack of both adaptability and specificity. To overcome these shortcomings of current antibiotic treatments, there has been a renewed interest in bacteriophage research. Bacteriophages are naturally-occurring viruses that lyse bacteria. They are highly specific, with each bacteriophage type lysing a narrow range of bacteria strains. Bacteriophages are also ubiquitous biological entities, populating environments where bacterial growth is supported. Just as humans are exposed to bacteria in their daily lives, we are exposed to bacteriophages as well. To use bacteriophages in practical applications, they must be delivered to the site of an infection in a controlled-release system. Two systems were studied to observe their support of bacteriophage lytic activity, as well as investigate the possibility of controlling bacteriophage release rates. First, hydrogels were studied, using crosslinking and blending techniques to achieve a range of release profiles. Second, polyanhydride microparticles were studied, evaluating release rates as a function of monomer chemistries.

  11. K. OXYTOCA BACTERIOPHAGES ISOLATION METHODS IMPROVEMENT

    Directory of Open Access Journals (Sweden)

    G. R. Sadrtdinova

    2017-01-01

    Full Text Available The article presents the results of a study related to increasing the efficiency of phage isolation of bacteria of the species K. oxytoca, by developing the optimal composition of the medium used in the work. In scientific research, in almost all methods associated with the isolation of bacteriophages, meat-peptone broth and meat-peptone agar are used as the nutrient basis. The peculiarities of growth and cultivation of microorganisms create certain difficulties for the isolation of phages active against bacteria of the species K. oxytoca. The selection of components and the creation of an environment that would ensure the optimal growth of both the bacterial culture and the reproduction of the virus makes it possible to facilitate the isolation of bacteriophages. The number of bacterial strains used in the work was 7. All strains of cultures were obtained from the Museum of the Department of Microbiology, Virology, Epizootology and Veterinary and Sanitary Expertise of the Federal State Budget Educational Institution of Higher Education “Ulyanovsk State Agrarian University named after P.A. Stolypin”. The studies included 2 main stages. The first stage consisted in isolation of bacteriophages by the method of isolation from the external environment by the method of Adelson L.I., Lyashenko E.A. The material for the studies were samples: soil, sewage sample, fecal samples (2. Only 4 samples. According to the chosen method, the sowing of the putative phagolysate was carried out on meat-peptone agar (1.5% and the agar for isolating bacteriophages (Aph (1.5%. A positive result was the presence on the environment of negative colonies, clearly visible on the matt background of deep growth of bacteria. A negative result is a continuous growth (“lawn” of bacterial culture. As a control, the culture of the microorganism studied was used for the media. In the course of the conducted studies for the first stage, 2 bacteriophages were isolated, active

  12. Rapid and Sensitive Detection of Yersinia pestis Using Amplification of Plague Diagnostic Bacteriophages Monitored by Real-Time PCR

    Science.gov (United States)

    2010-06-01

    2004) Development of a bacteriophage phage replication assay for diagnosis of pulmonary tuberculosis . J Clin Microbiol 42: 2115–2120. 30. Abshire TG...Mycobacterium tuberculosis in Mexico . Clin Microbiol 39: 3883–3888. 38. Schofield DA, Westwater C (2009) Phage-mediated bioluminescent detection of

  13. Bacteria vs. bacteriophages: parallel evolution of immune arsenals

    Directory of Open Access Journals (Sweden)

    Muhammad Abu Bakr Shabbir

    2016-08-01

    Full Text Available Bacteriophages are the most common entities on earth and represent a constant challenge to bacterial populations. To fend off bacteriophage infection, bacteria evolved immune systems to avert phage adsorption and block invader DNA entry. They developed restriction-modification systems and mechanisms to abort infection and interfere with virion assembly, as well as newly recognized clustered regularly interspaced short palindromic repeats (CRISPR. In response to bacterial immune systems, bacteriophages synchronously evolved resistance mechanisms, such as the anti-CRISPR systems to counterattack bacterial CRISPR-cas systems, in a continuing evolutionary arms race between virus and host. In turn, it is fundamental to the survival of the bacterial cell to evolve a system to combat bacteriophage immune strategies.

  14. Are the Major Agglutinative Languages Genetically Related?

    Science.gov (United States)

    Hakola, H. P. A.

    1989-01-01

    Examination of accidental CVC and CV correspondences among languages representing 5 large families of agglutinative languages found that comparison pairs had much more similarity between basic 100-word vocabularies than would have been possible by mere chance, supporting the hypothesis that those 5 language families were mutually related.…

  15. Jaburetox-induced toxic effects on the hemocytes of Rhodnius prolixus (Hemiptera: Reduviidae).

    Science.gov (United States)

    Moyetta, Natalia R; Broll, Valquiria; Perin, Ana Paula A; Uberti, Augusto F; Coste Grahl, Matheus V; Staniscuaski, Fernanda; Carlini, Celia R; Fruttero, Leonardo L

    2017-10-01

    Jaburetox is a recombinant peptide derived from a Canavalia ensiformis urease that presents toxic effects upon several species of insects, phytopathogenic fungi and yeasts of medical importance. So far, no toxicity of Jaburetox to mammals has been shown. Previous reports have identified biochemical targets of this toxic peptide in insect models, although its mechanism of action is not completely understood. In this work, we aimed to characterize the effects of Jaburetox in hemolymphatic insect cells. For this purpose, the model insect and Chagas' disease vector Rhodnius prolixus was used. In vivo and in vitro experiments indicated that Jaburetox interacts with a subset of hemocytes and it can be found in various subcellular compartments. In insects injected with Jaburetox there was an increase in the gene expression of the enzymes UDP-N-acetylglucosamine pyrophosphorylase (UAP), chitin synthase and nitric oxide synthase (NOS). Nevertheless, the expression of NOS protein, the enzyme activities of UAP and acid phosphatase (a possible link between UAP and NOS) as well as the phosphorylation state of proteins remained unchanged upon the in vivo Jaburetox treatment. Nitric oxide (NO) imaging using fluorescent probes showed that Jaburetox augmented NO production in the hemocyte aggregates when compared to controls. Even though Jaburetox activated the hemocytes, as demonstrated by wheat germ agglutinin binding assays, the peptide did not lead to an increase of their phagocytic behavior. Taken together, these findings contribute to our understanding of toxic effects of Jaburetox, a peptide with biotechnological applications and a prospective tool for rational insect control. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Virus-like particles in venom of Meteorus pulchricornis induce host hemocyte apoptosis.

    Science.gov (United States)

    Suzuki, M; Tanaka, T

    2006-06-01

    Ultrastructural studies on the reproductive tract and venom apparatus of a female braconid, Meteorus pulchricornis, revealed that the parasitoid lacks the calyx region in its oviduct, but possesses a venom gland with two venom gland filaments and a venom reservoir filled with white and cloudy fluid. Its venom gland cell is concaved and has a lumen filled with numerous granules. Transmisson electron microscopic (TEM) observation revealed that virus-like particles (VLPs) were produced in venom gland cells. The virus-like particle observed in M. pulchricornis (MpVLP) is composed of membranous envelopes with two different parts: a high-density core and a whitish low-density part. The VLPs of M. pulchricornis is also found assembling ultimately in the lumen of venom gland cell. Microvilli were found thrusting into the lumen of the venom gland cell and seem to aid in driving the matured MpVLPs to the common duct of the venom gland filament. Injection of MpVLPs into non-parasitized Pseudaletia separata hosts induced apoptosis in hemocytes, particularly granulocytes (GRs). Rate of apoptosis induced in GRs peaked 48h after VLP injection. While a large part of the GR population collapsed due to apoptosis caused by MpVLPs, the plasmatocyte population was minimally affected. The capacity of MpVLPs to cause apoptosis in host's hemocytes was further demonstrated by a decrease ( approximately 10-fold) in ability of host hemocytes to encapsulate fluorescent latex beads when MpVLPs were present. Apparently, the reduced encapsulation ability was due to a decrease in the GR population resulting from MpVLP-induced apoptosis.

  17. TAT improves in vitro transportation of fortilin through midgut and into hemocytes of white shrimp Litopenaeus vannamei

    Science.gov (United States)

    Zhou, Yi; Zhang, Wenbing; Mai, Kangsen; Xu, Wei; Zhang, Yanjiao; Ai, Qinghui; Wang, Xiaojie

    2012-06-01

    Fortilin is a multifunctional protein implicated in many important cellular processes. Since injection of Pm-fortilin reduces shrimp mortality caused by white spot syndrome virus (WSSV), there is potential application of fortilin in shrimp culture. In the present study, in order to improve trans-membrane transportation efficiency, the protein transduction domain of the transactivator of transcription (TAT) peptide was fused to fortilin. The Pichia pastoris yeast expression system, which is widely accepted in animal feeds, was used for production of recombinant fusion protein. Green fluorescence protein (GFP) was selected as a reporter because of its intrinsic visible fluorescence. The fortilin, TAT and GFP fusion protein were constructed. Their trans-membrane transportation efficiency and effects on immune response of shrimp were analyzed in vitro. Results showed that TAT peptide improved in vitro uptake of fortilin into the hemocytes and midgut of Litopenaeus vannamei. The phenoloxidase (PO) activity of hemocytes incubated with GFP-Fortilin or GFP-Fortilin-TAT was significantly increased compared with that in the control without expressed fortilin. The PO activity of hemocytes incubated with 200 μg mL-1 GFP-Fortilin-TAT was significantly higher than that in the group with the same concentration of GFP-Fortilin. Hemocytes incubated with GFP-Fortilin-TAT at all concentrations showed significantly higher nitric oxide synthase (NOS) activity than those in the control or in the GFP-Fortilin treatment. The present in vitro study indicated that TAT fusion protein improved the immune effect of fortilin.

  18. Bacteriophage Infectivity Against Pseudomonas aeruginosa in Saline Conditions

    KAUST Repository

    Scarascia, Giantommaso

    2018-05-02

    Pseudomonas aeruginosa is a ubiquitous member of marine biofilm, and reduces thiosulfate to produce toxic hydrogen sulfide gas. In this study, lytic bacteriophages were isolated and applied to inhibit the growth of P. aeruginosa in planktonic mode at different temperature, pH, and salinity. Bacteriophages showed optimal infectivity at a multiplicity of infection of 10 in saline conditions, and demonstrated lytic abilities over all tested temperature (25, 30, 37, and 45°C) and pH 6–9. Planktonic P. aeruginosa exhibited significantly longer lag phase and lower specific growth rates upon exposure to bacteriophages. Bacteriophages were subsequently applied to P. aeruginosa-enriched biofilm and were determined to lower the relative abundance of Pseudomonas-related taxa from 0.17 to 5.58% in controls to 0.01–0.61% in treated microbial communities. The relative abundance of Alphaproteobacteria, Pseudoalteromonas, and Planococcaceae decreased, possibly due to the phage-induced disruption of the biofilm matrix. Lastly, when applied to mitigate biofouling of ultrafiltration membranes, bacteriophages were determined to reduce the transmembrane pressure increase by 18% when utilized alone, and by 49% when used in combination with citric acid. The combined treatment was more effective compared with the citric acid treatment alone, which reported ca. 30% transmembrane pressure reduction. Collectively, the findings demonstrated that bacteriophages can be used as a biocidal agent to mitigate undesirable P. aeruginosa-associated problems in seawater applications.

  19. The hemocytes of Panstrogyllus Megistus (Hemiptera: Reduviidae Os hemócitos de Panstrongylus megistus (Hemiptera: Reduviidae

    Directory of Open Access Journals (Sweden)

    Margherita Anna Barracco

    1987-09-01

    Full Text Available Five hemocyte types were identified in the hemolymph of Panstrongylus megistus by phase contrast and common light microscopy using some histochemical methods. These are: Prohemocytes, small cells presenting a great nucleus/cytoplasm ratio; Plasmatocytes, the most numerous hemocytes, are polymorphic cells mainly characterized by a large amount of lysosomes; Granulocytes, hemocytes very similar to plasmatocytes which contain cytoplasmic granules and are especially rich in polysaccharides; Oenocytoids, cells presenting a small nucleus and a thick cytoplasm; they show many small round vacuoles when observed in Giemsa smears and many cytoplasmic granules under phase microscopy; Adipohemocytes, very large hemocytes, presenting many fat droplet inclusions which could correspond to free fat bodies which entered the hemolymph. Only prohemocytes and plasmatocytes can be clearly classified; all the other hemocyte types have a more ambiguous classification.Cinco tipos de hemócitos foram identificados na hemolinfa de Panstrongylus megistus através da microscopia de constraste de fase e de luz, usando alguns testes histoquímicos: Pró-hemócitos-células pequenas que mostram uma grande relação núcleocitoplasmática; Plasmotócitos-células polimporficas, que se caracterizam principalmente pela sua grande abundância em lisossomos - são os hemócitos mais numerosos; Granulócitos-células muito semelhantes aos plasmatócitos que contêm grânulos citoplasmáticos particularmente ricos em polissacarídeos; Enocitóides-hemócitos que apresentam um núcleo pequeno e cujo citoplasma basofílico revela-se muito denso e homogênio - mostram uma série de pequenos vacúolos esféricos quando observados nos esfregaços corados pelo Giemsa, mas a microscopia de fase revela uma grande quantidade de pequenos grânulo ao invés de vacúolos; Adipo-hemócitos-hemócitos muito grandes que contêm uma grande quantidade de inclusões lípicas - poderiam corresponder a

  20. Effect of dopamine injection on the hemocyte count and prophenoloxidase system of the white shrimp Litopenaeus vannamei

    Science.gov (United States)

    Pan, Luqing; Hu, Fawen; Zheng, Debin

    2011-09-01

    Effects of dopamine injection on the hemocyte count, phenoloxidase activity, serine proteinase activity, proteinase inhibitor activity and α2-macroglobulin-like activity in L. vannamei were studied. Results showed that dopamine injection resulted in a significant effect on the parameters measured ( P < 0.05), while no significant difference was observed in the control group (0.85% NaCl). In the experimental groups, the hemocyte count reached the minimum in 3 h; granular and semi-granular cells became stable after 12 h and hyaline cells and the total hemocyte count became stable after 18 h. Phenoloxidase activity reached the minimum in 6 h, and then became stable after 9 h. Serine protease activity and proteinase inhibitor activity reached the minimum in 3 h, and α2-macroglobulin-like activity reached the maximum in 3 h, and all the three parameters became stable after 12 h. The results suggest that the activating mechanisms of the proPO system triggered by dopamine are different from those triggered by invasive agents or spontaneously activated under a normal physical condition.

  1. Comparative experiments for tube agglutination test of pullorum antiserum with gamma ray Co/sup 60/ irradiated salmonella pullorum

    Energy Technology Data Exchange (ETDEWEB)

    Seo, B K [City Univ. of Seoul, Seoul (Republic of Korea)

    1976-01-01

    An agglutinability between naturally infected positive chicken serum of pullorum disease and hyperimmunized rabbit antiserum was compared. And the following results were obtained and summarized. On the agglutinability, Salmonella pullorum antigen which irradiated gamma-ray was better than another both formalized and heated antigen. Time of judgemented as positive titer in the tube agglutination test to the naturally infected positive chicken serum was it most suitable for 12 hours at 37/sup 0/C. Agglutination titer of positive immune chicken serum against gamma-ray irradiate Salmonella pullorum were as 320 approximately 640x.

  2. The secret life of the anthrax agent Bacillus anthracis: bacteriophage-mediated ecological adaptations.

    Directory of Open Access Journals (Sweden)

    Raymond Schuch

    2009-08-01

    Full Text Available Ecological and genetic factors that govern the occurrence and persistence of anthrax reservoirs in the environment are obscure. A central tenet, based on limited and often conflicting studies, has long held that growing or vegetative forms of Bacillus anthracis survive poorly outside the mammalian host and must sporulate to survive in the environment. Here, we present evidence of a more dynamic lifecycle, whereby interactions with bacterial viruses, or bacteriophages, elicit phenotypic alterations in B. anthracis and the emergence of infected derivatives, or lysogens, with dramatically altered survival capabilities. Using both laboratory and environmental B. anthracis strains, we show that lysogeny can block or promote sporulation depending on the phage, induce exopolysaccharide expression and biofilm formation, and enable the long-term colonization of both an artificial soil environment and the intestinal tract of the invertebrate redworm, Eisenia fetida. All of the B. anthracis lysogens existed in a pseudolysogenic-like state in both the soil and worm gut, shedding phages that could in turn infect non-lysogenic B. anthracis recipients and confer survival phenotypes in those environments. Finally, the mechanism behind several phenotypic changes was found to require phage-encoded bacterial sigma factors and the expression of at least one host-encoded protein predicted to be involved in the colonization of invertebrate intestines. The results here demonstrate that during its environmental phase, bacteriophages provide B. anthracis with alternatives to sporulation that involve the activation of soil-survival and endosymbiotic capabilities.

  3. Bacteriophages as indicators of faecal pollution and enteric virus removal.

    Science.gov (United States)

    McMinn, B R; Ashbolt, N J; Korajkic, A

    2017-07-01

    Bacteriophages are an attractive alternative to faecal indicator bacteria (FIB), particularly as surrogates of enteric virus fate and transport, due to their closer morphological and biological properties. Based on a review of published data, we summarize densities of coliphages (F+ and somatic), Bacteroides spp. and enterococci bacteriophages (phages) in individual human waste, raw wastewater, ambient fresh and marine waters and removal through wastewater treatment processes utilizing traditional treatments. We also provide comparisons with FIB and enteric viruses whenever possible. Lastly, we examine fate and transport characteristics in the aquatic environment and provide an overview of the environmental factors affecting their survival. In summary, concentrations of bacteriophages in various sources were consistently lower than FIB, but more reflective of infectious enteric virus levels. Overall, our investigation indicates that bacteriophages may be adequate viral surrogates, especially in built systems, such as wastewater treatment plants. Bacteriophage are alternative fecal indicators that may be better surrogates for viral pathogens than fecal indicator bacteria (FIB). This report offers a summary of the existing literature concerning the utility of bacteriophage as indicators of viral presence (fecal sources and surface waters) and persistence (in built infrastructure and aquatic environments). Our findings indicate that bacteriophage levels in all matrices examined are consistently lower than FIB, but similar to viral pathogens. Furthermore, in built infrastructure (e.g. wastewater treatment systems) bacteriophage closely mimic viral pathogen persistence suggesting they may be adequate sentinels of enteric virus removal. © 2017 The Society for Applied Microbiology.

  4. A bacteriophage endolysin that eliminates intracellular streptococci

    Science.gov (United States)

    Shen, Yang; Barros, Marilia; Vennemann, Tarek; Gallagher, D Travis; Yin, Yizhou; Linden, Sara B; Heselpoth, Ryan D; Spencer, Dennis J; Donovan, David M; Moult, John; Fischetti, Vincent A; Heinrich, Frank; Lösche, Mathias; Nelson, Daniel C

    2016-01-01

    PlyC, a bacteriophage-encoded endolysin, lyses Streptococcus pyogenes (Spy) on contact. Here, we demonstrate that PlyC is a potent agent for controlling intracellular Spy that often underlies refractory infections. We show that the PlyC holoenzyme, mediated by its PlyCB subunit, crosses epithelial cell membranes and clears intracellular Spy in a dose-dependent manner. Quantitative studies using model membranes establish that PlyCB interacts strongly with phosphatidylserine (PS), whereas its interaction with other lipids is weak, suggesting specificity for PS as its cellular receptor. Neutron reflection further substantiates that PlyC penetrates bilayers above a PS threshold concentration. Crystallography and docking studies identify key residues that mediate PlyCB–PS interactions, which are validated by site-directed mutagenesis. This is the first report that a native endolysin can traverse epithelial membranes, thus substantiating the potential of PlyC as an antimicrobial for Spy in the extracellular and intracellular milieu and as a scaffold for engineering other functionalities. DOI: http://dx.doi.org/10.7554/eLife.13152.001 PMID:26978792

  5. Isolation of lytic bacteriophage against Vibrio harveyi.

    Science.gov (United States)

    Crothers-Stomps, C; Høj, L; Bourne, D G; Hall, M R; Owens, L

    2010-05-01

    The isolation of lytic bacteriophage of Vibrio harveyi with potential for phage therapy of bacterial pathogens of phyllosoma larvae from the tropical rock lobster Panulirus ornatus. Water samples from discharge channels and grow-out ponds of a prawn farm in northeastern Australia were enriched for 24 h in a broth containing four V. harveyi strains. The bacteriophage-enriched filtrates were spotted onto bacterial lawns demonstrating that the bacteriophage host range for the samples included strains of V. harveyi, Vibrio campbellii, Vibrio rotiferianus, Vibrio parahaemolyticus and Vibrio proteolyticus. Bacteriophage were isolated from eight enriched samples through triple plaque purification. The host range of purified phage included V. harveyi, V. campbellii, V. rotiferianus and V. parahaemolyticus. Transmission electron microscope examination revealed that six purified phage belonged to the family Siphoviridae, whilst two belonged to the family Myoviridae. The Myoviridae appeared to induce bacteriocin production in a limited number of host bacterial strains, suggesting that they were lysogenic rather than lytic. A purified Siphoviridae phage could delay the entry of a broth culture of V. harveyi strain 12 into exponential growth, but could not prevent the overall growth of the bacterial strain. Bacteriophage with lytic activity against V. harveyi were isolated from prawn farm samples. Purified phage of the family Siphoviridae had a clear lytic ability and no apparent transducing properties, indicating they are appropriate for phage therapy. Phage resistance is potentially a major constraint to the use of phage therapy in aquaculture as bacteria are not completely eliminated. Phage therapy is emerging as a potential antibacterial agent that can be used to control pathogenic bacteria in aquaculture systems. The development of phage therapy for aquaculture requires initial isolation and determination of the bacteriophage host range, with subsequent creation of

  6. Sequence and comparative analysis of Leuconostoc dairy bacteriophages

    DEFF Research Database (Denmark)

    Kot, Witold; Hansen, Lars Henrik; Neve, Horst

    2014-01-01

    Bacteriophages attacking Leuconostoc species may significantly influence the quality of the final product. There is however limited knowledge of this group of phages in the literature. We have determined the complete genome sequences of nine Leuconostoc bacteriophages virulent to either Leuconostoc...

  7. [Functional morphology of blowfly Calliphora vicina hemocytes].

    Science.gov (United States)

    Kind, T V

    2012-01-01

    In the hemolymph of Calliphora seven types of hemocytes were revealed. These are prohemocytes, which are the stem cells, stable and unstable hyaline cells, thrombocytoids, spindle cells, juvenile plasmatocytes and plasmatocytes I-IV, which represent sequential stages of one cell line differentiation were registered. The margin between them is completion of the crop emptying and beginning of wandering stage. In the feeding and crop emptying larvae take place rising of hyaline cells, thrombocytoids and hyaline cells amount with parallel growth of their defense function. The second wave of hemogenesis occur in the end of crop emptying period. It is accompanied by burst of plasmatocyte I production with their subsequent differentiation to plasmatocytes II-IV. Production of stable hyaline cells and respectively prothrombocytoids may be regulated not only by hormonal background but also by inorganic or organic particles invaded into the hemocel. Three types of hemocytes are involved in loosing of hemolymph from alien particles, notably thrombocytoids, juvenile plasmatocytes and plasmatocytes I and II. Thrombocytoids are responsible for parasitic eggs encapsulation. In addition they can phagocytize tiny organic and inorganic particles. Juvenile plasmatocytes respond to alien invasion almost as quickly as thrombocytoids at the onset of invasion. Plasmatocytes I and II start phagocytosis more slowly, hours post invasion, frequently accumulating the particles previously catched by thrombocytoids. Plasmatocytes I can absorb foreign particles and group in morules and can also surround filled thrombocytoids forming distinctive capsules. Both morules and capsules are temporary structures and disintegrate some hours lately. It is supposed the existence of three levels of immune defence: the fast response reaction of thrombocytoids and juvenile plasmatocytes and slow cellular reactions of plasmatocytes I. They are prerequisites for more extensive humoral response.

  8. Bacteriophages of Yersinia pestis.

    Science.gov (United States)

    Zhao, Xiangna; Skurnik, Mikael

    2016-01-01

    Bacteriophage play many varied roles in microbial ecology and evolution. This chapter collates a vast body of knowledge and expertise on Yersinia pestis phages, including the history of their isolation and classical methods for their isolation and identification. The genomic diversity of Y. pestis phage and bacteriophage islands in the Y. pestis genome are also discussed because all phage research represents a branch of genetics. In addition, our knowledge of the receptors that are recognized by Y. pestis phage, advances in phage therapy for Y. pestis infections, the application of phage in the detection of Y. pestis, and clustered regularly interspaced short palindromic repeats (CRISPRs) sequences of Y. pestis from prophage DNA are all reviewed here.

  9. Bacteriophage-based synthetic biology for the study of infectious diseases

    Science.gov (United States)

    Lu, Timothy K.

    2014-01-01

    Since their discovery, bacteriophages have contributed enormously to our understanding of molecular biology as model systems. Furthermore, bacteriophages have provided many tools that have advanced the fields of genetic engineering and synthetic biology. Here, we discuss bacteriophage-based technologies and their application to the study of infectious diseases. New strategies for engineering genomes have the potential to accelerate the design of novel phages as therapies, diagnostics, and tools. Though almost a century has elapsed since their discovery, bacteriophages continue to have a major impact on modern biological sciences, especially with the growth of multidrug-resistant bacteria and interest in the microbiome. PMID:24997401

  10. In vivo effects of metaldehyde on Pacific oyster, Crassostrea gigas: comparing hemocyte parameters in two oyster families.

    Science.gov (United States)

    Moreau, Pierrick; Burgeot, Thierry; Renault, Tristan

    2015-06-01

    Pollutants via run-off into the ocean represent a potential threat to marine organisms, especially bivalves such as oysters living in coastal environments. These organisms filter large volumes of seawater and may accumulate contaminants within their tissues. Pesticide contamination in water could have a direct or indirect toxic action on tissues or cells and could induce alteration of immune system. Bivalve immunity is mainly supported by hemocytes and participates directly by phagocytosis to eliminate pathogens. Some studies have shown that pesticides can reduce immune defences and/or modify genomes in vertebrates and invertebrates. Metaldehyde is used to kill slugs, snails and other terrestrial gastropods. Although metaldehyde has been detected in surface waters, its effects on marine bivalves including the Pacific oyster, Crassostrea gigas, have never been studied. Given the mode of action of this molecule and its targets (molluscs), it could be potentially more toxic to oysters than other pesticides (herbicides, fungicides, insecticides, etc.). Effects of metaldehyde on oyster hemocyte parameters were thus monitored through in vivo experiments based on a short-term exposure. In this work, metaldehyde at 0.1 μg/L, which corresponds to an average concentration detected in the environment, modulated hemocyte activities of Pacific oysters after an in vivo short-term contact. Individuals belonging to two families showed different behaviours for some hemocyte activities after contamination by metaldehyde. These results suggested that effects of pollutants on oysters may differ from an individual to another in relation to genetic diversity. Finally, it appears essential to take an interest in the effects of metaldehyde on a wide variety of aquatic invertebrates including those that have a significant economic impact.

  11. Apoptosis of hemocytes from lions-paw scallop Nodipecten subnodosus induced with paralyzing shellfish poison from Gymnodinium catenatum.

    Science.gov (United States)

    Estrada, Norma; Ascencio, Felipe; Shoshani, Liora; Contreras, Rubén G

    2014-12-01

    The toxic dinoflagellate Gymnodinium catenatum produces paralyzing shellfish poisons (PSPs) that are consumed and accumulated by bivalves. Previously, we recorded a decrease in hemocytes 24h after injection of PSPs (gonyautoxin 2/3 epimers, GTX2/3) in the adductor muscle in the lions-paw scallop Nodipecten subnodosus. In this work, qualitative and quantitative analyses, in in vivo and in vitro experiments, revealed that the lower count of hemocytes results from cells undergoing typical apoptosis when exposed to GTX 2/3 epimers. This includes visible morphological alterations of the cytoplasmic membrane, damage to the nuclear membrane, condensation of chromatin, DNA fragmentation, and release of DNA fragments into the cytoplasm. Induction of apoptosis was accompanied by phosphatidylserine exposure to the outer cell membrane and activation of cysteine-aspartic proteases, caspase 3 and caspase 8. Addition of an inhibitor of caspase to the medium suppressed activation in hemocytes exposed to the toxins, suggesting that cell death was induced by a caspase-dependent apoptotic pathway. The results are important for future investigation of the scallop's immune system and should provide new insights into apoptotic processes in immune cells of scallops exposed to PSPs. Copyright © 2014 Elsevier GmbH. All rights reserved.

  12. Hemocyte parameters of the Pacific oyster Crassostrea gigas a year after the Hebei Spirit oil spill off the west coast of Korea

    Science.gov (United States)

    Donaghy, Ludovic; Hong, Hyun-Ki; Lee, Hee-Jung; Jun, Je-Cheon; Park, Young-Je; Choi, Kwang-Sik

    2010-12-01

    In marine bivalves, hemocytes support various physiological functions, including immune defense, nutrient transport, shell repair, and homeostatic maintenance. Although the effects of marine contaminants on the immunological functions of bivalves have been extensively investigated, the impacts of oil spills are not well understood. Therefore, we investigated hemocyte parameters in the Pacific oyster Crassostrea gigas 13 months after the Hebei Spirit oil spill (December 2007) off the west coast of Korea. The parameters studied included hemocyte concentration and mortality, relative proportion of hemocyte populations, and immunological functions such as phagocytosis and oxidative activity using flow cytometry. These immune-related parameters in oysters damaged by the oil spill were also compared to control oysters that were collected from an area unaffected by the spill. The flow cytometry study indicated that granulocyte population, phagocytic capacity, and reactive oxygen species production in oysters exposed to crude oil 13 months prior were depressed compared to the unexposed control oysters. Our data suggest that immunocompetence in oysters affected by the oil spill had not fully recovered 1 year after the accident, although more detailed studies on the physiology and disease resistance should be performed.

  13. Evolution and the complexity of bacteriophages.

    Science.gov (United States)

    Serwer, Philip

    2007-03-13

    The genomes of both long-genome (> 200 Kb) bacteriophages and long-genome eukaryotic viruses have cellular gene homologs whose selective advantage is not explained. These homologs add genomic and possibly biochemical complexity. Understanding their significance requires a definition of complexity that is more biochemically oriented than past empirically based definitions. Initially, I propose two biochemistry-oriented definitions of complexity: either decreased randomness or increased encoded information that does not serve immediate needs. Then, I make the assumption that these two definitions are equivalent. This assumption and recent data lead to the following four-part hypothesis that explains the presence of cellular gene homologs in long bacteriophage genomes and also provides a pathway for complexity increases in prokaryotic cells: (1) Prokaryotes underwent evolutionary increases in biochemical complexity after the eukaryote/prokaryote splits. (2) Some of the complexity increases occurred via multi-step, weak selection that was both protected from strong selection and accelerated by embedding evolving cellular genes in the genomes of bacteriophages and, presumably, also archaeal viruses (first tier selection). (3) The mechanisms for retaining cellular genes in viral genomes evolved under additional, longer-term selection that was stronger (second tier selection). (4) The second tier selection was based on increased access by prokaryotic cells to improved biochemical systems. This access was achieved when DNA transfer moved to prokaryotic cells both the more evolved genes and their more competitive and complex biochemical systems. I propose testing this hypothesis by controlled evolution in microbial communities to (1) determine the effects of deleting individual cellular gene homologs on the growth and evolution of long genome bacteriophages and hosts, (2) find the environmental conditions that select for the presence of cellular gene homologs, (3) determine

  14. Evolution and the complexity of bacteriophages

    Directory of Open Access Journals (Sweden)

    Serwer Philip

    2007-03-01

    Full Text Available Abstract Background The genomes of both long-genome (> 200 Kb bacteriophages and long-genome eukaryotic viruses have cellular gene homologs whose selective advantage is not explained. These homologs add genomic and possibly biochemical complexity. Understanding their significance requires a definition of complexity that is more biochemically oriented than past empirically based definitions. Hypothesis Initially, I propose two biochemistry-oriented definitions of complexity: either decreased randomness or increased encoded information that does not serve immediate needs. Then, I make the assumption that these two definitions are equivalent. This assumption and recent data lead to the following four-part hypothesis that explains the presence of cellular gene homologs in long bacteriophage genomes and also provides a pathway for complexity increases in prokaryotic cells: (1 Prokaryotes underwent evolutionary increases in biochemical complexity after the eukaryote/prokaryote splits. (2 Some of the complexity increases occurred via multi-step, weak selection that was both protected from strong selection and accelerated by embedding evolving cellular genes in the genomes of bacteriophages and, presumably, also archaeal viruses (first tier selection. (3 The mechanisms for retaining cellular genes in viral genomes evolved under additional, longer-term selection that was stronger (second tier selection. (4 The second tier selection was based on increased access by prokaryotic cells to improved biochemical systems. This access was achieved when DNA transfer moved to prokaryotic cells both the more evolved genes and their more competitive and complex biochemical systems. Testing the hypothesis I propose testing this hypothesis by controlled evolution in microbial communities to (1 determine the effects of deleting individual cellular gene homologs on the growth and evolution of long genome bacteriophages and hosts, (2 find the environmental conditions that

  15. Simulated hatchery system to assess bacteriophage efficacy against Vibrio harveyi.

    Science.gov (United States)

    Raghu Patil, J; Desai, Srividya Narayanamurthy; Roy, Panchali; Durgaiah, Murali; Saravanan, R Sanjeev; Vipra, Aradhana

    2014-12-02

    Vibriosis caused by luminous Vibrio harveyi commonly contributes to poor survival in shrimp hatcheries and aquaculture ponds. Lytic bacteriophages pathogenic for V. harveyi are currently being investigated as an alternative to antibiotics to prevent vibriosis. Here, 8 bacteriophages were isolated from oysters and clams using V. harveyi strains as baiting hosts. Among these bacteriophages, 1 strain (VHP6b) identified as broadly pathogenic for 27 V. harveyi strains examined was further characterized by electron microscopy and genome sequence analysis. Phage VHP6b possessed a tail and morphology consistent with it being a member of the family Siphoviridae, and its genome and proteome were most closely related to the Vibrio phages SSP02 and MAR10. An integrase gene essential for lysogeny was not evident. The ability of bacteriophage VHP6b to protect shrimp postlarvae against vibriosis caused by V. harveyi strain VH6 was demonstrated in a model system designed to simulate typical hatchery conditions. Bacteriophage treatment improved survival of postlarvae by 40 to 60% under these conditions, so therapies based on this or other bacteriophages may be useful in shrimp hatcheries.

  16. Characterization of myosin light chain in shrimp hemocytic phagocytosis.

    Science.gov (United States)

    Han, Fang; Wang, Zhiyong; Wang, Xiaoqing

    2010-11-01

    Myosin light chain, a well-known cytoskeleton gene, regulates multiple processes that are involved in material transport, muscle shrink and cell division. However, its function in phagocytosis against invading pathogens in crustacean remains unknown. In this investigation, a myosin light chain gene was obtained from Marsupenaeus japonicus shrimp. The full-length cDNA of this gene was of 766 bp and an open reading frame (ORF) of 462 bp encoding a polypeptide of 153 amino acids. The myosin light chain protein was expressed in Escherichia coli and purified. Subsequently the specific antibody was raised using the purified GST fusion protein. As revealed by immuno-electron microscopy, the myosin light chain protein was only expressed in the dark bands of muscle. In the present study, the myosin light chain gene was up-regulated in the WSSV-resistant shrimp as revealed by real-time PCR and western blot. And the phagocytic percentage and phagocytic index using FITC-labeled Vibrio parahemolyticus were remarkably increased in the WSSV-resistant shrimp, suggesting that the myosin light chain protein was essential in hemocytic phagocytosis. On the other hand, RNAi assays indicated that the phagocytic percentage and phagocytic index were significantly decreased when the myosin light chain gene was silenced by sequence-specific siRNA. These findings suggested that myosin light chain protein was involved in the regulation of hemocytic phagocytosis of shrimp. Copyright 2010 Elsevier Ltd. All rights reserved.

  17. Characteristic and functional analysis of toll-like receptors (TLRs in the lophotrocozoan, Crassostrea gigas, reveals ancient origin of TLR-mediated innate immunity.

    Directory of Open Access Journals (Sweden)

    Yang Zhang

    Full Text Available The evolution of TLR-mediated innate immunity is a fundamental question in immunology. Here, we report the characterization and functional analysis of four TLR members in the lophotrochozoans Crassostreagigas (CgTLRs. All CgTLRs bear a conserved domain organization and have a close relationship with TLRs in ancient non-vertebrate chordates. In HEK293 cells, every CgTLR could constitutively activate NF-κB responsive reporter, but none of the PAMPs tested could stimulate CgTLR-activated NF-κB induction. Subcellular localization showed that CgTLR members have similar and dual distribution on late endosomes and plasma membranes. Moreover, CgTLRs and CgMyD88 mRNA show a consistent response to multiple PAMP challenges in oyster hemocytes. As CgTLR-mediated NF-κB activation is dependent on CgMyD88, we designed a blocking peptide for CgTLR signaling that would inhibit CgTLR-CgMyD88 dependent NF-κB activation. This was used to demonstrate that a Vibrio parahaemolyticus infection-induced enhancement of degranulation and increase of cytokines TNF mRNA in hemocytes, could be inhibited by blocking CgTLR signaling. In summary, our study characterized the primitive TLRs in the lophotrocozoan C. gigas and demonstrated a fundamental role of TLR signaling in infection-induced hemocyte activation. This provides further evidence for an ancient origin of TLR-mediated innate immunity.

  18. T4 bacteriophage conjugated magnetic particles for E. coli capturing: Influence of bacteriophage loading, temperature and tryptone.

    Science.gov (United States)

    Liana, Ayu Ekajayanthi; Marquis, Christopher P; Gunawan, Cindy; Gooding, J Justin; Amal, Rose

    2017-03-01

    This work demonstrates the use of bacteriophage conjugated magnetic particles (Fe 3 O 4 ) for the rapid capturing and isolation of Escherichia coli. The investigation of T4 bacteriophage adsorption to silane functionalised Fe 3 O 4 with amine (NH 2 ), carboxylic (COOH) and methyl (CH 3 ) surface functional groups reveals the domination of net electrostatic and hydrophobic interactions in governing bacteriophage adsorption. The bare Fe 3 O 4 and Fe 3 O 4 -NH 2 with high T4 loading captured 3-fold more E. coli (∼70% capturing efficiency) compared to the low loading T4 on Fe 3 O 4 -COOH, suggesting the significance of T4 loading in E. coli capturing efficiency. Importantly, it is further revealed that E. coli capture is highly dependent on the incubation temperature and the presence of tryptone in the media. Effective E. coli capturing only occurs at 37°C in tryptone-containing media with the absence of either conditions resulted in poor bacteria capture. The incubation temperature dictates the capturing ability of Fe 3 O 4 /T4, whereby T4 and E. coli need to establish an irreversible binding that occurred at 37°C. The presence of tryptophan-rich tryptone in the suspending media was also critical, as shown by a 3-fold increase in E. coli capture efficiency of Fe 3 O 4 /T4 in tryptone-containing media compared to that in tryptone-free media. This highlights for the first time that successful bacteria capturing requires not only an optimum tailoring of the particle's surface physicochemical properties for favourable bacteriophage loading, but also an in-depth understanding of how factors, such as temperature and solution chemistry influence the subsequent bacteriophage-bacteria interactions. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Serotyping of Actinobacillus pleuropneumoniae serotype 5 strains using a monoclonal-based polystyrene agglutination test

    DEFF Research Database (Denmark)

    Dubreuil, J.D.; Letellier, A.; Stenbæk, Eva

    1996-01-01

    A polystyrene agglutination test has been developed for serotyping Actinobacillus pleuropneumoniae serotype 5a and 5b strains. Protein A-coated polystyrene microparticles were sensitized with a murine monoclonal antibody recognizing an epitope on serotype 5 LPS-O chain as shown by SDS-PAGE and We......A polystyrene agglutination test has been developed for serotyping Actinobacillus pleuropneumoniae serotype 5a and 5b strains. Protein A-coated polystyrene microparticles were sensitized with a murine monoclonal antibody recognizing an epitope on serotype 5 LPS-O chain as shown by SDS...... suspension of bacterial cells grown for 18 h. All A, pleuropneumoniae strains had been previously serotyped using standard procedures, The polystyrene agglutination test was rapid (less than 3 min) and easy to perform. Overall a very good correlation (97.3%) with the standard techniques was found...

  20. Comparison of agglutination test, microscopy and nPCR for ...

    African Journals Online (AJOL)

    action (nPCR) for the detection of T. gondii infection in mice (n=399) inoculated with heart .... Direct Agglutination Test (DAT) (Toxo screen DA, Biomerieux®, France) fol- ... 0.3 µl external forward Primer (50 µM), 0.3 µl external reverse Primer.

  1. Immune responses of mussel hemocyte subpopulations are differentially regulated by enzymes of the PI 3-K, PKC, and ERK kinase families.

    Science.gov (United States)

    García-García, Erick; Prado-Alvarez, Maria; Novoa, Beatriz; Figueras, Antonio; Rosales, Carlos

    2008-01-01

    Various hemocyte cell types have been described in invertebrates, but for most species a functional characterization of different hemocyte cell types is still lacking. In order to characterize some immunological properties of mussel (Mytilus galloprovincialis) hemocytes, cells were separated by flow cytometry and their capacity for phagocytosis, production of reactive oxygen species (ROS), and production of nitric oxide (NO), was examined. Phosphatidylinositol 3-kinase (PI 3-K), protein kinase C (PKC), and extracellular signal-regulated kinase (ERK) inhibitors were also used to biochemically characterize these cell responses. Four morphologically distinct subpopulations, designated R1-R4, were detected. R1, R2, and R3 cells presented different levels of phagocytosis towards zymosan, latex beads, and two bacteria species. Similarly, R1 to R3, but not R4, cells produced ROS, while all subpopulations produced NO, in response to zymosan. Internalization of all phagocytic targets was blocked by PI 3-K inhibition. In addition, internalization of latex particles, but not of bacteria, was partially blocked by PKC or ERK inhibition. Interestingly, phagocytosis of zymosan was impaired by PKC, or ERK inhibitors, only in R2 cells. Zymosan-induced ROS production was blocked by PI 3-K inhibition, but not by PKC, or ERK inhibition. In addition, zymosan-stimulated NO production was affected by PI 3-K inhibition in R1 and R2, but not in R3 or R4 cells. NO production in all cell types was unaffected by PKC inhibition, but ERK inhibition blocked it in R2 cells. These data reveal the existence of profound functional and biochemical differences in mussel hemocytes and indicate that M. galloprovincialis hemocytes are specialized cells fulfilling specific tasks in the context of host defense.

  2. The isolation and characterization of Campylobacter jejuni bacteriophages from free range and indoor poultry.

    Science.gov (United States)

    Owens, Jane; Barton, Mary D; Heuzenroeder, Michael W

    2013-02-22

    Six hundred and sixty one samples - primarily fresh chicken faeces - were processed to isolate wild type Campylobacter jejuni bacteriophages, via overlay agar methods using C. jejuni NCTC 12662. The aims of this study were to isolate and purify bacteriophages and then test for their ability to lyse field strains of C. jejuni in vitro. Of all samples processed, 130 were positive for bacteriophages. A distinct difference was observed between samples from different poultry enterprises. No bacteriophages could be isolated from indoor broilers. The majority of bacteriophages were isolated from free range poultry - both broilers and egg layers. Bacteriophages were purified and then selected for characterization based on their ability to produce clear lysis on plaque assay, as opposed to turbid plaques. Two hundred and forty one C. jejuni field isolates were tested for sensitivity to the bacteriophages. Lysis was graded subjectively and any minimal lysis was excluded. Using this system, 59.0% of the C. jejuni isolates showed significant sensitivity to at least one bacteriophage. The sensitivity to individual bacteriophages ranged from 10.0% to 32.5% of the C. jejuni isolates. Five bacteriophages were examined by electron microscopy and determined to belong to the Myoviridae family. The physical size, predicted genetic composition and genome size of the bacteriophages correlated well with other reported Campylobacter bacteriophages. The reasons for the observed difference between indoor broilers and free range poultry is unknown, but are postulated to be due to differences in the Campylobacter population in birds under different rearing conditions. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. Whole-genome sequence of the bacteriophage-sensitive strain Campylobacter jejuni NCTC12662

    DEFF Research Database (Denmark)

    Gencay, Yilmaz Emre; Sørensen, Martine C.H.; Brøndsted, Lone

    2017-01-01

    Campylobacter jejuni NCTC12662 has been the choice bacteriophage isolation strain due to its susceptibility to C. jejuni bacteriophages. This trait makes it a good candidate for studying bacteriophage-host interactions. We report here the whole-genome sequence of NCTC12662, allowing future...

  4. Effects of Copper on Hemocyte Apoptosis, ROS Production, and Gene Expression in White Shrimp Litopenaeus vannamei.

    Science.gov (United States)

    Guo, Hui; Li, Kexu; Wang, Wei; Wang, Chenggui; Shen, Yuchun

    2017-10-01

    Copper, a common chemical contaminant in aquatic environment, is known to be toxic to aquatic life at high concentrations. In the present study, we evaluated the apoptotic cell ratio and ROS production in hemocytes of the white shrimp Litopenaeus vannamei exposed to 1 or 5 mg L -1 Cu for 0, 3, 6, 12, 24, and 48 h. The expression changes of antioxidant biomarker genes, i.e., copper-zinc superoxide dismutase (Cu-Zn SOD) and catalase (CAT), apoptosis-related genes, i.e., caspase-3 and inhibitor of apoptosis protein (IAP), and a specific biomarker gene of heavy metal pollution, i.e., metallothionein (MT), were also determined in hemocytes. Significant increases in ROS production were observed in both treatment groups at each time points. The apoptotic cell ratios were significantly increased at 6-48 h among shrimp exposed to 1 mg L -1 Cu and at each time points in 5 mg L -1 Cu group. These results indicated that Cu would induce oxidative stress and apoptosis in the hemocyte of L. vannamei. Quantitative real-time PCR analysis revealed that the relative expression levels of Cu-Zn SOD, CAT, caspase-3, IAP, and MT were upregulated in a dose-dependent and time-dependent manner, suggesting the involvement of these genes in stress response against Cu exposure.

  5. Study of the reactivation of X-ray inactivated lambda bacteriophages by irradiated Escherichia coli bacteria

    International Nuclear Information System (INIS)

    Kiessling, W.

    1980-01-01

    Bacteriophages lambda and E.coli cells were exposed to X-rays in LB medium. Host cells exposed to a dose of 85 to 765 Gy had a reactivation factor 1.3 to 3.0 for bacteriophages inactivated by X-rays. The capacity of the bacteria for bacteriophage mutliplication remained apparently unchanged in this dose range. After UV-irradiation of the host cells, only a reactivation factor of 1.3 was found for bacteriophages exposed to X-radiation. The comparatively low Weigle reactivation of bacteriophages exposed to X-radiation - as compared with bacteriophages exposed to UV radiation was analyzed by counting free, non-adsorbed bacteriophages determined by filtration of radioactively labelled bacteriophage-host complexes, it was found to be due to a reduced adsorptivity. Reactivation experiments with bacteriophages exposed to X-rays and host bacterias with different degrees of radiosensitivity proved this assumption to be correct. (orig.) [de

  6. Potential of a lytic bacteriophage to disrupt Acinetobacter baumannii biofilms in vitro.

    Science.gov (United States)

    Liu, Yannan; Mi, Zhiqiang; Niu, Wenkai; An, Xiaoping; Yuan, Xin; Liu, Huiying; Wang, Yong; Feng, Yuzhong; Huang, Yong; Zhang, Xianglilan; Zhang, Zhiyi; Fan, Hang; Peng, Fan; Li, Puyuan; Tong, Yigang; Bai, Changqing

    2016-10-01

    The ability of Acinetobacter baumannii to form biofilms and develop antibiotic resistance makes it difficult to control infections caused by this bacterium. In this study, we explored the potential of a lytic bacteriophage to disrupt A. baumannii biofilms. The potential of the lytic bacteriophage to disrupt A. baumannii biofilms was assessed by performing electron microscopy, live/dead bacterial staining, crystal violet staining and by determining adenosine triphosphate release. The bacteriophage inhibited the formation of and disrupted preformed A. baumannii biofilms. Results of disinfection assay showed that the lytic bacteriophage lysed A. baumannii cells suspended in blood or grown on metal surfaces. These results suggest the potential of the lytic bacteriophage to disrupt A. baumannii biofilms.

  7. A simple system for in-droplet incubation and quantification of agglutination assays

    KAUST Repository

    Castro, David

    2013-10-28

    This work reports on a simple system for quantitative sensing of a target analyte based on agglutination in micro-channels. Functionalized microbeads and analyte with no prior incubation are flowed in droplets (~2μL) through a thin silicone tube filled with mineral oil at a flow rate of 150 μL/min. Hydrodynamic forces alone produce a highly efficient mixing of the beads within the droplet, without the need of complex mixing structures or magnetic actuation. The setup allows rapid observation of agglutination (<2 min), which is quantified using image analysis, and has potential application to high-throughput analysis.

  8. A simple system for in-droplet incubation and quantification of agglutination assays

    KAUST Repository

    Castro, David; Kodzius, Rimantas; Foulds, Ian G.

    2013-01-01

    This work reports on a simple system for quantitative sensing of a target analyte based on agglutination in micro-channels. Functionalized microbeads and analyte with no prior incubation are flowed in droplets (~2μL) through a thin silicone tube filled with mineral oil at a flow rate of 150 μL/min. Hydrodynamic forces alone produce a highly efficient mixing of the beads within the droplet, without the need of complex mixing structures or magnetic actuation. The setup allows rapid observation of agglutination (<2 min), which is quantified using image analysis, and has potential application to high-throughput analysis.

  9. Synthesis for Lunar Simulants: Glass, Agglutinate, Plagioclase, Breccia

    Science.gov (United States)

    Weinstein, Michael; Wilson, Stephen A.; Rickman, Douglas L.; Stoeser, Douglas

    2012-01-01

    The video describes a process for making glass for lunar regolith simulants that was developed from a patented glass-producing technology. Glass composition can be matched to simulant design and specification. Production of glass, pseudo agglutinates, plagioclase, and breccias is demonstrated. The system is capable of producing hundreds of kilograms of high quality glass and simulants per day.

  10. Ocean Acidification Affects the Cytoskeleton, Lysozymes, and Nitric Oxide of Hemocytes: A Possible Explanation for the Hampered Phagocytosis in Blood Clams, Tegillarca granosa.

    Science.gov (United States)

    Su, Wenhao; Rong, Jiahuan; Zha, Shanjie; Yan, Maocang; Fang, Jun; Liu, Guangxu

    2018-01-01

    An enormous amount of anthropogenic carbon dioxide (CO 2 ) has been dissolved into the ocean, leading to a lower pH and changes in the chemical properties of seawater, which has been termed ocean acidification (OA). The impacts of p CO 2 -driven acidification on immunity have been revealed recently in various marine organisms. However, the mechanism causing the reduction in phagocytosis still remains unclear. Therefore, the impacts of p CO 2 -driven OA at present and near-future levels (pH values of 8.1, 7.8, and 7.4) on the rate of phagocytosis, the abundance of cytoskeleton components, the levels of nitric oxide (NO), and the concentration and activity of lysozymes (LZM) of hemocytes were investigated in a commercial bivalve species, the blood clam ( Tegillarca granosa ). In addition, the effects of OA on the expression of genes regulating actin skeleton and nitric oxide synthesis 2 ( NOS2 ) were also analyzed. The results obtained showed that the phagocytic rate, cytoskeleton component abundance, concentration and activity of LZM of hemocytes were all significantly reduced after a 2-week exposure to the future OA scenario of a pH of 7.4. On the contrary, a remarkable increase in the concentration of NO compared to that of the control was detected in clams exposed to OA. Furthermore, the expression of genes regulating the actin cytoskeleton and NOS were significantly up-regulated after OA exposure. Though the mechanism causing phagocytosis seemed to be complicated based on the results obtained in the present study and those reported previously, our results suggested that OA may reduce the phagocytosis of hemocytes by (1) decreasing the abundance of cytoskeleton components and therefore hampering the cytoskeleton-mediated process of engulfment, (2) reducing the concentration and activity of LZM and therefore constraining the degradation of the engulfed pathogen through an oxygen-independent pathway, and (3) inducing the production of NO, which may negatively

  11. Reduction of Salmonella in ground chicken using a bacteriophage.

    Science.gov (United States)

    Grant, Ar'Quette; Parveen, Salina; Schwarz, Jurgen; Hashem, Fawzy; Vimini, Bob

    2017-08-01

    This study's goal was to ascertain the effectiveness of a commercially available Salmonella bacteriophage during ground chicken production focusing on: water source, different Salmonella serovars, and time. Salmonella-free boneless, skinless chicken meat was inoculated with 4.0 Log CFU/cm2 of either a cocktail of 3 Salmonella isolates derived from ground chicken (GC) or a cocktail of 3 Salmonella strains not isolated from ground chicken (non-GC). Bacteriophages were spread onto the chicken using sterile tap or filtered water for 30 min or 8 h. Salmonella was recovered using standard plating method. Greater Salmonella reduction was observed when the bacteriophage was diluted in sterile tap water than in sterile filtered water: 0.39 Log CFU/cm2 and 0.23 Log CFU/cm2 reduction after 30 min, respectively (P Salmonella's susceptibility to the bacteriophage, and treatment time. © 2017 Poultry Science Association Inc.

  12. Photodynamic Inactivation of Mammalian Viruses and Bacteriophages

    Directory of Open Access Journals (Sweden)

    Liliana Costa

    2012-06-01

    Full Text Available Photodynamic inactivation (PDI has been used to inactivate microorganisms through the use of photosensitizers. The inactivation of mammalian viruses and bacteriophages by photosensitization has been applied with success since the first decades of the last century. Due to the fact that mammalian viruses are known to pose a threat to public health and that bacteriophages are frequently used as models of mammalian viruses, it is important to know and understand the mechanisms and photodynamic procedures involved in their photoinactivation. The aim of this review is to (i summarize the main approaches developed until now for the photodynamic inactivation of bacteriophages and mammalian viruses and, (ii discuss and compare the present state of the art of mammalian viruses PDI with phage photoinactivation, with special focus on the most relevant mechanisms, molecular targets and factors affecting the viral inactivation process.

  13. Genome Sequences of 19 Novel Erwinia amylovora Bacteriophages.

    Science.gov (United States)

    Esplin, Ian N D; Berg, Jordan A; Sharma, Ruchira; Allen, Robert C; Arens, Daniel K; Ashcroft, Cody R; Bairett, Shannon R; Beatty, Nolan J; Bickmore, Madeline; Bloomfield, Travis J; Brady, T Scott; Bybee, Rachel N; Carter, John L; Choi, Minsey C; Duncan, Steven; Fajardo, Christopher P; Foy, Brayden B; Fuhriman, David A; Gibby, Paul D; Grossarth, Savannah E; Harbaugh, Kala; Harris, Natalie; Hilton, Jared A; Hurst, Emily; Hyde, Jonathan R; Ingersoll, Kayleigh; Jacobson, Caitlin M; James, Brady D; Jarvis, Todd M; Jaen-Anieves, Daniella; Jensen, Garrett L; Knabe, Bradley K; Kruger, Jared L; Merrill, Bryan D; Pape, Jenny A; Payne Anderson, Ashley M; Payne, David E; Peck, Malia D; Pollock, Samuel V; Putnam, Micah J; Ransom, Ethan K; Ririe, Devin B; Robinson, David M; Rogers, Spencer L; Russell, Kerri A; Schoenhals, Jonathan E; Shurtleff, Christopher A; Simister, Austin R; Smith, Hunter G; Stephenson, Michael B; Staley, Lyndsay A; Stettler, Jason M; Stratton, Mallorie L; Tateoka, Olivia B; Tatlow, P J; Taylor, Alexander S; Thompson, Suzanne E; Townsend, Michelle H; Thurgood, Trever L; Usher, Brittian K; Whitley, Kiara V; Ward, Andrew T; Ward, Megan E H; Webb, Charles J; Wienclaw, Trevor M; Williamson, Taryn L; Wells, Michael J; Wright, Cole K; Breakwell, Donald P; Hope, Sandra; Grose, Julianne H

    2017-11-16

    Erwinia amylovora is the causal agent of fire blight, a devastating disease affecting some plants of the Rosaceae family. We isolated bacteriophages from samples collected from infected apple and pear trees along the Wasatch Front in Utah. We announce 19 high-quality complete genome sequences of E. amylovora bacteriophages. Copyright © 2017 Esplin et al.

  14. Development of a fast agglutination screening test (FAST) for the detection of anti-Leishmania antibodies in dogs

    NARCIS (Netherlands)

    Schallig, H. D. F. H.; Schoone, G. J.; Beijer, E. G. M.; Kroon, C. C. M.; Hommers, M.; Ozbel, Y.; Ozensoy, S.; da Silva, E. S.; Cardoso, L. M.; da Silva, E. D.

    2002-01-01

    A fast agglutination screening test (FAST) for the detection of anti-Leishinania antibodies in serum samples from dogs with visceral leishmamosis was developed. The test is based on the direct agglutination test (DAT), but combines a higher parasite concentration with a smaller test volume. In

  15. Typhoid fever in a Tertiary Hospital in Nigeria: Another look at the Widal agglutination test as a preferred option for diagnosis.

    Science.gov (United States)

    Enabulele, Osahon; Awunor, Simeon Nyemike

    2016-01-01

    Single Widal agglutination test rather than blood culture, is commonly employed to diagnose typhoid fever in Nigeria. We took another look at the Widal agglutination test as a preferred option for diagnosis of typhoid fever by determining the specificity and sensitivity of Widal agglutination test in febrile adult patients. Two hundred and seventy-one blood samples from consecutive adults (>18 years) with febrile illness attending the General Practice Clinic of the University of Benin Teaching Hospital were tested using the Widal agglutination test, blood culture, and malaria parasite test on each sample to establish the diagnosis of typhoid fever. Of the 271 blood samples 124 (45.76%) were positive following a Widal agglutination test, 60 (22.10%) blood samples grew Salmonella organisms on blood culture while 55 (20.29%) blood samples showed a co-infection of typhoid fever and malaria. A sensitivity of 35%, specificity of 51%, positive predictive value of 17%, and a negative predictive value of 73% were observed for Widal agglutination test as a diagnostic modality for typhoid fever infection. A single Widal agglutination test is not a valid diagnostic option for typhoid fever while co-infection with malaria parasite is the preponderant microbiological finding in typhoid fever infections. The severity of malaria parasitemia is associated with positive titers on Widal test.

  16. Hemocyte responses of Dreissena polymorpha following a short-term in vivo exposure to titanium dioxide nanoparticles: Preliminary investigations

    Energy Technology Data Exchange (ETDEWEB)

    Couleau, Nicolas; Techer, Didier [Universite de Lorraine, Laboratoire des Interactions Ecotoxicologie, Biodiversite, Ecosystemes (LIEBE), CNRS UMR 7146, IUT Thionville-Yutz, Espace Cormontaigne, Yutz, F-57970 (France); Pagnout, Christophe [Universite de Lorraine, Laboratoire des Interactions Ecotoxicologie, Biodiversite, Ecosystemes (LIEBE), UMR 7146, Campus Bridoux, rue du General Delestraint, Metz, F-57070 (France); International Consortium for the Environmental Implications of Nanotechnology, iCEINT, http://www.i-ceint.org (France); Jomini, Stephane [Universite de Lorraine, Laboratoire des Interactions Ecotoxicologie, Biodiversite, Ecosystemes (LIEBE), UMR 7146, Campus Bridoux, rue du General Delestraint, Metz, F-57070 (France); Foucaud, Laurent; Laval-Gilly, Philippe; Falla, Jairo [Universite de Lorraine, Laboratoire des Interactions Ecotoxicologie, Biodiversite, Ecosystemes (LIEBE), CNRS UMR 7146, IUT Thionville-Yutz, Espace Cormontaigne, Yutz, F-57970 (France); Bennasroune, Amar, E-mail: amar.bennasroune@univ-metz.fr [Universite de Lorraine, Laboratoire des Interactions Ecotoxicologie, Biodiversite, Ecosystemes (LIEBE), CNRS UMR 7146, IUT Thionville-Yutz, Espace Cormontaigne, Yutz, F-57970 (France)

    2012-11-01

    The widespread use of titanium-based nanoparticles and their environmental release may pose a significant risk to aquatic organisms within freshwater ecosystems. Suspension-feeder invertebrates like bivalve molluscs represent a unique target group for nanoparticle toxicology. The aim of this work was to investigate the short-term responses of Dreissena polymorpha hemocytes after in vivo exposure to titanium dioxide nanoparticles (TiO{sub 2} NP). For this purpose, freshwater mussels were exposed to P25 TiO{sub 2} NP at the concentrations of 0.1, 1, 5 and 25 mg/L during 24 h. Viability, phagocytosis activity and mitogen activated protein kinase (MAPK) phosphorylation level of ERK 1/2 and p38 in hemocytes extracted from exposed mussels were compared to those from control specimens. Results demonstrated an inhibition of the phagocytosis activity after exposure to TiO{sub 2} NP at 0.1 and 1 mg/L. Similar trends, albeit less pronounced, were reported for higher concentrations of NP. Transmission electron microscopy showed for the first time the internalization of TiO{sub 2} NP into Dreissena polymorpha hemocytes. Besides, exposure to NP increased the ERK 1/2 phosphorylation levels in all treatments. Concerning the phosphorylation level of p38, only exposures to 5 and 25 mg/L of NP induced significant p38 activation in comparison to that of the control. Finally, these short-term effects observed at environmentally relevant concentrations highlighted the need for further studies concerning ecotoxicological evaluation of nanoparticle release into an aquatic environment. -- Highlights: Black-Right-Pointing-Pointer Phagocytosis inhibition at TiO{sub 2} NP exposure concentrations of 0.1 and 1 mg/L. Black-Right-Pointing-Pointer Internalization of TiO{sub 2} NP in freshwater mussel hemocytes. Black-Right-Pointing-Pointer Increased phosphorylation level of p38 and ERK 1/2 after in vivo exposure to TiO{sub 2} NP.

  17. Evaluation of Anti- Bacteriophage as Feed Additives to Prevent (SE in Broiler

    Directory of Open Access Journals (Sweden)

    K. H. Kim

    2013-03-01

    Full Text Available This experiment was conducted to evaluate anti-Salmonella enteritidis (anti-SE bacteriophage as feed additives to prevent Salmonella enteritidis in broilers. The experimental diets were formulated for 2 phases feeding trial, and 3 different levels (0.05, 0.1 and 0.2% of anti-SE bacteriophage were supplemented in basal diet. The basal diet was regarded as the control treatment. A total of 320 1-d-old male broilers (Ross 308 were allotted by randomized complete block (RCB design in 8 replicates with 10 chicks per pen. All birds were raised on rice hull bedding in ambient controlled environment and free access to feed and water. There were no significant differences in body weight gain, feed intake and feed conversion ratio (FCR at terminal period among treatments (p>0.05. Relative weights of liver, spleen, abdominal fat and tissue muscle of breast obtained from each anti-SE bacteriophage treatment were similar to control, with a slightly higher value in anti-SE bacteriophage 0.2%. In addition, a numerical difference of glutamic-oxaloacetic transaminase (GOT, glutamic-pyruvic transaminase (GPT and LDL cholesterol level was observed in the 0.2% anti-SE bacteriophage application even though blood profiles were not significantly affected by supplemented levels of anti-SE bacteriophage (p>0.05. In the result of a 14 d record after Salmonella enteritidis challenge of 160 birds from 4 previous treatments, mortality was linearly decreased with increasing anti-SE bacteriophage level (p<0.05, and Salmonella enteritidis concentration in the cecum was decreased with increasing levels of anti-SE bacteriophage (p<0.05. Based on the results of this study, it is considered that supplementation of 0.2% anti-SE bacteriophage may not cause any negative effect on growth, meat production, and it reduces mortality after Salmonella enteritidis challenge. These results imply to a possible use of anti-SE bacteriophage as an alternative feed additive instead of antibiotics

  18. Aligning the unalignable: bacteriophage whole genome alignments.

    Science.gov (United States)

    Bérard, Sèverine; Chateau, Annie; Pompidor, Nicolas; Guertin, Paul; Bergeron, Anne; Swenson, Krister M

    2016-01-13

    In recent years, many studies focused on the description and comparison of large sets of related bacteriophage genomes. Due to the peculiar mosaic structure of these genomes, few informative approaches for comparing whole genomes exist: dot plots diagrams give a mostly qualitative assessment of the similarity/dissimilarity between two or more genomes, and clustering techniques are used to classify genomes. Multiple alignments are conspicuously absent from this scene. Indeed, whole genome aligners interpret lack of similarity between sequences as an indication of rearrangements, insertions, or losses. This behavior makes them ill-prepared to align bacteriophage genomes, where even closely related strains can accomplish the same biological function with highly dissimilar sequences. In this paper, we propose a multiple alignment strategy that exploits functional collinearity shared by related strains of bacteriophages, and uses partial orders to capture mosaicism of sets of genomes. As classical alignments do, the computed alignments can be used to predict that genes have the same biological function, even in the absence of detectable similarity. The Alpha aligner implements these ideas in visual interactive displays, and is used to compute several examples of alignments of Staphylococcus aureus and Mycobacterium bacteriophages, involving up to 29 genomes. Using these datasets, we prove that Alpha alignments are at least as good as those computed by standard aligners. Comparison with the progressive Mauve aligner - which implements a partial order strategy, but whose alignments are linearized - shows a greatly improved interactive graphic display, while avoiding misalignments. Multiple alignments of whole bacteriophage genomes work, and will become an important conceptual and visual tool in comparative genomics of sets of related strains. A python implementation of Alpha, along with installation instructions for Ubuntu and OSX, is available on bitbucket (https://bitbucket.org/thekswenson/alpha).

  19. Comparison of Rose Bengal Plate Agglutination, Standard tube agglutination and Indirect ELISA tests for detection of Brucella antibodies in Cows and Buffaloes

    Directory of Open Access Journals (Sweden)

    S. N. Ghodasara

    2010-04-01

    Full Text Available A total of 180 serum samples (107 cows, 73 buffaloes from cases of abortion and various reproductive disorders were collected for detection of Brucella antibody by Rose Bengal Plate Agglutination Test (RBPT, Serum Tube Agglutination Test (STAT and indirect- ELISA (i-ELISA. The overall prevalence of brucellosis by RBPT, STAT and i-ELISA were 11.21%, 16.00% and 24.30% in cows 9.59%, 12.33% and 26.03% in buffaloes respectively. Overall seroprevalence of Brucellosis in cases of abortion, R.O.P. by RBPT, STAT and i-ELISA were 11.32%, 16.04% and 32.08% respectively. When three serological tests were compared, seropositivity was found highest by i-ELISA (25%, followed by STAT (14.45% and RBPT (10.56%. The results shows higher prevalence of brucellosis in cases of abortion and R.O.P., while at lower level from various reproductive disorders as detected serologically indicating endemicity of the infection in villages around Anand city, Gujarat. [Vet. World 2010; 3(2.000: 61-64

  20. Molecular characterization of a new efficiently transducing bacteriophage identified in meticillin-resistant Staphylococcus aureus.

    Science.gov (United States)

    Varga, Marian; Pantůček, Roman; Růžičková, Vladislava; Doškař, Jirˇí

    2016-01-01

    In Staphylococcus aureus, generalized transduction mediated by temperate bacteriophages represents a highly efficient way of transferring antibiotic resistance genes between strains. In the present study, we identified and characterized in detail a new efficiently transducing bacteriophage of the family Siphoviridae, designated ϕJB, which resides as a prophage in the meticillin-resistant S. aureus (MRSA) strain Jevons B. Whole-genome sequencing followed by detailed in silico analysis uncovered a linear dsDNA genome consisting of 43 ,12 bp and comprising 70 ORFs, of which ∼40 encoded proteins with unknown function. A global genome alignment of ϕJB and other efficiently transducing phages ϕ11, ϕ53, ϕ80, ϕ80α and ϕNM4 showed a high degree of homology with ϕNM4 and substantial differences with regard to other phages. Using a model transduction system with a well-defined donor and recipient, ϕJB transferred the tetracycline resistance plasmid pT181 and a penicillinase plasmid with outstanding frequencies, beating most of the above-mentioned phages by an order of magnitude. Moreover, ϕJB demonstrated high frequencies of transferring antibiotic resistance plasmids even upon induction from a lysogenic donor strain. Considering such transducing potential, ϕJB and related bacteriophages may serve as a suitable tool for elucidating the nature of transduction and its contribution to the spread of antibiotic resistance genes in naturally occurring MRSA populations.

  1. Bacteriophage cocktail for biocontrol of Salmonella in dried pet food.

    Science.gov (United States)

    Heyse, Serena; Hanna, Leigh Farris; Woolston, Joelle; Sulakvelidze, Alexander; Charbonneau, Duane

    2015-01-01

    Human salmonellosis has been associated with contaminated pet foods and treats. Therefore, there is interest in identifying novel approaches for reducing the risk of Salmonella contamination within pet food manufacturing environments. The use of lytic bacteriophages shows promise as a safe and effective way to mitigate Salmonella contamination in various food products. Bacteriophages are safe, natural, highly targeted antibacterial agents that specifically kill bacteria and can be targeted to kill food pathogens without affecting other microbiota. In this study, we show that a cocktail containing six bacteriophages had a broadspectrum activity in vitro against a library of 930 Salmonella enterica strains representing 44 known serovars. The cocktail was effective against 95% of the strains in this tested library. In liquid culture dose-ranging experiments, bacteriophage cocktail concentrations of ≥10(8) PFU/ml inactivated more than 90% of the Salmonella population (10(1) to 10(3) CFU/ml). Dried pet food inoculated with a mixture containing equal proportions of Salmonella serovars Enteritidis (ATCC 4931), Montevideo (ATCC 8387), Senftenberg (ATCC 8400), and Typhimurium (ATCC 13311) and then surface treated with the six-bacteriophage cocktail (≥2.5 ± 1.5 × 10(6) PFU/g) achieved a greater than 1-log (P contamination in samples taken from an undistributed lot of commercial dried dog food that tested positive for Salmonella. Our results indicate that bacteriophage biocontrol of S. enterica in dried pet food is technically feasible.

  2. Genotoxic and immunotoxic potential effects of selected psychotropic drugs and antibiotics on blue mussel (Mytilus edulis) hemocytes

    International Nuclear Information System (INIS)

    Lacaze, Emilie; Pédelucq, Julie; Fortier, Marlène; Brousseau, Pauline; Auffret, Michel; Budzinski, Hélène; Fournier, Michel

    2015-01-01

    The potential toxicity of pharmaceuticals towards aquatic invertebrates is still poorly understood and sometimes controversial. This study aims to document the in vitro genotoxicity and immunotoxicity of psychotropic drugs and antibiotics on Mytilus edulis. Mussel hemocytes were exposed to fluoxetine, paroxetine, venlafaxine, carbamazepine, sulfamethoxazole, trimethoprim and erythromycin, at concentrations ranging from μg/L to mg/L. Paroxetine at 1.5 μg/L led to DNA damage while the same concentration of venlafaxine caused immunomodulation. Fluoxetine exposure resulted in genotoxicity, immunotoxicity and cytotoxicity. In the case of antibiotics, trimethoprim was genotoxic at 200 μg/L and immunotoxic at 20 mg/L whereas erythromycin elicited same detrimental effects at higher concentrations. DNA metabolism seems to be a highly sensitive target for psychotropic drugs and antibiotics. Furthermore, these compounds affect the immune system of bivalves, with varying intensity. This attests the relevance of these endpoints to assess the toxic mode of action of pharmaceuticals in the aquatic environment. - Highlights: • Psychotropic drugs and antibiotics affect the immune system of Mytilus edulis. • Genotoxic and immunotoxic endpoints were relevant to assess pharmaceuticals toxicity. • DNA metabolism is a highly sensitive target for pharmaceuticals. • Fluoxetine and paroxetine were the most toxic compounds on mussel hemocytes. - Psychotropic drugs and antibiotics have the potential to cause immune toxicity and genotoxicity on Mytilus edulis hemocytes

  3. Bacteriophage-based Probiotic Preparation for Managing Shigella Infections

    Science.gov (United States)

    2015-04-16

    The preparation (designated “ShigActive”) is a bacteriophage cocktail that specifically targets Shigella spp. (significant diarrhea-causing pathogens...phages lytic for Shigella , and we have developed a murine model in which the in vivo efficacy of our 1. REPORT DATE (DD-MM-YYYY) 4. TITLE AND...10-Apr-2013 Approved for Public Release; Distribution Unlimited Final Report: Bacteriophage-based Probiotic Preparation for Managing Shigella

  4. Methods for initial characterization of Campylobacter jejuni bacteriophages

    DEFF Research Database (Denmark)

    Sørensen, Martine Camilla Holst; Gencay, Yilmaz Emre; Brøndsted, Lone

    2017-01-01

    Here we describe an initial characterization of Campylobacter jejuni bacteriophages by host range analysis, genome size determination by pulsed-field gel electrophoresis, and receptor-type identification by screening mutants for phage sensitivity.......Here we describe an initial characterization of Campylobacter jejuni bacteriophages by host range analysis, genome size determination by pulsed-field gel electrophoresis, and receptor-type identification by screening mutants for phage sensitivity....

  5. Latex agglutination test (LAT) for the diagnosis of typhoid fever.

    Science.gov (United States)

    Sahni, Gopal Shankar

    2013-06-01

    The efficacy of latex agglutination test in the rapid diagnosis of typhoid fever was studied and the result compared with that of blood culture. This study included 80 children suffering from typhoid fever, among which 40 were confirmed by blood culture isolation and 40 had possible typhoid fever based on high Widal's titre (a four-fold rise in the titre of antibody to typhi "O" and "H" antigen was considered as a positive Widal's test result). Eighty children, 40 with febrile illness confirmed to be other than typhoid and 40 normal healthy children were used as negative controls. The various groups were: (i) Study group ie, group I had 40 children confirmed by culture isolation of Salmonella typhi(confirmed typhoid cases). (ii) Control groups ie, (a) group II with 40 febrile controls selected from paediatrics ward where cause other than S typhi has been established, (b) group III with 40 afebrile healthy controls that were siblings of the children admitted in paediatric ward for any reason with no history of fever and TAB vaccination in the last one year, and (c) group IV with 40 children with high Widal's titre in paired sera sample. Widal's test with paired sera with a one week interval between collections were done in all 40 patients. Latex aggtutination test which could detect 900 ng/ml of antigen as observed in checker board titration, was positive in all 40 children from group I who had positive blood culture and in 30 children from group IV who had culture negative and had high Widal's titre positive. Latex agglutination test was positive in 4 children in group II and none in group III. Using blood culture positive cases as true positive and children in groups II and III as true negative, the test had a sensitivity of 100% and specificity of 96%. Latex agglutination test was found to be significantly sensitive (100%) and specific (96%) and could detect 75% more cases in group IV (possible typhoid cases). Thus latex agglutination test can be used for rapid

  6. Evaluation of radiosensitivity hemocytes of Biomphalaria glabrata exposed to gamma radiation

    International Nuclear Information System (INIS)

    Silva, L.R.S.; Amaral, A.J.; Silva, E.B.; Amancio, F.F.; Melo, A.M.M.A.

    2013-01-01

    The mollusc Biomphalaria glabrata have characteristics that allow them to be identified as an animal model ideal for monitoring areas exposed to chemical agents and physical. This study evaluated the effect of ionizing radiation from Cobalt-60 in haemocytes present in the hemolymph of Biomphalaria glabrata, with the goal of using these cells as indicators of the presence of radiation in aquatic environments. The mollusks were divided into five groups: one control and four subjected doses of 25, 35, 45 and 55 Gy of gamma radiation. After 48 hours of irradiation, the clam hemolymph was collected and slides were prepared and stained with Giemsa for analyses under a light microscope. Statistical analysis was performed using ANOVA and Tukey's test, p <0.05. The results showed that the total number of cells after irradiation reduced compared to control except at a dose of 55 Gy. During data analysis, morphological changes were observed in haemocytes of mollusks subjected to doses of 35, 45 and 55 Gy. These modifications consisted of nucleus bilobulated and nucleo plasmatic bridges. Another change was exclusively observed in the cellular exposure of 55 Gy, where hemocytes showed misshapen nuclei and cytoplasm vacuolisation, suggestive of apoptosis. It is concluded that hemocytes are sensitive to radiation and can be used as indicators of the presence of high doses of ionizing radiation in aquatic environments. (author)

  7. Ultrastructural studies of the hemocytes of Panstrongylus megistus (Hemiptera: Reduvidae Ultra-estrutura dos hemócitos de Panstrongylus megistus (Hemiptera: Reduviidae

    Directory of Open Access Journals (Sweden)

    Margherita A. Barracco

    1989-06-01

    Full Text Available Ultrastructural analyses revealed the presence of six hemocyte types in the hemolymph of Panstrogylus megistus, partially confirming our previous results obtained through light microscopy. Prohemocytes: small, round hemocytes with a thin cytoplasm layer, espcieally rich in free ribosomes and poor in membranous systems. Plasmatocytes: polymorphic cells, whose cytoplasm contains many lysosomes and a well developed rough endoplasmic reticulum (RER.They are extremely phagocytic. Sometimes, they show a large vacuolation. Granulocytes: granular hemocytes whose granules show different degrees of electrondensity. Most of them, have an internal structuration. Coagulocytes: oval or elongated hemocytes, which show pronounced perinuclear cisternae as normally observed in coagulocytes. The cytoplasm is usually electrondense, poor in membranous systems and contains many labile granules. Oenocytoids: large and very stable hemocytes, whose homogeneous cytoplasme is rich in loose ribosomes and poor in membranous systems. Adipohemocytes: large cells, containing several characteristic lipid droplets. The cytoplasm is also rich in glycogen, RER and large mitochondria. The total and differential hemocyte count (THC and DHC were also calculated for this reduviid. THC increases from 2,900 hemocytes/cubic millimeter of hemolymph in the 4th intar to 4,350 in the 5th and then, decreases to 1,950 in the adults. Plasmatocytes and coagulocytes are the predominant hemocyte types.Estudos ao microscópio eletrônico de transmissão revelaram a presença de seis tipos de hemócitos na hemolinfa de Panstrongylus megistus. Estes resultados confirmam parcialmente os obtidos anteriormente através da microscopia de luz. Pró-hemócitos: células pequenas e arredondadas, cuja delgada faixa citoplasmática é especialmente rica em ribossomos livres e pobre em sistemas membranosos. Plasmatócitos: células polimórficas, cujo citoplasma caracteriza-se por um retículo endoplasm

  8. Rapid detection of microalbuminuria in diabetic patients by an agglutination inhibition test

    International Nuclear Information System (INIS)

    Giampietro, O.; Miccoli, R.; Di Palma, L.; Bertolotto, A.; Anichini, R.; Navalesi, R.; Clerico, A.

    1986-01-01

    Subclinical elevation of urinary albumin excretion (UAlbE) early in the course of diabetes mellitus has been suggested to predict later clinical proteinuria and mortality. UAlbE is currently measured using radioimmunoassay (RIA) or radial immunodiffusion methods. However, these procedures are expensive and time-consuming and cannot be used as screening methods. Recently, an agglutination test (AT) has been suggested as a routinary method for the screening of microalbuminuria in diabetic patients. In this paper the results obtained are compared with an AT procedure and RIA method in a screening program of microproteinuria in diabetic patients. An immunological test (a latex agglutination assay) for the analysis of albuminura is used, which human albumin was adsorbed to latex beads (about 0.3 μl of a urine sample. Urine samples containing an albumin concentration >40 μg/ml were found to inhibit the agglutination of latex beads with antiserum. The RIA and AT results showed good agreement when urine samples were assayed soon after collection or after a short period of storage (≤3 weeks at -20 grade centigrades). The AT procedure has been adjusted in order to give a positive response (no agglutination) over the range of supranormal concentrations of urinary albumin (>40 μg/ml), which are on the other hand undetectable by Albustix. In addition, it is possible to perform a semiquantitative test using various dilutions of urine samples with albumin concentration > 40 μg/ml, so to estimate approximately the UAlbE. The AT method is simple, fast and specific, and has proved to be useful for the identification of diabetic patients at risk for developing clinical nephropathy. Therefore, it may be used in screening programs for diabetic microproteinuria

  9. Genetic diversity among five T4-like bacteriophages

    Directory of Open Access Journals (Sweden)

    Bertrand Claire

    2006-05-01

    Full Text Available Abstract Background Bacteriophages are an important repository of genetic diversity. As one of the major constituents of terrestrial biomass, they exert profound effects on the earth's ecology and microbial evolution by mediating horizontal gene transfer between bacteria and controlling their growth. Only limited genomic sequence data are currently available for phages but even this reveals an overwhelming diversity in their gene sequences and genomes. The contribution of the T4-like phages to this overall phage diversity is difficult to assess, since only a few examples of complete genome sequence exist for these phages. Our analysis of five T4-like genomes represents half of the known T4-like genomes in GenBank. Results Here, we have examined in detail the genetic diversity of the genomes of five relatives of bacteriophage T4: the Escherichia coli phages RB43, RB49 and RB69, the Aeromonas salmonicida phage 44RR2.8t (or 44RR and the Aeromonas hydrophila phage Aeh1. Our data define a core set of conserved genes common to these genomes as well as hundreds of additional open reading frames (ORFs that are nonconserved. Although some of these ORFs resemble known genes from bacterial hosts or other phages, most show no significant similarity to any known sequence in the databases. The five genomes analyzed here all have similarities in gene regulation to T4. Sequence motifs resembling T4 early and late consensus promoters were observed in all five genomes. In contrast, only two of these genomes, RB69 and 44RR, showed similarities to T4 middle-mode promoter sequences and to the T4 motA gene product required for their recognition. In addition, we observed that each phage differed in the number and assortment of putative genes encoding host-like metabolic enzymes, tRNA species, and homing endonucleases. Conclusion Our observations suggest that evolution of the T4-like phages has drawn on a highly diverged pool of genes in the microbial world. The T4

  10. Genomics of three new bacteriophages useful in the biocontrol of Salmonella

    Directory of Open Access Journals (Sweden)

    Carlota eBardina

    2016-04-01

    Full Text Available Non-typhoid Salmonella is the principal pathogen related to food-borne diseases throughout the world. Widespread antibiotic resistance has adversely affected human health and has encouraged the search for alternative antimicrobial agents. The advances in bacteriophage therapy highlight their use in controlling a broad spectrum of food-borne pathogens. One requirement for the use of bacteriophages as antibacterials is the characterization of their genomes. In this work, complete genome sequencing and molecular analyses were carried out for three new virulent Salmonella-specific bacteriophages (UAB_Phi20, UAB_Phi78, and UAB_Phi87 able to infect a broad range of Salmonella strains. Sequence analysis of the genomes of UAB_Phi20, UAB_Phi78, and UAB_Phi87 bacteriophages did not evidence the presence of known virulence-associated and antibiotic resistance genes, and potential immunoreactive food allergens. The UAB_Phi20 genome comprised 41,809 base pairs with 80 open reading frames (ORFs; 24 of them with assigned function. Genome sequence showed a high homology of UAB_Phi20 with Salmonella bacteriophage P22 and other P22likeviruses genus of the Podoviridae family, including ST64T and ST104. The DNA of UAB_Phi78 contained 44,110 bp including direct terminal repeats of 179 bp and 58 putative ORFs were predicted and 20 were assigned function. This bacteriophage was assigned to the SP6likeviruses genus of the Podoviridae family based on its high similarity not only with SP6 but also with the K1-5, K1E, and K1F bacteriophages, all of which infect Escherichia coli. The UAB_Phi87 genome sequence consisted of 87,669 bp with terminal direct repeats of 608 bp; although 148 ORFs were identified, putative functions could be assigned to only 29 of them. Sequence comparisons revealed the mosaic structure of UAB_Phi87 and its high similarity with bacteriophages Felix O1 and wV8 of E. coli with respect to genetic content and functional organization. Phylogenetic

  11. Antibacterial Efficacy of Lytic Bacteriophages against Antibiotic-Resistant Klebsiella Species

    Directory of Open Access Journals (Sweden)

    M. Khajeh Karamoddini

    2011-01-01

    Full Text Available Bacterial resistance to antibiotics is a leading and highly prevalent problem in the treatment of infectious diseases. Bacteriophages (phages appear to be effective and safe alternatives for the treatment of resistant infections because of their specificity for bacterial species and lack of infectivity in eukaryotic cells. The present study aimed to isolate bacteriophages against Klebsiella spp. and evaluate their efficacy against antibiotic-resistant species. Seventy-two antibiotic-resistant Klebsiella spp. were isolated from samples of patients who referred to the Ghaem Hospital (Mashhad, Iran. Lytic bacteriophages against Klebsiella spp. were isolated from wastewater of the septic tank of the same hospital. Bactericidal activity of phages against resistant Klebsiella spp. was tested in both liquid (tube method; after 1 and 24 h of incubation and solid (double-layer agar plate method; after 24 h of incubation phases. In each method, three different concentrations of bacteriophages (low: 107 PFU/mL were used. Bacteriophages showed promising bactericidal activity at all assessed concentrations, regardless of the test method and duration of incubation. Overall, bactericidal effects were augmented at higher concentrations. In the tube method, higher activity was observed after 24 h of incubation compared to the 1-h incubation. The bactericidal effects were also higher in the tube method compared to the double-layer agar plate method after 24 h of incubation. The findings of the present study suggest that bacteriophages possess effective bactericidal activity against resistant Klebsiella spp. These bactericidal activities are influenced by phage concentration, duration of incubation, and test method.

  12. A hemocyte gene expression signature correlated with predictive capacity of oysters to survive Vibrio infections

    Directory of Open Access Journals (Sweden)

    Rosa Rafael

    2012-06-01

    Full Text Available Abstract Background The complex balance between environmental and host factors is an important determinant of susceptibility to infection. Disturbances of this equilibrium may result in multifactorial diseases as illustrated by the summer mortality syndrome, a worldwide and complex phenomenon that affects the oysters, Crassostrea gigas. The summer mortality syndrome reveals a physiological intolerance making this oyster species susceptible to diseases. Exploration of genetic basis governing the oyster resistance or susceptibility to infections is thus a major goal for understanding field mortality events. In this context, we used high-throughput genomic approaches to identify genetic traits that may characterize inherent survival capacities in C. gigas. Results Using digital gene expression (DGE, we analyzed the transcriptomes of hemocytes (immunocompetent cells of oysters able or not able to survive infections by Vibrio species shown to be involved in summer mortalities. Hemocytes were nonlethally collected from oysters before Vibrio experimental infection, and two DGE libraries were generated from individuals that survived or did not survive. Exploration of DGE data and microfluidic qPCR analyses at individual level showed an extraordinary polymorphism in gene expressions, but also a set of hemocyte-expressed genes whose basal mRNA levels discriminate oyster capacity to survive infections by the pathogenic V. splendidus LGP32. Finally, we identified a signature of 14 genes that predicted oyster survival capacity. Their expressions are likely driven by distinct transcriptional regulation processes associated or not associated to gene copy number variation (CNV. Conclusions We provide here for the first time in oyster a gene expression survival signature that represents a useful tool for understanding mortality events and for assessing genetic traits of interest for disease resistance selection programs.

  13. Bacteriophages in the control of pathogenic vibrios

    DEFF Research Database (Denmark)

    Plaza, Nicolás; Castillo Bermúdez, Daniel Elías; Perez-Reytor, Diliana

    2018-01-01

    constitute a continuing threat for aquaculture. Moreover, the continuous use of antibiotics has been accompanied by an emergence of antibiotic resistance in Vibrio species, implying a necessity for efficient treatments. One promising alternative that emerges is the use of lytic bacteriophages; however......, there are some drawbacks that should be overcome to make phage therapy a widely accepted method. In this work, we discuss about the major pathogenic Vibrio species and the progress, benefits and disadvantages that have been detected during the experimental use of bacteriophages to their control....

  14. Determination of degree of RBC agglutination for blood typing using a small quantity of blood sample in a microfluidic system.

    Science.gov (United States)

    Chang, Yaw-Jen; Ho, Ching-Yuan; Zhou, Xin-Miao; Yen, Hsiu-Rong

    2018-04-15

    Blood typing assay is a critical test to ensure the serological compatibility of a donor and an intended recipient prior to a blood transfusion. This paper presents a microfluidic blood typing system using a small quantity of blood sample to determine the degree of agglutination of red blood cell (RBC). Two measuring methods were proposed: impedimetric measurement and electroanalytical measurement. The charge transfer resistance in the impedimetric measurement and the power parameter in the electroanalytical measurement were used for the analysis of agglutination level. From the experimental results, both measuring methods provide quantitative results, and the parameters are linearly and monotonically related to the degree of RBC agglutination. However, the electroanalytical measurement is more reliable than the impedimetric technique because the impedimetric measurement may suffer from many influencing factors, such as chip conditions. Five levels from non-agglutination (level 0) to strong agglutination (level 4+) can be discriminated in this study, conforming to the clinical requirement to prevent any risks in transfusion. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Multixenobiotic resistance in Mytilus edulis: Molecular and functional characterization of an ABCG2- type transporter in hemocytes and gills.

    Science.gov (United States)

    Ben Cheikh, Yosra; Xuereb, Benoit; Boulangé-Lecomte, Céline; Le Foll, Frank

    2018-02-01

    Among the cellular protection arsenal, ABC transporters play an important role in xenobiotic efflux in marine organisms. Two pumps belonging to B and C subfamily has been identified in Mytilus edulis. In this study, we investigated the presence of the third major subtype ABCG2/BCRP protein in mussel tissues. Transcript was expressed in hemocytes and with higher level in gills. Molecular characterization revealed that mussel ABCG2 transporter shares the sequence and organizational structure with mammalian and molluscan orthologs. Overall identity of the predicted amino acid sequence with corresponding homologs from other organisms was between 49% and 98%. Moreover, protein efflux activity was demonstrated using a combination of fluorescent allocrites and specific inhibitors. The accumulation of bodipy prazosin and pheophorbide A was heterogeneous in gills and hemocytes. Most of the used blockers enhanced probe accumulation at different levels, most significantly for bodipy prazosin. Moreover, Mrp classical blocker MK571 showed a polyspecificity. In conclusion, our data demonstrate that several ABC transporters contribute to MXR phenotype in the blue mussel including ABCG2 that forms an active pump in hemocytes and gills. Efforts are needed to distinguish between the different members and to explore their single function and specificity towards allocrites and chemosensitizers. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Bacteriophages: The viruses for all seasons of molecular biology

    Directory of Open Access Journals (Sweden)

    Karam Jim D

    2005-03-01

    Full Text Available Abstract Bacteriophage research continues to break new ground in our understanding of the basic molecular mechanisms of gene action and biological structure. The abundance of bacteriophages in nature and the diversity of their genomes are two reasons why phage research brims with excitement. The pages of Virology Journal will reflect the excitement of the "New Phage Biology."

  17. Improved bacteriophage genome data is necessary for integrating viral and bacterial ecology.

    Science.gov (United States)

    Bibby, Kyle

    2014-02-01

    The recent rise in "omics"-enabled approaches has lead to improved understanding in many areas of microbial ecology. However, despite the importance that viruses play in a broad microbial ecology context, viral ecology remains largely not integrated into high-throughput microbial ecology studies. A fundamental hindrance to the integration of viral ecology into omics-enabled microbial ecology studies is the lack of suitable reference bacteriophage genomes in reference databases-currently, only 0.001% of bacteriophage diversity is represented in genome sequence databases. This commentary serves to highlight this issue and to promote bacteriophage genome sequencing as a valuable scientific undertaking to both better understand bacteriophage diversity and move towards a more holistic view of microbial ecology.

  18. Isolating E.Coli Bacteriophage from Raw Sewage and Determining its Selectivity to the Host Cell

    Directory of Open Access Journals (Sweden)

    SM Imeni

    2016-05-01

    Full Text Available Introduction: Bacteriophages are viruses that infect and destroy prokaryote cells, specifically the bacteria. They act too selective, so as each bacteriophage affects only on specific type of bacteria. Due to their specific features, bacteriophages can be used as an appropriate substitute for antibiotics in infectious diseases treatment. Therefore, this study aimed to isolate E. coli-specific bacteriophage from raw sewage. Methods: Eight samples of raw sewage, each containing approximately 50 ml of raw sewage with 10 minute gap, were prepared from Zargandeh wastewater treatment plant, Tehran, Iran. The sewages were mixed with Brain-heart infusion medium (BHI as a liquid culture medium in order to let the microorganisms grow. Incubation, purification and determination of bacteria were followed repeatedly to isolate the bacteriophage. Then it was tested on E.coli (ATCC 25922, Enterococcus faecalis (ATCC 19433, Staphylococcus aureus (ATCC 2392, and Yersinia enterocolitica (ATCC 9610 in order to determine the bacteriophage selectivity. Results: The E.coli bacteriophages were successfully isolated from all the eight samples, that were completely able to lyse and destroy E.coli bacterial cells, though no effect was observed on other types of bacteria. Conclusion: The study findings revealed that bacteriophages act selectively. Considering the raise of antibiotic resistance in the world, bacteriophages can serve as a good substitute for antibiotics in treating infectious diseases.

  19. Two-Phase Microfluidic Systems for High Throughput Quantification of Agglutination Assays

    KAUST Repository

    Castro, David

    2018-01-01

    assay, with a minimum detection limit of 50 ng/mL using optical image analysis. We compare optical image analysis and light scattering as quantification methods, and demonstrate the first light scattering quantification of agglutination assays in a two

  20. Bacteriophages for detection of bacterial pathogens

    International Nuclear Information System (INIS)

    Kutateladze, M.

    2009-01-01

    The G. Eliava Institute of Bacteriophages, Microbiology and Virology (Tbilisi, Georgia) is one of the most famous institutions focused on bacteriophage research for the elaboration of appropriate phage methodologies for human and animal protection. The main direction of the institute is the study and production of bacteriophages against intestinal disorders (dysentery, typhoid, intesti) and purulent-septic infections (staphylococcus, streptococcus, pyophage, etc.). These preparations were successfully introduced during the Soviet era, and for decades were used throughout the former Soviet Union and in other Socialist countries for the treatment, prophylaxis, and diagnosis of various infectious diseases, including those caused by antibiotic-resistant bacterial strains. Bacteriophages were widely used for identifying and detecting infections caused by the most dangerous pathogens and causative agents of epidemiological outbreaks. The specific topic of this presentation is the phage typing of bacterial species, which can be an important method for epidemiological diagnostics. Together with different genetic methodologies - such as PCR-based methods, PFGE, plasmid fingerprinting, and ribosomal typing - phage typing is one method for identifying bacterial pathogens. The method has a high percentage of determination of phage types, high specificity of reaction, and is easy for interpretation and use by health workers. Phage typing was applied for inter-species differentiation of different species of Salmonella, S. typhi, Brucella spp, Staphylococcus aureus, E. col,i Clostridium deficile, Vibrio cholerae, Yersinia pestis, Yersinia enterocolitica, Lysteria monocytogenes, Clostridium perfringens, Clostridium tetani, plant pathogens, and other bacterial pathogens. In addition to addressing the utility and efficacy of phage typing, the paper will discuss the isolation and selection of diagnostic typing phages for interspecies differentiation of pathogens that is necessary

  1. Application of bacteriophages in post-harvest control of human pathogenic and food spoiling bacteria.

    Science.gov (United States)

    Pérez Pulido, Rubén; Grande Burgos, Maria José; Gálvez, Antonio; Lucas López, Rosario

    2016-10-01

    Bacteriophages have attracted great attention for application in food biopreservation. Lytic bacteriophages specific for human pathogenic bacteria can be isolated from natural sources such as animal feces or industrial wastes where the target bacteria inhabit. Lytic bacteriophages have been tested in different food systems for inactivation of main food-borne pathogens including Listeria monocytogenes, Staphylococcus aureus, Escherichia coli O157:H7, Salmonella enterica, Shigella spp., Campylobacter jejuni and Cronobacter sakazkii, and also for control of spoilage bacteria. Application of lytic bacteriophages could selectively control host populations of concern without interfering with the remaining food microbiota. Bacteriophages could also be applied for inactivation of bacteria attached to food contact surfaces or grown as biofilms. Bacteriophages may receive a generally recognized as safe status based on their lack of toxicity and other detrimental effects to human health. Phage preparations specific for L. monocytogenes, E. coli O157:H7 and S. enterica serotypes have been commercialized and approved for application in foods or as part of surface decontamination protocols. Phage endolysins have a broader host specificity compared to lytic bacteriophages. Cloned endolysins could be used as natural preservatives, singly or in combination with other antimicrobials such as bacteriocins.

  2. Silkworm (Bombyx mori) hemocytes do not produce reactive oxygen metabolites as a part of defense mechanisms

    Czech Academy of Sciences Publication Activity Database

    Hyršl, P.; Číž, Milan; Kubala, Lukáš; Lojek, Antonín

    2004-01-01

    Roč. 49, č. 3 (2004), s. 315-319 ISSN 0015-5632 R&D Projects: GA AV ČR IBS5004009 Institutional research plan: CEZ:AV0Z5004920 Keywords : hemocytes * Bombyx mori * reactive oxygen species Subject RIV: BO - Biophysics Impact factor: 1.034, year: 2004

  3. Agglutinating antibodies against pathogenic Leptospira in healthy dogs and horses indicate common exposure and regular occurrence of subclinical infections

    NARCIS (Netherlands)

    D.J. Houwers; M.G.A. Goris (Marga); T.H. Abdoel (Theresia); J.A. Kas (Jeroen); S.S. Knobbe (Sandra); A.M. van Dongen (Astrid); F.E. Westerduin (Fenna); W.R. Klein (Wim); R.A. Hartskeerl (Rudy)

    2011-01-01

    textabstractIn order to get insight in the level of exposure to pathogenic Leptospira under the moderate sea climate conditions in the Netherlands, healthy dogs and horses were tested for antibodies using the Microscopic Agglutination Test (MAT). 55% of 198 dogs tested had agglutinating antibodies

  4. Agglutinating and bactericidal properties of fractions of rabbit anti-Vibrio cholerae serum.

    Science.gov (United States)

    Pike, R M; Chandler, C H

    1969-06-01

    The major portion of the agglutinating and bactericidal activity of the sera of rabbits immunized with live Vibrio cholerae or with cholera vaccine was found in the gammaM fractions during the early stages of immunization. After 5 weeks or more, gammaG fractions accounted for more than half of the agglutinating activity. When late antibody was measured as the amount of protein precipitated by somatic antigens, nearly 3 times as much gammaG as gammaM was required for agglutination, and about 30 times as much gammaG as gammaM was required to kill 50% of a standard inoculum in the presence of complement. The ratio of vibriocidal to agglutinin titer of gammaG fractions at different stages of immunization was more variable than that of gammaM fractions. More complement was required for a vibriocidal effect by gammaG than by gammaM. Increasing the amount of complement decreased the amount of both gammaG and gammaM required to kill, but smaller amounts of gammaM required disproportionately larger amounts of complement. Less time was required by gammaM than by gammaG to kill 50% of the inoculum. Removal of the group-reactive antibody from anti-Ogawa serum and serum fractions by absorption with Inaba reduced the vibriocidal titer by more than one-half.

  5. Diagnosis of Trichomonous vaginalis by microscopy, latex agglutination, diamond's media, and PCR in symptomatic women, Khartoum, Sudan.

    Science.gov (United States)

    Saleh, Amir M; Abdalla, Hamid S; Satti, Abdelsalam B; Babiker, Suad M; Gasim, Gasim I; Adam, Ishag

    2014-03-06

    Trichomoniasis is the most common sexually transmitted disease. However, limited data are available on an effective technique for the diagnosis of Trichomonas vaginalis. A cross-sectional study was conducted to evaluate the accuracy of wet mount microscopy, latex agglutination, Diamond's media, and polymerase chain reaction (PCR) for detection of T. vaginalis among symptomatic women who attended the gynecological clinic at Khartoum, Sudan. Of the 297 women studied, 252 (84.8%) were positive for T. vaginalis by wet mount microscopy, 257 (86.5%) by latex agglutination, 253 (85.2%) by Diamond's media, and 253 (85.2%) by PCR. The sensitivity and specificity of wet mount microscopy were 99.2% and 97.7%, respectively, compared with PCR. The sensitivity and specificity of latex agglutination and Diamond's media were 99.6% and 88.6%, and 100.0% and 86.4%, respectively, compared with PCR. In this study, wet mount microscopy, latex agglutination, and Diamond's media were found to be highly sensitive and specific. However, the availability and cost effectiveness might limit the use of Diamond's media and PCR in routine practice. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/7859723851211496.

  6. 75 - 78 Samira - BACTERIOPHAGES FINAL

    African Journals Online (AJOL)

    DR. AMIN

    Bayero Journal of Pure and Applied Sciences, 4(1): 75 - 78. Received: ... It involves the use of bacteriophages (small viruses that predate bacteria) to ..... Since the 1940s, research with ... phages is recognized by the appearance of plaques or.

  7. Silk Route to the Acceptance and Re-Implementation of Bacteriophage Therapy—Part II

    Directory of Open Access Journals (Sweden)

    Expert round table on acceptance and re-implementation of bacteriophage therapy

    2018-04-01

    Full Text Available This perspective paper follows up on earlier communications on bacteriophage therapy that we wrote as a multidisciplinary and intercontinental expert-panel when we first met at a bacteriophage conference hosted by the Eliava Institute in Tbilisi, Georgia in 2015. In the context of a society that is confronted with an ever-increasing number of antibiotic-resistant bacteria, we build on the previously made recommendations and specifically address how the Nagoya Protocol might impact the further development of bacteriophage therapy. By reviewing a number of recently conducted case studies with bacteriophages involving patients with bacterial infections that could no longer be successfully treated by regular antibiotic therapy, we again stress the urgency and significance of the development of international guidelines and frameworks that might facilitate the legal and effective application of bacteriophage therapy by physicians and the receiving patients. Additionally, we list and comment on several recently started and ongoing clinical studies, including highly desired double-blind placebo-controlled randomized clinical trials. We conclude with an outlook on how recently developed DNA editing technologies are expected to further control and enhance the efficient application of bacteriophages.

  8. [Determination of Azospirillum Brasilense Cells With Bacteriophages via Electrooptical Analysis of Microbial Suspensions].

    Science.gov (United States)

    Gulii, O I; Karavayeva, O A; Pavlii, S A; Sokolov, O I; Bunin, V D; Ignatov, O V

    2015-01-01

    The dependence-of changes in the electrooptical properties of Azospirillum brasilense cell suspension Sp7 during interaction with bacteriophage ΦAb-Sp7 on the number and time of interactions was studied. Incubation of cells with bacteriophage significantly changed the electrooptical signal within one minute. The selective effect of bacteriophage ΦAb on 18 strains of bacteria of the genus Azospirillum was studied: A. amazonense Ami4, A. brasilense Sp7, Cd, Sp107, Sp245, Jm6B2, Brl4, KR77, S17, S27, SR55, SR75, A. halopraeferans Au4, A. irakense KBC1, K A3, A. lipoferum Sp59b, SR65 and RG20a. We determined the limit of reliable determination of microbial cells infected with bacteriophage: - 10(4) cells/mL. The presence of foreign cell cultures of E. coli B-878 and E. coli XL-1 did not complicate the detection of A brasilense Sp7 cells with the use of bacteriophage ΦAb-Sp7. The results demonstrated that bacteriophage (ΦAb-Sp7 can be used for the detection of Azospirillum microbial cells via t electrooptical analysis of cell suspensions.

  9. Bacteriophage ecology in environmental biotechnology processes.

    Science.gov (United States)

    Shapiro, Orr H; Kushmaro, Ariel

    2011-06-01

    Heterotrophic bacteria are an integral part of any environmental biotechnology process (EBP). Therefore, factors controlling bacterial abundance, activity, and community composition are central to the understanding of such processes. Among these factors, top-down control by bacteriophage predation has so far received very limited attention. With over 10(8) particles per ml, phage appear to be the most numerous biological entities in EBP. Phage populations in EBP appear to be highly dynamic and to correlate with the population dynamics of their hosts and genomic evidence suggests bacteria evolve to avoid phage predation. Clearly, there is much to learn regarding bacteriophage in EBP before we can truly understand the microbial ecology of these globally important systems. Copyright © 2011 Elsevier Ltd. All rights reserved.

  10. 40 CFR 180.1261 - Xanthomonas campestris pv. vesicatoria and Pseudomonas syringae pv. tomato specific Bacteriophages.

    Science.gov (United States)

    2010-07-01

    ... and Pseudomonas syringae pv. tomato specific Bacteriophages. 180.1261 Section 180.1261 Protection of.... vesicatoria and Pseudomonas syringae pv. tomato specific Bacteriophages. An exemption from the requirement of... syringae pv. tomato specific bacteriophages in or on pepper and tomato. [74 FR 26536, June 3, 2009] ...

  11. Isolation of Dickeya dadantii strains from potato disease and biocontrol by their bacteriophages

    Directory of Open Access Journals (Sweden)

    Abbas Soleimani-Delfan

    2015-09-01

    Full Text Available One of the most economically important bacterial pathogens of plants and plant products is Dickeya dadantii. This bacterium causes soft rot disease in tubers and other parts of the potato and other plants of the Solanaceae family. The application of restricted host range bacteriophages as biocontrol agents has recently gained widespread interest. This study purposed to isolate the infectious agent of the potato and evaluate its biocontrol by bacteriophages. Two phytopathogenic strains were isolated from infected potatoes, identified based on biochemical and 16S rRNA gene sequencing, and submitted to GenBank as D. dadantii strain pis3 (accession no. HQ423668 and D. dadantii strain sip4 (accession no. HQ423669. Their bacteriophages were isolated from Caspian Sea water by enriching the water filtrate with D. dadantii strains as hosts using spot or overlay methods. On the basis of morphotypes, the isolated bacteriophages were identified as members of the Myoviridae and Siphoviridae families and could inhibit the growth of antibiotic resistant D. dadantii strains in culture medium. Moreover, in Dickeya infected plants treated with bacteriophage, no disease progression was detected. No significant difference was seen between phage-treated and control plants. Thus, isolated bacteriophages can be suggested for the biocontrol of plant disease caused by Dickeya strains.

  12. Bacteriophages of Soft Rot Enterobacteriaceae-a minireview.

    Science.gov (United States)

    Czajkowski, Robert

    2016-01-01

    Soft rot Enterobacteriaceae (Pectobacterium spp. and Dickeya spp., formerly pectinolytic Erwinia spp.) are ubiquitous necrotrophic bacterial pathogens that infect a large number of different plant species worldwide, including economically important crops. Despite the fact that these bacteria have been studied for more than 50 years, little is known of their corresponding predators: bacteriophages, both lytic and lysogenic. The aim of this minireview is to critically summarize recent ecological, biological and molecular research on bacteriophages infecting Pectobacterium spp. and Dickeya spp. with the main focus on current and future perspectives in that field. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. The Classroom-Friendly ABO Blood Types Kit: Blood Agglutination Simulation

    Science.gov (United States)

    Arnold, Savittree Rochanasmita; Kruatong, Tussatrin; Dahsah, Chanyah; Suwanjinda, Duongdearn

    2012-01-01

    The classroom-friendly ABO blood type kit was developed by combining advantages of modelling and a simulation laboratory to teach the topics of ABO blood types and blood transfusion. Teachers can easily simulate the agglutination reaction on a blood type testing plate in the classroom, and show the students how this reaction occurs by using the…

  14. 9 CFR 147.1 - The standard tube agglutination test. 1

    Science.gov (United States)

    2010-01-01

    ... may be done with a suitable pencil on etched portions of the tube, or by means of fast-gum labels. (3..., high agglutinability, but are not sensitive to negative and nonspecific sera. The stock cultures may be... for at least 20 hours at 37 °C. (h) The results shall be recorded as: N, or − (negative) when the...

  15. Metagenomic Analysis of Dairy Bacteriophages

    DEFF Research Database (Denmark)

    Muhammed, Musemma K.; Kot, Witold; Neve, Horst

    2017-01-01

    Despite their huge potential for characterizing the biodiversity of phages, metagenomic studies are currently not available for dairy bacteriophages, partly due to the lack of a standard procedure for phage extraction. We optimized an extraction method that allows to remove the bulk protein from...

  16. Comparative determination of the rheumatic factor by means of agglutination, immunofluorescence and radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Jaeger, L.; Storz, H.; Hein, G.; Schlenvoigt, G. (Friedrich-Schiller-Universitaet, Jena (German Democratic Republic). Bereich Medizin)

    1982-01-01

    The rheumatic factor (RF) was determined by means of agglutination, immunofluorescence (IF) test and radioimmunoassay (RIPEGA) in random groups of 56 patients with rheumatoid arthritis (RA), 13 patients with seronegative RA and 39 patients with psoriasis arthropathica. All three methods are of equal value with regard to the number of positive results. Further classification of seronegative patients, i.e. patients with a negative agglutination reaction and the clinical symptoms of RA is possible with the IF method and, above all, by means of RIPEGA. But because of the comprehensive test devices the two methods are only an alternative. Titer differences are attributed to the different indication principles and the immunological heterogeneity of RF. An improvement of the diagnosis of activity was not possible.

  17. Comparative determination of the rheumatic factor by means of agglutination, immunofluorescence and radioimmunoassay

    International Nuclear Information System (INIS)

    Jaeger, L.; Storz, H.; Hein, G.; Schlenvoigt, G.

    1982-01-01

    The rheumatic factor (RF) was determined by means of agglutination, immunofluorescence (IF) test and radioimmunoassay (RIPEGA) in random groups of 56 patients with rheumatoid arthritis (RA), 13 patients with seronegative RA and 39 patients with psoriasis arthropathica. All three methods are of equal value with regard to the number of positive results. Further classification of seronegative patients, i.e. patients with a negative agglutination reaction and the clinical symptoms of RA is possible with the IF method and, above all, by means of RIPEGA. But because of the comprehensive test devices the two methods are only an alternative. Titer differences are attributed to the different indication principles and the immunological heterogeneity of RF. An improvement of the diagnosis of activity was not possible. (author)

  18. Comparison of agglutinating and neutralizing antibodies to serovar hardjo in sows immunized with two commercial whole culture polivalent anti-leptospira bacterins

    Science.gov (United States)

    Soto, Francisco Rafael Martins; Pinheiro, Sônia Regina; Morais, Zenaide Maria; Gonçales, Amane Paldês; de Azevedo, Sérgio Santos; Bernardi, Fernanda; Camargo, Sebastião Rodrigues; Vasconcellos, Silvio Arruda

    2008-01-01

    It was performed the comparison of the intensity and duration of agglutinating and neutralizing antibodies to serovar Hardjo in swines vaccinated with two commercial anti-leptospira bacterins. Sows no reactive to 24 Leptospira sp serovars in the microscopic agglutination test (MAT) were divided in three groups: Group A (n=08): received two vaccine A doses with 30 days interval, Group B (n=08) two vaccine B doses with 30 days interval and Group C (n=08): control no vaccinated against leptospirosis.Blood samples were collected each 30 days during six months following the first vaccination. The sera were tested by MAT and growth inhibition test (GIT) to serovar Hardjo in order to evaluate respectively agglutinating and neutralizing antibodies. It was found that neutralizing antibodies persisted for a longer time than the agglutinating ones and that the absence of agglutinating antibodies does not means in the absence of the neutralizing. The peaks of agglutinating antibodies was obtained at least 30 days earlier than that produced by neutralizing. The duration of both kinds of antibodies measured differed between the two bacterines tested. The period for inducing neutralizing antibodies against serovar Hardjo indicated that gilts must be immunized with two doses of whole culture anti-leptospira bacterines applied 30 days each other at least 90 days before the first mating. For the maintenance of hight levels of neutralizing antibodies the revaccinations must be performed every six months after the first vaccination. PMID:24031250

  19. Bacteriophage-Based Bacterial Wilt Biocontrol for an Environmentally Sustainable Agriculture

    Directory of Open Access Journals (Sweden)

    Belén Álvarez

    2017-07-01

    Full Text Available Bacterial wilt diseases caused by Ralstonia solanacearum, R. pseudosolanacearum, and R. syzygii subsp. indonesiensis (former R. solanacearum species complex are among the most important plant diseases worldwide, severely affecting a high number of crops and ornamentals. Difficulties of bacterial wilt control by non-biological methods are related to effectiveness, bacterial resistance and environmental impact. Alternatively, a great many biocontrol strategies have been carried out, with the advantage of being environmentally friendly. Advances in bacterial wilt biocontrol include an increasing interest in bacteriophage-based treatments as a promising re-emerging strategy. Bacteriophages against the bacterial wilt pathogens have been described with either lytic or lysogenic effect but, they were proved to be active against strains belonging to R. pseudosolanacearum and/or R. syzygii subsp. indonesiensis, not to the present R. solanacearum species, and only two of them demonstrated successful biocontrol potential in planta. Despite the publication of three patents on the topic, until now no bacteriophage-based product is commercially available. Therefore, there is still much to be done to incorporate valid bacteriophages in an integrated management program to effectively fight bacterial wilt in the field.

  20. Bacteriophage-Based Bacterial Wilt Biocontrol for an Environmentally Sustainable Agriculture.

    Science.gov (United States)

    Álvarez, Belén; Biosca, Elena G

    2017-01-01

    Bacterial wilt diseases caused by Ralstonia solanacearum , R. pseudosolanacearum , and R. syzygii subsp. indonesiensis (former R. solanacearum species complex) are among the most important plant diseases worldwide, severely affecting a high number of crops and ornamentals. Difficulties of bacterial wilt control by non-biological methods are related to effectiveness, bacterial resistance and environmental impact. Alternatively, a great many biocontrol strategies have been carried out, with the advantage of being environmentally friendly. Advances in bacterial wilt biocontrol include an increasing interest in bacteriophage-based treatments as a promising re-emerging strategy. Bacteriophages against the bacterial wilt pathogens have been described with either lytic or lysogenic effect but, they were proved to be active against strains belonging to R. pseudosolanacearum and/or R. syzygii subsp. indonesiensis , not to the present R. solanacearum species, and only two of them demonstrated successful biocontrol potential in planta . Despite the publication of three patents on the topic, until now no bacteriophage-based product is commercially available. Therefore, there is still much to be done to incorporate valid bacteriophages in an integrated management program to effectively fight bacterial wilt in the field.

  1. Targeting Antibacterial Agents by Using Drug-Carrying Filamentous Bacteriophages

    OpenAIRE

    Yacoby, Iftach; Shamis, Marina; Bar, Hagit; Shabat, Doron; Benhar, Itai

    2006-01-01

    Bacteriophages have been used for more than a century for (unconventional) therapy of bacterial infections, for half a century as tools in genetic research, for 2 decades as tools for discovery of specific target-binding proteins, and for nearly a decade as tools for vaccination or as gene delivery vehicles. Here we present a novel application of filamentous bacteriophages (phages) as targeted drug carriers for the eradication of (pathogenic) bacteria. The phages are genetically modified to d...

  2. An Insight into the proteome of Crithidia fasciculata choanomastigotes as a comparative approach to axenic growth, peanut lectin agglutination and differentiation of Leishmania spp. promastigotes.

    Directory of Open Access Journals (Sweden)

    Pedro J Alcolea

    Full Text Available The life cycle of the trypanosomatid Crithidia fasciculata is monogenetic, as the unique hosts of these parasites are different species of culicids. The comparison of these non-pathogenic microorganisms evolutionary close to other species of trypanosomatids that develop digenetic life cycles and cause chronic severe sickness to millions of people worldwide is of outstanding interest. A ground-breaking analysis of differential protein abundance in Crithidia fasciculata is reported herein. The comparison of the outcome with previous gene expression profiling studies developed in the related human pathogens of the genus Leishmania has revealed substantial differences between the motile stages of these closely related organisms in abundance of proteins involved in catabolism, redox homeostasis, intracellular signalling, and gene expression regulation. As L. major and L. infantum agglutinate with peanut lectin and non-agglutinating parasites are more infective, the agglutination properties were evaluated in C. fasciculata. The result is that choanomastigotes are able to agglutinate with peanut lectin and a non-agglutinating subpopulation can be also isolated. As a difference with L. infantum, the non-agglutinating subpopulation over-expresses the whole machinery for maintenance of redox homeostasis and the translation factors eIF5a, EF1α and EF2, what suggests a relationship between the lack of agglutination and a differentiation process.

  3. Bacteriophage therapy for safeguarding animal and human health: a review.

    Science.gov (United States)

    Tiwari, Ruchi; Dhama, Kuldeep; Kumar, Amit; Rahal, Anu; Kapoor, Sanjay

    2014-02-01

    Since the discovery of bacteriophages at the beginning of the 19th century their contribution to bacterial evolution and ecology and use in a variety of applications in biotechnology and medicine has been recognized and understood. Bacteriophages are natural bacterial killers, proven as best biocontrol agents due to their ability to lyse host bacterial cells specifically thereby helping in disease prevention and control. The requirement of such therapeutic approach is straight away required in view of the global emergence of Multidrug Resistant (MDR) strains of bacteria and rapidly developing resistance to antibiotics in both animals and humans along with increasing food safety concerns including of residual antibiotic toxicities. Phage typing is a popular tool to differentiate bacterial isolates and to identify and characterize outbreak-associated strains of Salmonella, Campylobacter, Escherichia and Listeria. Numerous methods viz. plaque morphology, ultracentrifugation in the density gradient of CsCl2, and random amplified polymorphic DNA (RAPD) have been found to be effective in detection of various phages. Bacteriophages have been isolated and recovered from samples of animal waste products of different livestock farms. High titer cocktails of broad spectrum lytic bacteriophages are usually used for clinical trial for assessing their therapeutic efficacy against antibiotic unresponsive infections in different animals. Bacteriophage therapy also helps to fight various bacterial infections of poultry viz. colibacillosis, salmonellosis and listeriosis. Moreover, the utility of phages concerning biosafety has raised the importance to explore and popularize the therapeutic dimension of this promising novel therapy which forms the topic of discussion of the present review.

  4. Interactions of cationic polystyrene nanoparticles with marine bivalve hemocytes in a physiological environment: Role of soluble hemolymph proteins

    Energy Technology Data Exchange (ETDEWEB)

    Canesi, Laura, E-mail: Laura.Canesi@unige.it [Dept. of Earth, Environmental and Life Sciences – DISTAV, University of Genoa (Italy); Ciacci, Caterina [Dept. of Biomolecular Sciences – DIBS, University of Urbino (Italy); Fabbri, Rita; Balbi, Teresa [Dept. of Earth, Environmental and Life Sciences – DISTAV, University of Genoa (Italy); Salis, Annalisa; Damonte, Gianluca [Centre of Excellence for Biomedical Research – CEBR, University of Genoa (Italy); Cortese, Katia [Department of Experimental Medicine – DIMES, University of Genoa (Italy); Caratto, Valentina [Dept. of Earth, Environmental and Life Sciences – DISTAV, University of Genoa (Italy); Monopoli, Marco P. [Centre for BioNanoInteractions, School of Chemistry and Chemical Biology, University College Dublin (Ireland); Department of Pharmaceutical and Medical Chemistry, Royal College of Surgeons, 123 St. Stephen Green, Dublin (Ireland); Dawson, Kenneth [Centre for BioNanoInteractions, School of Chemistry and Chemical Biology, University College Dublin (Ireland); Bergami, Elisa; Corsi, Ilaria [Dept. of Physical, Earth and Environmental Sciences, University of Siena (Italy)

    2016-10-15

    The bivalve Mytilus galloprovincialis has proven as a suitable model invertebrate for evaluating the potential impact of nanoparticles (NPs) in the marine environment. In particular, in mussels, the immune system represents a sensitive target for different types of NPs. In environmental conditions, both NP intrinsic properties and those of the receiving medium will affect particle behavior and consequent bioavailability/uptake/toxicity. However, the evaluation of the biological effects of NPs requires additional understanding of how, once within the organism, NPs interact at the molecular level with cells in a physiological environment. In mammalian systems, different NPs associate with serum soluble components, organized into a “protein corona”, which affects particle interactions with target cells. However, no information is available so far on the interactions of NPs with biological fluids of aquatic organisms. In this work, the influence of hemolymph serum (HS) on the in vitro effects of amino modified polystyrene NPs (PS-NH{sub 2}) on Mytilus hemocytes was investigated. Hemocytes were incubated with PS-NH{sub 2} suspensions in HS (1, 5 and 50 µg/mL) and the results were compared with those obtained in ASW medium. Cell functional parameters (lysosomal membrane stability, oxyradical production, phagocytosis) were evaluated, and morphological changes were investigated by TEM. The activation state of the signalling components involved in Mytilus immune response (p38 MAPK and PKC) was determined. The results show that in the presence of HS, PS-NH{sub 2} increased cellular damage and ROS production with respect to ASW medium. The effects were apparently mediated by disregulation of p38 MAPK signalling. The formation of a PS-NH{sub 2}-protein corona in HS was investigated by centrifugation, and 1D- gel electrophoresis and nano-HPLC-ESI-MS/MS. The results identified the Putative C1q domain containing protein (MgC1q6) as the only component of the PS-NH{sub 2} hard

  5. Interactions of cationic polystyrene nanoparticles with marine bivalve hemocytes in a physiological environment: Role of soluble hemolymph proteins

    International Nuclear Information System (INIS)

    Canesi, Laura; Ciacci, Caterina; Fabbri, Rita; Balbi, Teresa; Salis, Annalisa; Damonte, Gianluca; Cortese, Katia; Caratto, Valentina; Monopoli, Marco P.; Dawson, Kenneth; Bergami, Elisa; Corsi, Ilaria

    2016-01-01

    The bivalve Mytilus galloprovincialis has proven as a suitable model invertebrate for evaluating the potential impact of nanoparticles (NPs) in the marine environment. In particular, in mussels, the immune system represents a sensitive target for different types of NPs. In environmental conditions, both NP intrinsic properties and those of the receiving medium will affect particle behavior and consequent bioavailability/uptake/toxicity. However, the evaluation of the biological effects of NPs requires additional understanding of how, once within the organism, NPs interact at the molecular level with cells in a physiological environment. In mammalian systems, different NPs associate with serum soluble components, organized into a “protein corona”, which affects particle interactions with target cells. However, no information is available so far on the interactions of NPs with biological fluids of aquatic organisms. In this work, the influence of hemolymph serum (HS) on the in vitro effects of amino modified polystyrene NPs (PS-NH 2 ) on Mytilus hemocytes was investigated. Hemocytes were incubated with PS-NH 2 suspensions in HS (1, 5 and 50 µg/mL) and the results were compared with those obtained in ASW medium. Cell functional parameters (lysosomal membrane stability, oxyradical production, phagocytosis) were evaluated, and morphological changes were investigated by TEM. The activation state of the signalling components involved in Mytilus immune response (p38 MAPK and PKC) was determined. The results show that in the presence of HS, PS-NH 2 increased cellular damage and ROS production with respect to ASW medium. The effects were apparently mediated by disregulation of p38 MAPK signalling. The formation of a PS-NH 2 -protein corona in HS was investigated by centrifugation, and 1D- gel electrophoresis and nano-HPLC-ESI-MS/MS. The results identified the Putative C1q domain containing protein (MgC1q6) as the only component of the PS-NH 2 hard protein corona in

  6. Bacteriophages-potential for application in wastewater treatment processes

    International Nuclear Information System (INIS)

    Withey, S.; Cartmell, E.; Avery, L.M.; Stephenson, T.

    2005-01-01

    Bacteriophages are viruses that infect and lyse bacteria. Interest in the ability of phages to control bacterial populations has extended from medical applications into the fields of agriculture, aquaculture and the food industry. Here, the potential application of phage techniques in wastewater treatment systems to improve effluent and sludge emissions into the environment is discussed. Phage-mediated bacterial mortality has the potential to influence treatment performance by controlling the abundance of key functional groups. Phage treatments have the potential to control environmental wastewater process problems such as: foaming in activated sludge plants; sludge dewaterability and digestibility; pathogenic bacteria; and to reduce competition between nuisance bacteria and functionally important microbial populations. Successful application of phage therapy to wastewater treatment does though require a fuller understanding of wastewater microbial community dynamics and interactions. Strategies to counter host specificity and host cell resistance must also be developed, as should safety considerations regarding pathogen emergence through transduction

  7. Hemocytes of zebra mussels (Dreissena polymorpha are relevant cells for the monitoring of environmental genotoxicity by the comet assay.

    Directory of Open Access Journals (Sweden)

    Marc Bonnard

    2015-06-01

    Full Text Available The measure of DNA integrity by the single cell gel electrophoresis (SCGE or comet assay is especially recommended for its sensitivity and its capacity for detecting different types of damages. Therefore, it has been applied in environmental genotoxicity in a variety of organisms. It appears today necessary to define both reference and threshold levels of DNA damage, for their application in in situ biomonitoring. However, little is known about the influence of both biological (sex, reproduction status or external (temperature… confounding factors on the measure of DNA damage by the comet assay. These variables need to be taken into account if the robustness of the assay is to be established (Jha, 2008. In the zebra mussel Dreissena polymorpha (recommended as a sentinel species in the evaluation of freshwater quality the measure of DNA damage by the comet assay is mainly performed on hemocytes, which are circulating cells involved in key physiological functions such as immunity, homeostasis, detoxication…. This communication will present and discuss results from an innovative study about the variability of the baseline level of DNA damage in hemocytes of mussels encaged for one year in the canal de l’Aisne à la Marne (Reims, according to their sex and their reproductive status. The sensitivity and the suitability of hemocytes in the evaluation of environmental genotoxicity will also be discussed, referring to observations during a 6 month-exposure of mussels in mesocosms to environmentally realistic concentrations of carbamazepine.

  8. Identification and Characterization of T5-Like Bacteriophages Representing Two Novel Subgroups from Food Products

    Directory of Open Access Journals (Sweden)

    Domonkos Sváb

    2018-02-01

    Full Text Available During recent years, interest in the use of bacteriophages as biocontrol agents against foodborne pathogens has increased, particularly for members of the family Enterobacteriaceae, with pathogenic Escherichia coli, Shigella, and Salmonella strains among them. Here, we report the isolation and characterisation of 12 novel T5-like bacteriophages from confiscated food samples. All bacterophages effectively lysed E. coli K-12 strains and were able to infect pathogenic E. coli strains representing enterohaemorrhagic (EHEC, enteropathogenic (EPEC, enterotoxigenic (ETEC, and enteroinvasive (EIEC pathotypes, Shigella dysenteriae, S. sonnei strains, as well as multidrug-resistant (MDR E. coli and multiple strains representing different Salmonella enterica serovars. All the bacteriophages exhibited Siphoviridae morphology. Whole genome sequencing of the novel T5-like bacteriophages showed that they represent two distinct groups, with the genome-based grouping correlating to the different host spectra. As these bacteriophages are of food origin, their stability and lack of any virulence genes, as well as their broad and mutually complementary host spectrum makes these new T5-like bacteriophages valuable candidates for use as biocontrol agents against foodborne pathogenic enterobacteria.

  9. A comparison of titers of anti-Brucella antibodies of naturally infected and healthy vaccinated cattle by standard tube agglutination test, microtiter plate agglutination test, indirect hemagglutination assay, and indirect enzyme-linked immunosorbent assay

    Directory of Open Access Journals (Sweden)

    Anju Mohan

    2016-07-01

    Full Text Available Aim: We determined the antibody response in cattle naturally infected with brucellosis and normal healthy adult cattle vaccinated during calf hood with strain 19. Materials and Methods: The antibody titers were measured by standard tube agglutination test (STAT, microtiter plate agglutination test (MAT, indirect hemagglutination assay (IHA, and indirect enzyme-linked immunosorbent assay (iELISA as per standard protocols. Results: The mean STAT titers were 1.963±0.345 in infected cattle and 1.200±0.155 in healthy vaccinated cattle. The difference was extremely significant (p<0.0001. The mean MAT titers were 2.244±0.727 in infected cattle and 1.200±0.155 in healthy vaccinated cattle. The difference was very significant (p<0.005. The mean IHA titers in infected cattle were 2.284±0.574, and those in healthy vaccinated cattle were 1.200±0.155. The difference was extremely significant (p=0.0002. However, the difference in mean iELISA titers of infected cattle (1.3678±0.014 and healthy vaccinated cattle (1.367±0.014 was non-significant. The infected animals showed very high titers of agglutinating antibodies compared to the vaccinated animals. However, it cannot be ascertained whether these antibodies are due to vaccine or response to infection. Since the infected animals had been vaccinated earlier, the current infection may suggest that vaccination was unable to induce protective levels of antibody. The heightened antibody response after infection may also indicate a secondary immune response to the antigens common to the vaccine strain and wild Brucella organisms. Conclusion: The brucellosis infected animals showed very high titers of agglutinating antibodies compared to the vaccinated animals.

  10. Role of RecA protein in untargeted UV mutagenesis of bacteriophage lambda: evidence for the requirement for the dinB gene

    International Nuclear Information System (INIS)

    Brotcorne-Lannoye, A.; Maenhaut-Michel, G.

    1986-01-01

    Untargeted UV mutagenesis of bacteriophage lambda--i.e., the increased recovery of lambda mutants when unirradiated lambda infects UV-irradiated Escherichia coli--is thought to be mediated by a transient decrease in DNA replication fidelity, generating mutations in the newly synthesized strands. Using the bacteriophage lambda cI857----lambda c mutation system, we provide evidence that the RecA protein, shown previously to be required for this mutagenic pathway, is no longer needed when the LexA protein is inactivated by mutation. We suggest that the error-prone DNA replication responsible for UV-induced untargeted mutagenesis is turned on by the presence of replication-blocking lesions in the host cell DNA and that the RecA protein is required only to derepress the relevant din gene(s). This is in contrast to mutagenesis of irradiated bacteria or irradiated phage lambda, in which activated RecA protein has a second role in mutagenesis in addition to the cleavage of the LexA protein. Among the tested din genes, the dinB gene product (in addition to the uvrA and uvrB gene products) was found to be required for untargeted mutagenesis of bacteriophage lambda. To our knowledge, a phenotype associated with the dinB gene has not been reported previously

  11. Comparative Genomics of Bacteriophage of the Genus Seuratvirus

    DEFF Research Database (Denmark)

    Sazinas, Pavelas; Redgwell, Tamsin; Rihtman, Branko

    2017-01-01

    polB and terL showed these bacteriophages to be closely related to members of the genus Seuratvirus. We performed a core-gene analysis using the 14 new and four closely related genomes. A total of 58 core genes were identified, the majority of which has no known function. These genes were used...... to construct a core-gene phylogeny, the results of which confirmed the new isolates to be part of the genus Seuratvirus and expanded the number of species within this genus to four. All bacteriophages within the genus contained the genes queCDE encoding enzymes involved in queuosine biosynthesis. We suggest...

  12. Bacteriophage interactions with marine pathogenic Vibrios

    DEFF Research Database (Denmark)

    Kalatzis, Panagiotis

    development and spreading of antibiotic resistant bacteria in the environment. Bacteriophage therapy, constitutes a potent alternative not only for treatment but also for prevention of vibriosis in aquaculture and the current thesis addresses the potential and challenges of using phages to control Vibrio...... pathogens. The combinatory administration of virulent bacteriophages φSt2 and φGrn1, isolated against Vibrio alginolyticus significantly reduced the Vibrio load in cultures of Artemia salina live prey, decreasing subsequently the risk of a vibriosis outbreak in the marine hatchery. During infection...... therapy applications. Lytic phage vB_VspP_pVa5 that has been isolated against the rapidly emerging pathogen V. splendidus is also a promising candidate for phage therapy application according to its gene content and in vitro performance against its host. The genetic features of vB_VspP_pVa5 provide also...

  13. Pulmonary bacteriophage therapy on Pseudomonas aeruginosa cystic fibrosis strains: first steps towards treatment and prevention.

    Directory of Open Access Journals (Sweden)

    Eric Morello

    Full Text Available Multidrug-resistant bacteria are the cause of an increasing number of deadly pulmonary infections. Because there is currently a paucity of novel antibiotics, phage therapy--the use of specific viruses that infect bacteria--is now more frequently being considered as a potential treatment for bacterial infections. Using a mouse lung-infection model caused by a multidrug resistant Pseudomonas aeruginosa mucoid strain isolated from a cystic fibrosis patient, we evaluated bacteriophage treatments. New bacteriophages were isolated from environmental samples and characterized. Bacteria and bacteriophages were applied intranasally to the immunocompetent mice. Survival was monitored and bronchoalveolar fluids were analysed. Quantification of bacteria, bacteriophages, pro-inflammatory and cytotoxicity markers, as well as histology and immunohistochemistry analyses were performed. A curative treatment (one single dose administrated 2 h after the onset of the infection allowed over 95% survival. A four-day preventive treatment (one single dose resulted in a 100% survival. All of the parameters measured correlated with the efficacy of both curative and preventive bacteriophage treatments. We also showed that in vitro optimization of a bacteriophage towards a clinical strain improved both its efficacy on in vivo treatments and its host range on a panel of 20 P. aeruginosa cystic fibrosis strains. This work provides an incentive to develop clinical studies on pulmonary bacteriophage therapy to combat multidrug-resistant lung infections.

  14. Nematocytes: Discovery and characterization of a novel anculeate hemocyte in Drosophila falleni and Drosophila phalerata.

    Directory of Open Access Journals (Sweden)

    Julianna Bozler

    Full Text Available Immune challenges, such as parasitism, can be so pervasive and deleterious that they constitute an existential threat to a species' survival. In response to these ecological pressures, organisms have developed a wide array of novel behavioral, cellular, and molecular adaptations. Research into these immune defenses in model systems has resulted in a revolutionary understanding of evolution and functional biology. As the field has expanded beyond the limited number of model organisms our appreciation of evolutionary innovation and unique biology has widened as well. With this in mind, we have surveyed the hemolymph of several non-model species of Drosophila. Here we identify and describe a novel hemocyte, type-II nematocytes, found in larval stages of numerous Drosophila species. Examined in detail in Drosophila falleni and Drosophila phalerata, we find that these remarkable cells are distinct from previously described hemocytes due to their anucleate state (lacking a nucleus and unusual morphology. Type-II nematocytes are long, narrow cells with spindle-like projections extending from a cell body with high densities of mitochondria and microtubules, and exhibit the ability to synthesize proteins. These properties are unexpected for enucleated cells, and together with our additional characterization, we demonstrate that these type-II nematocytes represent a biological novelty. Surprisingly, despite the absence of a nucleus, we observe through live cell imaging that these cells remain motile with a highly dynamic cellular shape. Furthermore, these cells demonstrate the ability to form multicellular structures, which we suggest may be a component of the innate immune response to macro-parasites. In addition, live cell imaging points to a large nucleated hemocyte, type-I nematocyte, as the progenitor cell, leading to enucleation through a budding or asymmetrical division process rather than nuclear ejection: This study is the first to report such a

  15. R and G color component competition of RGB image decomposition as a criterion to register RBC agglutinates for blood group typing.

    Science.gov (United States)

    Doubrovski, Valeri A; Ganilova, Yuliya A; Zabenkov, Igor V

    2014-03-01

    A new approach of the criterion assignment for registration of erythrocyte agglutinates to instrumentally determine blood group type is suggested. The criterion is based on comparison of R and G components of RGB decomposition of microscopy digital image taken for the blood-serum mixture sample. For the chosen experimental conditions, the minimal size (area) of RBC agglutinate to be registered by the criterion suggested is estimated theoretically. The proposed method was tested experimentally on the example of monitoring agglutinates in flow. The encouraging experimental results were obtained for improvement of the resolving power of the method; the optimal experimental conditions were revealed for maximum resolution. Though the suggested method was realized for dynamic (flow) blood group determination, it could also be applied for diagnostics in a stationary environment. This approach increases the reliability of RBC agglutinates registration and, hence, blood group typing. The results may be used to develop the apparatus for automated determination of human blood group.

  16. Complete genome sequence of the Pectobacterium carotovorum subsp. carotovorum virulent bacteriophage PM1.

    Science.gov (United States)

    Lim, Jeong-A; Shin, Hakdong; Lee, Dong Hwan; Han, Sang-Wook; Lee, Ju-Hoon; Ryu, Sangryeol; Heu, Sunggi

    2014-08-01

    PM1, a novel virulent bacteriophage that infects Pectobacterium carotovorum subsp. carotovorum, was isolated. Its morphological features were examined by electron microscopy, which indicated that this phage belongs to the family Myoviridae. It has a 55,098-bp genome, including a 2,665-bp terminal repeat. A total of 63 open reading frames (ORFs) were predicted, but only 20 ORFs possessed homology with functional proteins. There is one tRNA coding region, and the GC-content of the genome is 44.9 %. Most ORFs in bacteriophage PM1 showed high homology to enterobacteria phage ΦEcoM-GJ1 and Erwinia phage νB EamM-Y2. Like these bacteriophages, PM1 encodes an RNA polymerase, which is a hallmark of T7-like phages. There is no integrase or repressor, suggesting that PM1 is a virulent bacteriophage.

  17. Occurrence and numbers of bacteriophages and bacterial indicators in faeces of yellow-legged seagull (Larus cachinnans).

    Science.gov (United States)

    Muniesa, M; Jofre, J; Lucena, F

    1999-12-01

    Faeces from feral populations of yellow-legged seagulls from the northern coastal area of Catalonia (North-eastern Spain) contained variable amounts of faecal coliforms, faecal streptococci, somatic coliphages, F-specific bacteriophages and Bacteroides fragilis bacteriophages. Occurrence and numbers of bacterial indicators and bacteriophages in the faeces of yellow-legged seagulls are in the ranges described in the faeces of different animals. The ratios between numbers of bacterial indicators and numbers of bacteriophages are much higher in faeces of seagulls than in treated or raw sewage contributed by out-falls of the same area.

  18. Methods for generation of reporter phages and immobilization of active bacteriophages on a polymer surface

    Science.gov (United States)

    Morgan, Mark Thomas (Inventor); Kothapalli, Aparna (Inventor); Applegate, Bruce Michael (Inventor); Perry, Lynda Louise (Inventor)

    2012-01-01

    Novel reporter bacteriophages are provided. Provided are compositions and methods that allow bacteriophages that are used for specific detection or killing of E. coli 0157:H7 to be propagated in nonpathogenic E. coli, thereby eliminating the safety and security risks of propagation in E. coli 0157:H7. Provided are compositions and methods for attaching active bacteriophages to the surface of a polymer in order to kill target bacteria with which the phage comes into contact. Provided are modified bacteriophages immobilized to a surface, which capture E. coli 0157:H7 and cause the captured cells to emit light or fluorescence, allowing detection of the bacteria in a sample.

  19. Genetically engineered bacteriophage delivers a tumor necrosis factor alpha antagonist coating on neural electrodes

    International Nuclear Information System (INIS)

    Kim, Young Jun; Nam, Chang-Hoon; Jin, Young-Hyun; Stieglitz, Thomas; Salieb-Beugelaar, Georgette B

    2014-01-01

    This paper reports a novel approach for the formation of anti-inflammatory surface coating on a neural electrode. The surface coating is realized using a recombinant f88 filamentous bacteriophage, which displays a short platinum binding motif and a tumor necrosis factor alpha antagonist (TNF-α antagonist) on p3 and p8 proteins, respectively. The recombinant bacteriophages are immobilized on the platinum surface by a simple dip coating process. The selective and stable immobilization of bacteriophages on a platinum electrode is confirmed by quartz crystal microbalance with dissipation monitoring, atomic force microscope and fluorescence microscope. From the in vitro cell viability test, the inflammatory cytokine (TNF-α) induced cell death was prevented by presenting recombinant bacteriophage coating, albeit with no significant cytotoxic effect. It is also observed that the bacteriophage coating does not have critical effects on the electrochemical properties such as impedance and charge storage capacities. Thus, this approach demonstrates a promising anti-apoptotic as well as anti-inflammatory surface coating for neural implant applications. (paper)

  20. What history tells us XLIII Bacteriophage

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Biosciences; Volume 42; Issue 3. What history tells us XLIII Bacteriophage: The contexts in which it was discovered. MICHEL MORANGE. Series Volume 42 Issue 3 September 2017 pp 359-362. Fulltext. Click here to view fulltext PDF. Permanent link:

  1. Bombyx mori and Aedes aegypti form multi-functional immune complexes that integrate pattern recognition, melanization, coagulants, and hemocyte recruitment.

    Science.gov (United States)

    Phillips, Dennis R; Clark, Kevin D

    2017-01-01

    The innate immune system of insects responds to wounding and pathogens by mobilizing multiple pathways that provide both systemic and localized protection. Key localized responses in hemolymph include melanization, coagulation, and hemocyte encapsulation, which synergistically seal wounds and envelop and destroy pathogens. To be effective, these pathways require a targeted deposition of their components to provide protection without compromising the host. Extensive research has identified a large number of the effectors that comprise these responses, but questions remain regarding their post-translational processing, function, and targeting. Here, we used mass spectrometry to demonstrate the integration of pathogen recognition proteins, coagulants, and melanization components into stable, high-mass, multi-functional Immune Complexes (ICs) in Bombyx mori and Aedes aegypti. Essential proteins common to both include phenoloxidases, apolipophorins, serine protease homologs, and a serine protease that promotes hemocyte recruitment through cytokine activation. Pattern recognition proteins included C-type Lectins in B. mori, while A. aegypti contained a protein homologous to Plasmodium-resistant LRIM1 from Anopheles gambiae. We also found that the B. mori IC is stabilized by extensive transglutaminase-catalyzed cross-linking of multiple components. The melanization inhibitor Egf1.0, from the parasitoid wasp Microplitis demolitor, blocked inclusion of specific components into the IC and also inhibited transglutaminase activity. Our results show how coagulants, melanization components, and hemocytes can be recruited to a wound surface or pathogen, provide insight into the mechanism by which a parasitoid evades this immune response, and suggest that insects as diverse as Lepidoptera and Diptera utilize similar defensive mechanisms.

  2. C-type lectin B (SpCTL-B) regulates the expression of antimicrobial peptides and promotes phagocytosis in mud crab Scylla paramamosain.

    Science.gov (United States)

    Wei, Xiaoyuan; Wang, Limin; Sun, Wanwei; Zhang, Ming; Ma, Hongyu; Zhang, Yueling; Zhang, Xinxu; Li, Shengkang

    2018-07-01

    As pattern recognition receptors, C-type lectins (CTLs) play important roles in immune system of crustaceans through identifying and binding to the conservative pathogen-associated molecular patterns (PAMPs) on pathogen surfaces. In this study, a new CTL, SpCTL-B, was identified from the hemocytes of mud crab Scylla paramamosain. The full-length of SpCTL-B cDNA was 1278 bp with an open reading frame (ORF) of 348 bp. The predicted SpCTL-B protein contains a single carbohydrate-recognition domain (CRD). SpCTL-B transcripts were distributed in all examined tissues with the highest levels in hepatopancreas. After challenged with Vibrio parahaemolyticus, LPS, polyI:C and white spot syndrome virus (WSSV), the mRNA levels of SpCTL-B in hemocytes and hepatopancreas were up-regulated. The recombinant SpCTL-B (rSpCTL-B) purified by Ni-affinity chromatography showed stronger binding activities with Staphylococcus aureus, β-hemolytic Streptococcus, Escherichia coli, Aeromonas hydrophila, Vibrio alginolyticus than those with V. parahaemolyticus and Saccharomyces cerevisiae. rSpCTL-B exhibited a broad spectrum of microorganism-agglutination activities against Gram-positive bacteria (S. aureus, β-hemolytic Streptococcus) and Gram-negative bacteria (E. coli, V. parahaemolyticus, A. hydrophila, V. alginolyticus) in a Ca 2+ -dependent manner. The agglutination activities of rSpCTL-B could be inhibited by D-mannose and LPS, but not by d-fructose and galactose. The antimicrobial assay showed that rSpCTL-B exhibited the growth inhibition against all examined gram-positive bacteria and gram-negative bacteria. When SpCTL-B was silenced by RNAi, the bacterial clearance ability in mud crab was decreased and the transcript levels of five antimicrobial peptides (AMPs) (SpCrustin, SpHistin, SpALF4 (anti-lipopolysaccharide factor), SpALF5 and SpALF6) were significantly decreased in hemocytes. In our study, knockdown of SpCTL-B could down-regulate the expression of SpSTAT at m

  3. Use of a bacteriophage cocktail to control Salmonella in food and the food industry.

    Science.gov (United States)

    Spricigo, Denis Augusto; Bardina, Carlota; Cortés, Pilar; Llagostera, Montserrat

    2013-07-15

    The use of lytic bacteriophages for the biocontrol of food-borne pathogens in food and in the food industry is gaining increasing acceptance. In this study, the effectiveness of a bacteriophage cocktail composed of three different lytic bacteriophages (UAB_Phi 20, UAB_Phi78, and UAB_Phi87) was determined in four different food matrices (pig skin, chicken breasts, fresh eggs, and packaged lettuce) experimentally contaminated with Salmonella enterica serovar Typhimurium and S. enterica serovar Enteritidis. A significant bacterial reduction (>4 and 2 log/cm(2) for S. Typhimurium and S. Enteritidis, respectively; p≤0.005) was obtained in pig skin sprayed with the bacteriophage cocktail and then incubated at 33 °C for 6h. Significant decreases in the concentration of S. Typhimurium and S. Enteritidis were also measured in chicken breasts dipped for 5 min in a solution containing the bacteriophage cocktail and then refrigerated at 4 °C for 7 days (2.2 and 0.9 log10 cfu/g, respectively; p≤0.0001) as well as in lettuce similarly treated for 60 min at room temperature (3.9 and 2.2 log10 cfu/g, respectively; p≤0.005). However, only a minor reduction of the bacterial concentration (0.9 log10 cfu/cm(2) of S. Enteritidis and S. Typhimurium; p≤0.005) was achieved in fresh eggs sprayed with the bacteriophage cocktail and then incubated at 25 °C for 2 h. These results show the potential effectiveness of this bacteriophage cocktail as a biocontrol agent of Salmonella in several food matrices under conditions similar to those used in their production. Copyright © 2013 Elsevier B.V. All rights reserved.

  4. Rapid detection of microalbuminuria in diabetic patients by an agglutination inhibition test. Comparison with radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Giampietro, O; Miccoli, R; Di Palma, L; Bertolotto, A; Anichini, R; Navalesi, R; Clerico, A

    1986-01-01

    Subclinical elevation of urinary albumin excretion (UAlbE) early in the course of diabetes mellitus has been suggested to predict later clinical proteinuria and mortality. UAlbE is currently measured using radioimmunoassay (RIA) or radial immunodiffusion methods. However, these procedures are expensive and time-consuming and cannot be used as screening methods. Recently, an agglutination test (AT) has been suggested as a routinary method for the screening of microalbuminuria in diabetic patients. In this paper the results obtained are compared with an AT procedure and RIA method in a screening program of microproteinuria in diabetic patients. An immunological test (a latex agglutination assay) for the analysis of albuminura is used, which human albumin was adsorbed to latex beads (about 0.3 ..mu..l of a urine sample. Urine samples containing an albumin concentration >40 ..mu..g/ml were found to inhibit the agglutination of latex beads with antiserum. The RIA and AT results showed good agreement when urine samples were assayed soon after collection or after a short period of storage (less than or equal to3 weeks at -20 grade centigrades). The AT procedure has been adjusted in order to give a positive response (no agglutination) over the range of supranormal concentrations of urinary albumin (>40 ..mu..g/ml), which are on the other hand undetectable by Albustix. In addition, it is possible to perform a semiquantitative test using various dilutions of urine samples with albumin concentration > 40 ..mu..g/ml, so to estimate approximately the UAlbE. The AT method is simple, fast and specific, and has proved to be useful for the identification of diabetic patients at risk for developing clinical nephropathy. Therefore, it may be used in screening programs for diabetic microproteinuria. 21 refs.

  5. Pasteurella haemolytica bacteriophage: identification, partial characterization, and relationship of temperate bacteriophages from isolates of Pasteurella haemolytica (biotype A, serotype 1)

    International Nuclear Information System (INIS)

    Richards, A.B.; Renshaw, H.W.; Sneed, L.W.

    1985-01-01

    Pasteurella haemolytica (biotype A, serotype 1) isolates (n = 15) from the upper respiratory tract of clinically normal cattle, as well as from lung lesions from cases of fatal bovine pasteurellosis, were examined for the presence of bacteriophage after irradiation with UV light. Treatment of all P haemolytica isolates with UV irradiation resulted in lysis of bacteria due to the induction of vegetative development of bacteriophages. The extent of growth inhibition and bacterial lysis in irradiated cultures was UV dose-dependent. Bacterial cultures exposed to UV light for 20 s reached peak culture density between 60 and 70 minutes after irradiation; thereafter, culture density declined rapidly, so that by 120 minutes, it was approximately 60% of the original value. When examined ultrastructurally, lytic cultures from each isolate revealed bacteriophages with an overall length of approximately 200 nm and that appeared to have a head with icosahedral symmetry and a contractile tail. Cell-free filtrate from each noninduced bacterial isolate was inoculated onto the other bacterial isolates in a cross-culture sensitivity assay for the presence of phages lytic for the host bacterial isolates. Zones of lysis (plaques) did not develop when bacterial lawns grown from the different isolates were inoculated with filtrates from the heterologous isolates

  6. Replication of bacteriophage lambda DNA

    International Nuclear Information System (INIS)

    Tsurimoto, T.; Matsubara, K.

    1983-01-01

    In this paper results of studies on the mechanism of bacteriophage lambda replication using molecular biological and biochemical approaches are reported. The purification of the initiator proteins, O and P, and the role of the O and P proteins in the initiation of lambda DNA replication through interactions with specific DNA sequences are described. 47 references, 15 figures

  7. Nitric oxide production by hemocytes of larva and pharate prepupa of Galleria mellonella in response to bacterial lipopolysaccharide: cytoprotective or cytotoxic?

    Czech Academy of Sciences Publication Activity Database

    Krishnan, Natraj; Hyršl, P.; Šimek, V.

    2006-01-01

    Roč. 142, 1-2 (2006), s. 103-110 ISSN 1532-0456 Institutional research plan: CEZ:AV0Z50070508 Keywords : nitric oxide * hemocytes * lipopolysaccharide Subject RIV: ED - Physiology Impact factor: 1.991, year: 2006

  8. Proposed method for agglutinating antibody titer analysis and its use as indicator of acquired immunity in pacu, Piaractus mesopotamicus

    Directory of Open Access Journals (Sweden)

    JD Biller-Takahashi

    Full Text Available Antibody can be assessed by agglutinating antibody titer which is a quantitative measure of circulating antibodies in serum from fish previously immunized. The antibody evaluation has been performed with different fish species, and is considered a reliable method that can be applied to confirm several hypothesis regarding acquired immunity, even in conjunction with precise methods to describe immune mechanisms. In order to provide appropriate analytical methods for future studies on the specific immune system of native fish, the present study standardized on assay to measure the serum agglutinating antibody titer produced after immunization with inactivated A. hydrophila and levamisole administration in pacu. It was possible to determine the agglutinating antibodies titer in a satisfactorily way in pacu immunized with inactive A. hydrophila, and the highest titers were observed on fish fed with levamisole.

  9. BRED: a simple and powerful tool for constructing mutant and recombinant bacteriophage genomes.

    Directory of Open Access Journals (Sweden)

    Laura J Marinelli

    Full Text Available Advances in DNA sequencing technology have facilitated the determination of hundreds of complete genome sequences both for bacteria and their bacteriophages. Some of these bacteria have well-developed and facile genetic systems for constructing mutants to determine gene function, and recombineering is a particularly effective tool. However, generally applicable methods for constructing defined mutants of bacteriophages are poorly developed, in part because of the inability to use selectable markers such as drug resistance genes during viral lytic growth. Here we describe a method for simple and effective directed mutagenesis of bacteriophage genomes using Bacteriophage Recombineering of Electroporated DNA (BRED, in which a highly efficient recombineering system is utilized directly on electroporated phage DNA; no selection is required and mutants can be readily detected by PCR. We describe the use of BRED to construct unmarked gene deletions, in-frame internal deletions, base substitutions, precise gene replacements, and the addition of gene tags.

  10. Novel platelet-agglutinating protein from a thrombotic thrombocytopenic purpura plasma.

    OpenAIRE

    Siddiqui, F A; Lian, E C

    1985-01-01

    A novel platelet-agglutinating protein (PAP) was purified approximately 2,000-fold from the plasma of a patient with thrombotic thrombocytopenic purpura (TTP) by ammonium sulfate fractionation, DEAE-Sephacel and concanavalin A-Sepharose chromatographies. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with and without reduction, this preparation revealed a major protein band with a molecular weight of 37,000, and a minor band with a molecular weight of 32,000-34,000. After eluti...

  11. Engineered enzymatically active bacteriophages and methods of uses thereof

    Energy Technology Data Exchange (ETDEWEB)

    Collins, James J [Newton, MA; Kobayashi, Hideki [Yokohama, JP; Kearn, Mads [Ottawa, CA; Araki, Michihiro [Minatoku, JP; Friedland, Ari [Boston, MA; Lu, Timothy Kuan-Ta [Palo Alto, CA

    2012-05-22

    The present invention provides engineered bacteriophages that express at least one biofilm degrading enzyme on their surface and uses thereof for degrading bacterial biofilms. The invention also provides genetically engineered bacteriophages expressing the biofilm degrading enzymes and proteins necessary for the phage to replicate in different naturally occurring biofilm producing bacteria. The phages of the invention allow a method of biofilm degradation by the use of one or only a few administration of the phage because the system using these phages is self perpetuating, and capable of degrading biofilm even when the concentration of bacteria within the biofilm is low.

  12. A Bacteriophage-Related Chimeric Marine Virus Infecting Abalone

    Science.gov (United States)

    Zhuang, Jun; Cai, Guiqin; Lin, Qiying; Wu, Zujian; Xie, Lianhui

    2010-01-01

    Marine viruses shape microbial communities with the most genetic diversity in the sea by multiple genetic exchanges and infect multiple marine organisms. Here we provide proof from experimental infection that abalone shriveling syndrome-associated virus (AbSV) can cause abalone shriveling syndrome. This malady produces histological necrosis and abnormally modified macromolecules (hemocyanin and ferritin). The AbSV genome is a 34.952-kilobase circular double-stranded DNA, containing putative genes with similarity to bacteriophages, eukaryotic viruses, bacteria and endosymbionts. Of the 28 predicted open reading frames (ORFs), eight ORF-encoded proteins have identifiable functional homologues. The 4 ORF products correspond to a predicted terminase large subunit and an endonuclease in bacteriophage, and both an integrase and an exonuclease from bacteria. The other four proteins are homologous to an endosymbiont-derived helicase, primase, single-stranded binding (SSB) protein, and thymidylate kinase, individually. Additionally, AbSV exhibits a common gene arrangement similar to the majority of bacteriophages. Unique to AbSV, the viral genome also contains genes associated with bacterial outer membrane proteins and may lack the structural protein-encoding ORFs. Genomic characterization of AbSV indicates that it may represent a transitional form of microbial evolution from viruses to bacteria. PMID:21079776

  13. A bacteriophage-related chimeric marine virus infecting abalone.

    Directory of Open Access Journals (Sweden)

    Jun Zhuang

    Full Text Available Marine viruses shape microbial communities with the most genetic diversity in the sea by multiple genetic exchanges and infect multiple marine organisms. Here we provide proof from experimental infection that abalone shriveling syndrome-associated virus (AbSV can cause abalone shriveling syndrome. This malady produces histological necrosis and abnormally modified macromolecules (hemocyanin and ferritin. The AbSV genome is a 34.952-kilobase circular double-stranded DNA, containing putative genes with similarity to bacteriophages, eukaryotic viruses, bacteria and endosymbionts. Of the 28 predicted open reading frames (ORFs, eight ORF-encoded proteins have identifiable functional homologues. The 4 ORF products correspond to a predicted terminase large subunit and an endonuclease in bacteriophage, and both an integrase and an exonuclease from bacteria. The other four proteins are homologous to an endosymbiont-derived helicase, primase, single-stranded binding (SSB protein, and thymidylate kinase, individually. Additionally, AbSV exhibits a common gene arrangement similar to the majority of bacteriophages. Unique to AbSV, the viral genome also contains genes associated with bacterial outer membrane proteins and may lack the structural protein-encoding ORFs. Genomic characterization of AbSV indicates that it may represent a transitional form of microbial evolution from viruses to bacteria.

  14. Pecularities of mutagenesis of T4Br bacteriophage under the direct and indirect radiation effects

    International Nuclear Information System (INIS)

    Yurov, S.S.

    1975-01-01

    Different lethal and mutagenic effects were shown when bacteriophage T4Br + (470 r/min) was irradiated in broth (direct effect) and a buffer solution (direct and indirect action). The survival rate of the bacteriophage in the buffer solution was 0.1 percent for a dose rate of 60 kr; in the broth it was 10 percent. The frequency of mutation of the bacteriophage also showed the greater effect of the irradiation in the buffer solution than in the broth (25 and 5 r-mutants respectively at a dose rate of 10 kr). An analysis of the ratio of the r-groups when the bacteriophage was treated in various ways revealed differences between mutagenesis produced in the broth and the buffer, and spontaneous mutagenesis. (V.A.P.)

  15. Removal of endotoxins from bacteriophage preparations by extraction with organic solvents.

    Directory of Open Access Journals (Sweden)

    Bożena Szermer-Olearnik

    Full Text Available Lipopolysaccharide (LPS, endotoxin, pyrogen constitutes a very troubling contaminant of crude phage lysates produced in Gram-negative bacteria. Toxicity of LPS depends on the strong innate immunity response including the cytokines. Therefore, its removal is important for bacteriophage applications. In this paper, we present a procedure for extractive removal of endotoxin from bacteriophage preparations with water immiscible solvents (1-octanol or 1-butanol. During extraction most of the phage lytic activity is retained in the aqueous phase, while endotoxin accumulates in the organic solvent. The levels of endotoxin (expressed as endotoxin units, EU in the aqueous bacteriophage-containing fraction determined by limulus amebocyte lysate or EndoLISA assay were exceptionally low. While the initial endotoxin levels in the crude phage lysates ranged between 10(3 and 10(5 EU/ml the average level after organic extraction remaining in the aqueous fraction was 5.3 EU/ml. These values when related to phage titers decreased from 10(3-10(5 EU/10(9 PFU (plaque forming units down to an average of 2.8 EU/10(9 PFU. The purification procedure is scalable, efficient and applicable to all the bacteriophages tested: T4, HAP1 (E. coli and F8 (P. aeruginosa.

  16. Effectiveness of cooking to reduce norovirus and infectious F-specific RNA bacteriophage concentrations in Mytilus edulis.

    Science.gov (United States)

    Flannery, J; Rajko-Nenow, P; Winterbourn, J B; Malham, S K; Jones, D L

    2014-08-01

    The aim of this study was to determine if domestic cooking practices can reduce concentrations of norovirus (NoV) and F-specific RNA (FRNA) bacteriophage in experimentally contaminated mussels. Mussels (n = 600) contaminated with NoV and FRNA bacteriophage underwent four different cooking experiments performed in triplicate at ~70°C and >90°C. Concentrations of infectious FRNA bacteriophage (using a plaque assay) were compared with concentrations of FRNA bacteriophage and NoV determined using a standardised RT-qPCR. Initial concentrations of infectious FRNA bacteriophage (7·05 log10  PFU g(-1) ) in mussels were not significantly reduced in simmering water (~70°C); however, cooking at higher temperatures (>90°C) reduced infectious FRNA bacteriophage to undetected levels within 3 min. Further investigation determined the time required for a 1-log reduction of infectious FRNA bacteriophage at 90°C to be 42 s therefore a >3-log reduction in infectious virus can be obtained by heating mussel digestive tissue to 90°C for 126 s. Domestic cooking practices based on shell opening alone do not inactivate infectious virus in mussels, however, cooking mussels at high temperatures is effective to reduce infectious virus concentrations and the risk of illness in consumers. The data will contribute towards evidence-based cooking recommendations for shellfish to provide a safe product for human consumption. © 2014 The Society for Applied Microbiology.

  17. M13 Bacteriophage Based Protein Sensors

    Science.gov (United States)

    Lee, Ju Hun

    Despite significant progress in biotechnology and biosensing, early detection and disease diagnosis remains a critical issue for improving patient survival rates and well-being. Many of the typical detection schemes currently used possess issues such as low sensitivity and accuracy and are also time consuming to run and expensive. In addition, multiplexed detection remains difficult to achieve. Therefore, developing advanced approaches for reliable, simple, quantitative analysis of multiple markers in solution that also are highly sensitive are still in demand. In recent years, much of the research has primarily focused on improving two key components of biosensors: the bio-recognition agent (bio-receptor) and the transducer. Particular bio-receptors that have been used include antibodies, aptamers, molecular imprinted polymers, and small affinity peptides. In terms of transducing agents, nanomaterials have been considered as attractive candidates due to their inherent nanoscale size, durability and unique chemical and physical properties. The key focus of this thesis is the design of a protein detection and identification system that is based on chemically engineered M13 bacteriophage coupled with nanomaterials. The first chapter provides an introduction of biosensors and M13 bacteriophage in general, where the advantages of each are provided. In chapter 2, an efficient and enzyme-free sensor is demonstrated from modified M13 bacteriophage to generate highly sensitive colorimetric signals from gold nanocrystals. In chapter 3, DNA conjugated M13 were used to enable facile and rapid detection of antigens in solution that also provides modalities for identification. Lastly, high DNA loadings per phage was achieved via hydrozone chemistry and these were applied in conjunction with Raman active DNA-gold/silver core/shell nanoparticles toward highly sensitive SERS sensing.

  18. Direct Quantitative Detection and Identification of Lactococcal Bacteriophages from Milk and Whey by Real-Time PCR: Application for the Detection of Lactococcal Bacteriophages in Goat's Raw Milk Whey in France

    Directory of Open Access Journals (Sweden)

    Mai Huong Ly-Chatain

    2011-01-01

    Full Text Available The presence of Lactococcus bacteriophages in milk can partly or completely inhibit milk fermentation. To prevent the problems associated with the bacteriophages, the real-time PCR was developed in this study for direct detection from whey and milk of three main groups of Lactococcus bacteriophages, c2, 936, and P335. The optimization of DNA extraction protocol from complex matrices such as whey and milk was optimized allowed the amplification of PCR without any matrix and nontarget contaminant interference. The real-time PCR program was specific and with the detection limit of 102 PFU/mL. The curve slopes were −3.49, −3.69, and −3.45 with the amplification efficiency estimated at 94%, 94%, and 98% and the correlation coefficient (2 of 0.999, 0.999, and 0.998 for c2, 936 and P335 group, respectively. This method was then used to detect the bacteriophages in whey and goat's raw milk coming from three farms located in the Rhône-Alpes region (France.

  19. In vitro Effectiveness of Commercial Bacteriophage Cocktails on Diverse Extended-Spectrum Beta-Lactamase Producing Escherichia coli Strains.

    Science.gov (United States)

    Gundogdu, Aycan; Bolkvadze, Darajen; Kilic, Huseyin

    2016-01-01

    The objective of this study is to determine the in vitro susceptibility of Georgian bacteriophage cocktails on multidrug resistant (MDR) extended-spectrum beta-lactamase producing Escherichia coli (ESBL-EC) isolated from patients' blood and urine cultures. A total of 615 E. coli isolates were included in this study. Phene Plate (PhP)-typing and phylogenetic grouping were used for the typing. Antimicrobial resistance profiles and ESBL production of all isolates were confirmed according to Clinical and Laboratory Standards Institute (CLSI) criteria. The activities of four bacteriophage cocktails (Enko-phage, SES-bacteriophage, Pyo-bacteriophage, and Intesti-bacteriophage) were determined against 142 ESBL-EC using in vitro spot tests. According to this, Enko-phage were active against 87.3% of the tested strains while that ratio was 81.7% for Intesti-bacteriophage, 81.7% for Pyo-bacteriophage, and 59.2% for SES-bacteriophage cocktails. Based on the contingency tests, the phage cocktails were observed to be statistically significantly ( p < 0.001) more effective on ESBL-EC strains belonging to phylogenetic groups D and B2. The employed phage cocktails were found to be affective against all tested resistant types. These results are promising especially for the infections that are caused by MDR pathogens that are difficult to treat. As this is a preliminary step to the potential clinical trials to be designed for the country, in vitro confirmation of their success on a MDR ESBL-EC collection should be accepted as an initial action, which is encouraging to consider clinical trials of phage therapy especially in countries which are not introduce phage therapy.

  20. A comparison of sperm agglutination and immobilization assays with a quantitative ELISA for anti-sperm antibody in serum.

    Science.gov (United States)

    Lynch, D M; Leali, B A; Howe, S E

    1986-08-01

    An enzyme-linked immunosorbent assay (ELISA) that quantitates antisperm antibody in serum was compared with standard sperm agglutination and immobilization assays with the use of sera from 40 normal and 292 subfertile individuals. Quantitation of the assay was accomplished by standardizing assay parameters, including the incorporation of a standard reference curve, the number of whole target sperm, the optimal dilution of serum, the selection of microtiter plate, and the time and temperatures involved in the adsorption and incubation phases. With this method, the level of antisperm antibody binding to target sperm in 40 normal fertile individuals was found to be 2.3 (+/- 1.1 standard deviation [SD]) fg immunoglobulin (Ig)/sperm. An increased mean level of 7.4 +/- 3.7 fg Ig/sperm was determined in 84 infertile patients with positive agglutination and/or immobilization tests. In 208 individuals with negative agglutination and immobilization tests the mean concentration of antisperm antibody was 2.5 +/- 1.3 fg Ig/sperm. Postvasectomy patients assayed by this method had a mean Ig binding value of 7.1 +/- 2.4 fg Ig/sperm. The infertile group with positive agglutination and/or immobilization tests had a significantly higher mean antisperm antibody level than the normal fertile group, according to the Student's t-test for independent samples (P less than 0.001). This indirect serum-based assay reproducibly quantitates antisperm antibody binding to whole target sperm, suggests the normal and abnormal levels of antisperm antibody, and correlates with standard functional assays.

  1. Guidelines for Bacteriophage Product Certification.

    Science.gov (United States)

    Fauconnier, Alan

    2018-01-01

    Following decades in the wilderness, bacteriophage therapy is now appearing as a credible antimicrobial strategy. However, this reemerging therapy does not rekindle without raising sensitive regulatory concerns. Indeed, whereas the European regulatory framework has been basically implemented to tackle ready-to-use pharmaceuticals produced on a large scale, bacteriophage therapy relies on a dynamic approach requiring a regulation on personalized medicine, nonexistent at present. Because of this, no guideline are currently available for addressing the scientific and regulatory issues specifically related to phage therapy medicinal products (PTMP).Pending to the implementation of an appropriate regulatory framework and to the development of ensuing guidelines, several avenues which might lead to PTMP regulatory compliance are explored here. Insights might come from the multi-strain dossier approach set up for particular animal vaccines, from the homologous group concept developed for the allergen products or from the licensing process for veterinary autogenous vaccines. Depending on national legislations, customized preparations prescribed as magistral formulas or to be used on a named-patient basis are possible regulatory approaches to be considered. However, these schemes are not optimal and should thus be regarded as transitional.

  2. Multiplex PCR for the detection and identification of dairy bacteriophages in milk.

    Science.gov (United States)

    del Rio, B; Binetti, A G; Martín, M C; Fernández, M; Magadán, A H; Alvarez, M A

    2007-02-01

    Bacteriophage infections of starter lactic acid bacteria are a serious risk in the dairy industry. Phage infection can lead to slow lactic acid production or even the total failure of fermentation. The associated economic losses can be substantial. Rapid and sensitive methods are therefore required to detect and identify phages at all stages of the manufacture of fermented dairy products. This study describes a simple and rapid multiplex PCR method that, in a single reaction, detects the presence of bacteriophages infecting Streptococcus thermophilus and Lactobacillus delbrueckii, plus three genetically distinct 'species' of Lactococcus lactis phages commonly found in dairy plants (P335, 936 and c2). Available bacteriophage genome sequences were examined and the conserved regions used to design five pairs of primers, one for each of the above bacteriophage species. These primers were designed to generate specific fragments of different size depending on the species. Since this method can detect the above phages in untreated milk and can be easily incorporated into dairy industry routines, it might be readily used to earmark contaminated milk for use in processes that do not involve susceptible starter organisms or for use in those that involve phage-deactivating conditions.

  3. Two-Stage Dynamics of In Vivo Bacteriophage Genome Ejection

    Science.gov (United States)

    Chen, Yi-Ju; Wu, David; Gelbart, William; Knobler, Charles M.; Phillips, Rob; Kegel, Willem K.

    2018-04-01

    Biopolymer translocation is a key step in viral infection processes. The transfer of information-encoding genomes allows viruses to reprogram the cell fate of their hosts. Constituting 96% of all known bacterial viruses [A. Fokine and M. G. Rossmann, Molecular architecture of tailed double-stranded DNA phages, Bacteriophage 4, e28281 (2014)], the tailed bacteriophages deliver their DNA into host cells via an "ejection" process, leaving their protein shells outside of the bacteria; a similar scenario occurs for mammalian viruses like herpes, where the DNA genome is ejected into the nucleus of host cells, while the viral capsid remains bound outside to a nuclear-pore complex. In light of previous experimental measurements of in vivo bacteriophage λ ejection, we analyze here the physical processes that give rise to the observed dynamics. We propose that, after an initial phase driven by self-repulsion of DNA in the capsid, the ejection is driven by anomalous diffusion of phage DNA in the crowded bacterial cytoplasm. We expect that this two-step mechanism is general for phages that operate by pressure-driven ejection, and we discuss predictions of our theory to be tested in future experiments.

  4. Two-Stage Dynamics of In Vivo Bacteriophage Genome Ejection

    Directory of Open Access Journals (Sweden)

    Yi-Ju Chen

    2018-05-01

    Full Text Available Biopolymer translocation is a key step in viral infection processes. The transfer of information-encoding genomes allows viruses to reprogram the cell fate of their hosts. Constituting 96% of all known bacterial viruses [A. Fokine and M. G. Rossmann, Molecular architecture of tailed double-stranded DNA phages, Bacteriophage 4, e28281 (2014], the tailed bacteriophages deliver their DNA into host cells via an “ejection” process, leaving their protein shells outside of the bacteria; a similar scenario occurs for mammalian viruses like herpes, where the DNA genome is ejected into the nucleus of host cells, while the viral capsid remains bound outside to a nuclear-pore complex. In light of previous experimental measurements of in vivo bacteriophage λ ejection, we analyze here the physical processes that give rise to the observed dynamics. We propose that, after an initial phase driven by self-repulsion of DNA in the capsid, the ejection is driven by anomalous diffusion of phage DNA in the crowded bacterial cytoplasm. We expect that this two-step mechanism is general for phages that operate by pressure-driven ejection, and we discuss predictions of our theory to be tested in future experiments.

  5. Gene expression profile of Bombyx mori hemocyte under the stress of destruxin A.

    Directory of Open Access Journals (Sweden)

    Liang Gong

    Full Text Available Destruxin A (DA is a cyclo-peptidic mycotoxin from the entomopathogenic fungus Metarhizium anisopliae. To uncover potential genes associated with its molecular mechanisms, a digital gene expression (DGE profiling analysis was used to compare differentially expressed genes in the hemocytes of silkworm larvae treated with DA. Ten DGE libraries were constructed, sequenced, and assembled, and the unigenes with least 2.0-fold difference were further analyzed. The numbers of up-regulated genes were 10, 20, 18, 74 and 8, as well as the numbers of down-regulated genes were 0, 1, 8, 13 and 3 at 1, 4, 8, 12 and 24 h post treatment, respectively. Totally, the expression of 132 genes were significantly changed, among them, 1, 3 and 12 genes were continually up-regulated at 4, 3 and 2 different time points, respectively, while 1 gene was either up or down-regulated continually at 2 different time points. Furthermore, 68 genes were assigned to one or multiple gene ontology (GO terms and 89 genes were assigned to specific Kyoto Encyclopedia of Genes and Genomes (KEGG Orthology. In-depth analysis identified that these genes putatively involved in insecticide resistance, cell apoptosis, and innate immune defense. Finally, twenty differentially expressed genes were randomly chosen and validated by quantitative real-time PCR (qRT-PCR. Our studies provide insights into the toxic effect of this microbial insecticide on silkworm's hemocytes, and are helpful to better understanding of the molecular mechanisms of DA as a biological insecticide.

  6. Use of the integration elements encoded by the temperate lactococcal bacteriophage TP901-1

    DEFF Research Database (Denmark)

    Brøndsted, Lone; Hammer, Karin

    1999-01-01

    Previously we showed that only one phage-expressed protein (Orf1), a 425-bp region upstream of the orf1 gene (presumably encoding a promoter), and the attP region are necessary and also sufficient for integration of the bacteriophage TP901-1 genome into the chromosome of Lactococcus lactis subsp......P region seem to be necessary for site-specific integration of the temperate bacteriophage TP901-1. By use of the integrative elements (attP and orf1) expressed by the temperate lactococcal bacteriophage TP901-1, a system for obtaining stable chromosomal single-copy transcriptional fusions in L. lactis...

  7. Bacteriophage-antibiotic synergism to control planktonic and biofilm ...

    African Journals Online (AJOL)

    Bacteriophage-antibiotic synergism to control planktonic and biofilm producing clinical isolates of Pseudomonas aeruginosa. Amina Amal Mahmoud Nouraldin, Manal Mohammad Baddour, Reem Abdel Hameed Harfoush, Sara AbdelAziz Mohamed Essa ...

  8. Document Categorization with Modified Statistical Language Models for Agglutinative Languages

    Directory of Open Access Journals (Sweden)

    Tantug

    2010-11-01

    Full Text Available In this paper, we investigate the document categorization task with statistical language models. Our study mainly focuses on categorization of documents in agglutinative languages. Due to the productive morphology of agglutinative languages, the number of word forms encountered in naturally occurring text is very large. From the language modeling perspective, a large vocabulary results in serious data sparseness problems. In order to cope with this drawback, previous studies in various application areas suggest modified language models based on different morphological units. It is reported that performance improvements can be achieved with these modified language models. In our document categorization experiments, we use standard word form based language models as well as other modified language models based on root words, root words and part-of-speech information, truncated word forms and character sequences. Additionally, to find an optimum parameter set, multiple tests are carried out with different language model orders and smoothing methods. Similar to previous studies on other tasks, our experimental results on categorization of Turkish documents reveal that applying linguistic preprocessing steps for language modeling provides improvements over standard language models to some extent. However, it is also observed that similar level of performance improvements can also be acquired by simpler character level or truncated word form models which are language independent.

  9. Coronatin-2 from the entomopathogenic fungus Conidiobolus coronatus kills Galleria mellonella larvae and incapacitates hemocytes.

    Science.gov (United States)

    Boguś, M I; Wieloch, W; Ligęza-Żuber, M

    2017-02-01

    Coronatin-2, a 14.5 kDa protein, was isolated from culture filtrates of the entomopathogenic fungus Conidiobolus coronatus (Costantin) Batko (Entomophthoramycota: Entomophthorales). After LC-MS/MS (liquid chromatography tandem mass spectrometry) analysis of the tryptic peptide digest of coronatin-2 and a mass spectra database search no orthologs of this protein could be found in fungi. The highest homology was observed to the partial translation elongation factor 1a from Sphaerosporium equinum (protein sequence coverage, 21%), with only one peptide sequence, suggesting that coronatin-2 is a novel fungal protein that has not yet been described. In contrast to coronatin-1, an insecticidal 36 kDa protein, which shows both elastolytic and chitinolytic activity, coronatin-2 showed no enzymatic activity. Addition of coronatin-2 into cultures of hemocytes taken from larvae of Galleria mellonella Linnaeus (Lepidoptera: Pyralidae), resulted in progressive disintegration of nets formed by granulocytes and plasmatocytes due to rapid degranulation of granulocytes, extensive vacuolization of plasmatocytes accompanied by cytoplasm expulsion, and cell disintegration. Spherulocytes remained intact, while oenocytes rapidly disintegrated. Coronatin-2 produced 80% mortality when injected into G. mellonella at 5 µg larva-1. Further study is warranted to determine the relevance of the acute toxicity of coronatin-2 and its effects on hemocytes in vitro to virulence of C. coronatus against its hosts.

  10. Comparative venom toxicity between Pteromalus puparum and Nasonia vitripennis (Hymenoptera: Pteromalidae) toward the hemocytes of their natural hosts, non-target insects and cultured insect cells.

    Science.gov (United States)

    Zhang, Zhong; Ye, Gong-Yin; Cai, Jun; Hu, Cui

    2005-09-01

    Crude venoms from two parasitoid species, Pteromalus puparum and Nasonia vitripennis (Hymenoptera: Pteromalidae) were assayed for biological activities toward hemocytes from two species of their natural hosts and eight species of their non-natural hosts as well as two lines of cultured Lepidoptera cells, respectively. By inhibiting the spreading and viability of insect hemocytes, the venom from P. puparum displayed significantly higher activities toward plasmatocytes and granular cells from both larvae and pupae of two natural hosts, Pieris rapae and Papilio xuthus, and lower activity toward those from Spodoptera litura, Musca domestica and Sarcophaga peregrina. However, no effect was found towards any type of hemocytes from other five insects tested, namely, Ectropis oblique, Galleria mellonella, Sesamia inferens, Bombyx mori and Parnara guttata. In contrast, the venom from N. vitripennis showed a narrower range of targeted insects. It appeared to have highly adverse effects on the spreading and viability of plasmatocytes and granular cells only from the natural hosts, M. domestica and S. peregrina, little toxicity to cells from P. rapae and P. xuthus, and no effect on any of the other insects tested. Pteromalus puparum venom also apparently presented a high ability to block the spreading of Tn-5B1-4 cells derived from Trichoplusia ni, and high cytotoxicity to the cells and Ha cells derived from Helicoverpa armigera. Nasonia vitripennis venom, however, only had a marked lethal effect to Ha cells. In addition, the possibility that the host range of a defined parasitoid could be assessed using our method of treating hemocytes from candidate insects with venom in vitro, and the potential of our venoms tested in the development of bio-insecticides, insect-resistant transgenic plants, are discussed.

  11. in vitro effectiveness of commercial bacteriophage cocktails on diverse extended spectrum beta-lactamase (ESBL producing Escherichia coli strains

    Directory of Open Access Journals (Sweden)

    Aycan Gundogdu

    2016-11-01

    Full Text Available The objective of this study is to determine the in vitro susceptibility of Georgian bacteriophage cocktails on multi-drug resistant extended-spectrum β-lactamase producing Escherichia coli (ESBL-EC isolated from patients' blood and urine cultures. 615 E. coli isolates were included in this study. PhP-typing and phylogenetic grouping were used for the typing. Antimicrobial resistance profiles and ESBL production of all isolates were confirmed according to CLSI criteria. The activities of four bacteriophage cocktails (Enko-phage, SES-bacteriophage, Pyo-bacteriophage and Intesti-bacteriophage were determined against 142 ESBL- EC using in vitro spot tests. According to this, Enko-phage were active against 87.3% of the tested strains while that ratio was 81.7% for intesti-bacteriophage, 81.7% for Pyo-bacteriophage and 59.2% for SES-bacteriophage cocktails. Based on the contingency tests, the phage cocktails were observed to be statistically significantly (p<0.001 more effective on ESBL-EC strains belonging to phylogenetic groups D and B2. The employed phage cocktails were found to be affective against all tested resistant types. These results are promising especially for the infections that are caused by multi-drug resistant pathogens that are difficult to treat. As this is a preliminary step to the potential clinical trials to be designed for the country, in vitro confirmation of their success on a multi-drug-resistant ESBL-EC collection should be accepted as an initial action, which is encouraging to consider clinical trials of phage therapy especially in countries which are not introduce phage therapy.

  12. Encapsulation Strategies of Bacteriophage (Felix O1) for Oral Therapeutic Application.

    Science.gov (United States)

    Islam, Golam S; Wang, Qi; Sabour, Parviz M

    2018-01-01

    Due to emerging antibiotic-resistant strains among the pathogens, a variety of strategies, including therapeutic application of bacteriophages, have been suggested as a possible alternative to antibiotics in food animal production. As pathogen-specific biocontrol agents, bacteriophages are being studied intensively. Primarily their applications in the food industry and animal production have been recognized in the USA and Europe, for pathogens including Salmonella, Campylobacter, Escherichia coli, and Listeria. However, the viability of orally administered phage may rapidly reduce under the harsh acidic conditions of the stomach, presence of enzymes and bile. It is evident that bacteriophages, intended for phage therapy by oral administration, require efficient protection from the acidic environment of the stomach and should remain active in the animal's gastrointestinal tract where pathogen colonizes. Encapsulation of phages by spray drying or extrusion methods can protect phages from the simulated hostile gut conditions and help controlled release of phages to the digestive system when appropriate formulation strategy is implemented.

  13. Bacteriophage use to control Salmonella biofilm on surfaces present in chicken slaughterhouses.

    Science.gov (United States)

    Garcia, Keila Carolina de Ornellas Dutka; Corrêa, Isadora Mainieri de Oliveira; Pereira, Larissa Quinto; Silva, Tarcísio Macedo; Mioni, Mateus de Souza Ribeiro; Izidoro, Ana Carolina de Moraes; Bastos, Igor Henrique Vellano; Gonçalves, Guilherme Augusto Marietto; Okamoto, Adriano Sakai; Andreatti Filho, Raphael Lucio

    2017-09-01

    Foodborne diseases represent a major risk to public health worldwide. Pathogenic bacteria can live in the form of biofilm within the food industry, providing a permanent source of contamination. The aim of this study was to evaluate the influence of the types of adhesion surfaces on Salmonella biofilm formation at eight different times, and analyze the action time of a bacteriophage pool on established biofilms. Most of the samples used were classified as weak biofilm producers, with serovars Enteritidis and Heidelberg showing the highest frequency of biofilm formation. Glass and stainless steel surfaces significantly favored biofilm formation at 60 and 36 h of incubation respectively, but the polyvinyl chloride surface did not favor biofilm production, suggesting that the type of material may interfere with production. The bacteriophage pool action period focused on 3 h, but treatment of 9 h on glass surface biofilms was superior to other treatments because it affected the largest number of samples. These results suggests that some surface types and Salmonella serotypes may promote biofilm formation and indicate bacteriophages as an alternative to control biofilms. But further studies are required to prove the effectiveness and safety of bacteriophage therapy as an alternative in the antimicrobial control in the processing plants. © 2017 Poultry Science Association Inc.

  14. Bacteriophage-antibiotic synergism to control planktonic and biofilm ...

    African Journals Online (AJOL)

    Amina Amal Mahmoud Nouraldin

    2015-07-11

    Jul 11, 2015 ... mote resistance to antimicrobial agents, and its occurrence during the infectious ... Biofilm is a structured community of bacterial cells adher- ent to an inert or ..... biofilms with bacteriophages and chlorine. Biotechnol Bioeng.

  15. MetaPhinder-Identifying Bacteriophage Sequences in Metagenomic Data Sets

    DEFF Research Database (Denmark)

    Jurtz, Vanessa Isabell; Villarroel, Julia; Lund, Ole

    2016-01-01

    genome structure of many bacteriophages. The method is demonstrated to outperform both BLAST methods based on single hits and methods based on k-mer comparisons. MetaPhinder is available as a web service at the Center for Genomic Epidemiology https://cge.cbs.dtu.dk/services/MetaPhinder/, while the source...... and understand them. Here we present MetaPhinder, a method to identify assembled genomic fragments (i.e. contigs) of phage origin in metage-nomic data sets. The method is based on a comparison to a database of whole genome bacteriophage sequences, integrating hits to multiple genomes to accomodate for the mosaic...... code can be downloaded from https://bitbucket.org/genomicepidemiology/metaphinder or https://github.com/vanessajurtz/MetaPhinder....

  16. De novo transcriptome sequencing of the Octopus vulgaris hemocytes using Illumina RNA-Seq technology: response to the infection by the gastrointestinal parasite Aggregata octopiana.

    Science.gov (United States)

    Castellanos-Martínez, Sheila; Arteta, David; Catarino, Susana; Gestal, Camino

    2014-01-01

    Octopus vulgaris is a highly valuable species of great commercial interest and excellent candidate for aquaculture diversification; however, the octopus' well-being is impaired by pathogens, of which the gastrointestinal coccidian parasite Aggregata octopiana is one of the most important. The knowledge of the molecular mechanisms of the immune response in cephalopods, especially in octopus is scarce. The transcriptome of the hemocytes of O. vulgaris was de novo sequenced using the high-throughput paired-end Illumina technology to identify genes involved in immune defense and to understand the molecular basis of octopus tolerance/resistance to coccidiosis. A bi-directional mRNA library was constructed from hemocytes of two groups of octopus according to the infection by A. octopiana, sick octopus, suffering coccidiosis, and healthy octopus, and reads were de novo assembled together. The differential expression of transcripts was analysed using the general assembly as a reference for mapping the reads from each condition. After sequencing, a total of 75,571,280 high quality reads were obtained from the sick octopus group and 74,731,646 from the healthy group. The general transcriptome of the O. vulgaris hemocytes was assembled in 254,506 contigs. A total of 48,225 contigs were successfully identified, and 538 transcripts exhibited differential expression between groups of infection. The general transcriptome revealed genes involved in pathways like NF-kB, TLR and Complement. Differential expression of TLR-2, PGRP, C1q and PRDX genes due to infection was validated using RT-qPCR. In sick octopuses, only TLR-2 was up-regulated in hemocytes, but all of them were up-regulated in caecum and gills. The transcriptome reported here de novo establishes the first molecular clues to understand how the octopus immune system works and interacts with a highly pathogenic coccidian. The data provided here will contribute to identification of biomarkers for octopus resistance against

  17. Detection of bacteriophage-infected cells of Lactococcus lactis using flow cytometry

    DEFF Research Database (Denmark)

    Michelsen, Ole; Cuesta-Dominguez, Álvaro; Albrektsen, Bjarne

    2007-01-01

    Bacteriophage infection in dairy fermentation constitutes a serious problem worldwide. We have studied bacteriophage infection in Lactococcus lactis by using the flow cytometer. The first effect of the infection of the bacterium is a change from cells in chains toward single cells. We interpret...... describe a new method for detection of phage infection in Lactococcus lactis dairy cultures. The method is based on flow cytometric detection of cells with low-density cell walls. The method allows fast and early detection of phage-infected bacteria, independently of which phage has infected the culture...

  18. The Silencing of a 14-3-3ɛ Homolog in Tenebrio molitor Leads to Increased Antimicrobial Activity in Hemocyte and Reduces Larval Survivability.

    Science.gov (United States)

    Seo, Gi Won; Jo, Yong Hun; Seong, Jeong Hwan; Park, Ki Beom; Patnaik, Bharat Bhusan; Tindwa, Hamisi; Kim, Sun-Am; Lee, Yong Seok; Kim, Yu Jung; Han, Yeon Soo

    2016-08-20

    The 14-3-3 family of phosphorylated serine-binding proteins acts as signaling molecules in biological processes such as metabolism, division, differentiation, autophagy, and apoptosis. Herein, we report the requirement of 14-3-3ɛ isoform from Tenebrio molitor (Tm14-3-3ɛ) in the hemocyte antimicrobial activity. The Tm14-3-3ɛ transcript is 771 nucleotides in length and encodes a polypeptide of 256 amino acid residues. The protein has the typical 14-3-3 domain, the nuclear export signal (NES) sequence, and the peptide binding residues. The Tm14-3-3ɛ transcript shows a significant three-fold expression in the hemocyte of T. molitor larvae when infected with Escherichia coli Tm14-3-3ɛ silenced larvae show significantly lower survival rates when infected with E. coli. Under Tm14-3-3ɛ silenced condition, a strong antimicrobial activity is elicited in the hemocyte of the host inoculated with E. coli. This suggests impaired secretion of antimicrobial peptides (AMP) into the hemolymph. Furthermore, a reduction in AMP secretion under Tm14-3-3ɛ silenced condition would be responsible for loss in the capacity to kill bacteria and might explain the reduced survivability of the larvae upon E. coli challenge. This shows that Tm14-3-3ɛ is required to maintain innate immunity in T. molitor by enabling antimicrobial secretion into the hemolymph and explains the functional specialization of the isoform.

  19. The Silencing of a 14-3-3ɛ Homolog in Tenebrio molitor Leads to Increased Antimicrobial Activity in Hemocyte and Reduces Larval Survivability

    Directory of Open Access Journals (Sweden)

    Gi Won Seo

    2016-08-01

    Full Text Available The 14-3-3 family of phosphorylated serine-binding proteins acts as signaling molecules in biological processes such as metabolism, division, differentiation, autophagy, and apoptosis. Herein, we report the requirement of 14-3-3ɛ isoform from Tenebrio molitor (Tm14-3-3ɛ in the hemocyte antimicrobial activity. The Tm14-3-3ɛ transcript is 771 nucleotides in length and encodes a polypeptide of 256 amino acid residues. The protein has the typical 14-3-3 domain, the nuclear export signal (NES sequence, and the peptide binding residues. The Tm14-3-3ɛ transcript shows a significant three-fold expression in the hemocyte of T. molitor larvae when infected with Escherichia coli Tm14-3-3ɛ silenced larvae show significantly lower survival rates when infected with E. coli. Under Tm14-3-3ɛ silenced condition, a strong antimicrobial activity is elicited in the hemocyte of the host inoculated with E. coli. This suggests impaired secretion of antimicrobial peptides (AMP into the hemolymph. Furthermore, a reduction in AMP secretion under Tm14-3-3ɛ silenced condition would be responsible for loss in the capacity to kill bacteria and might explain the reduced survivability of the larvae upon E. coli challenge. This shows that Tm14-3-3ɛ is required to maintain innate immunity in T. molitor by enabling antimicrobial secretion into the hemolymph and explains the functional specialization of the isoform.

  20. Isolation and in vitro evaluation of bacteriophages against MDR-bacterial isolates from septic wound infections.

    Directory of Open Access Journals (Sweden)

    Roja Rani Pallavali

    Full Text Available Multi-drug resistance has become a major problem for the treatment of pathogenic bacterial infections. The use of bacteriophages is an attractive approach to overcome the problem of drug resistance in several pathogens that cause fatal diseases. Our study aimed to isolate multi drug resistant bacteria from patients with septic wounds and then isolate and apply bacteriophages in vitro as alternative therapeutic agents. Pus samples were aseptically collected from Rajiv Gandhi Institute of Medical Science (RIMS, Kadapa, A.P., and samples were analyzed by gram staining, evaluating morphological characteristics, and biochemical methods. MDR-bacterial strains were collected using the Kirby-Bauer disk diffusion method against a variety of antibiotics. Bacteriophages were collected and tested in vitro for lytic activity against MDR-bacterial isolates. Analysis of the pus swab samples revealed that the most of the isolates detected had Pseudomonas aeruginosa as the predominant bacterium, followed by Staphylococcus aureus, Klebsiella pneumoniae and Escherichia coli. Our results suggested that gram-negative bacteria were more predominant than gram-positive bacteria in septic wounds; most of these isolates were resistant to ampicillin, amoxicillin, penicillin, vancomycin and tetracycline. All the gram-positive isolates (100% were multi-drug resistant, whereas 86% of the gram-negative isolates had a drug resistant nature. Further bacteriophages isolated from sewage demonstrated perfect lytic activity against the multi-drug resistant bacteria causing septic wounds. In vitro analysis of the isolated bacteriophages demonstrated perfect lysis against the corresponding MDR-bacteria, and these isolated phages may be promising as a first choice for prophylaxis against wound sepsis, Moreover, phage therapy does not enhance multi-drug resistance in bacteria and could work simultaneously on a wide variety of MDR-bacteria when used in a bacteriophage cocktail. Hence

  1. An Intramolecular Chaperone Inserted in Bacteriophage P22 Coat Protein Mediates Its Chaperonin-independent Folding*

    Science.gov (United States)

    Suhanovsky, Margaret M.; Teschke, Carolyn M.

    2013-01-01

    The bacteriophage P22 coat protein has the common HK97-like fold but with a genetically inserted domain (I-domain). The role of the I-domain, positioned at the outermost surface of the capsid, is unknown. We hypothesize that the I-domain may act as an intramolecular chaperone because the coat protein folds independently, and many folding mutants are localized to the I-domain. The function of the I-domain was investigated by generating the coat protein core without its I-domain and the isolated I-domain. The core coat protein shows a pronounced folding defect. The isolated I-domain folds autonomously and has a high thermodynamic stability and fast folding kinetics in the presence of a peptidyl prolyl isomerase. Thus, the I-domain provides thermodynamic stability to the full-length coat protein so that it can fold reasonably efficiently while still allowing the HK97-like core to retain the flexibility required for conformational switching during procapsid assembly and maturation. PMID:24126914

  2. Molecular and chemical engineering of bacteriophages for potential medical applications.

    Science.gov (United States)

    Hodyra, Katarzyna; Dąbrowska, Krystyna

    2015-04-01

    Recent progress in molecular engineering has contributed to the great progress of medicine. However, there are still difficult problems constituting a challenge for molecular biology and biotechnology, e.g. new generation of anticancer agents, alternative biosensors or vaccines. As a biotechnological tool, bacteriophages (phages) offer a promising alternative to traditional approaches. They can be applied as anticancer agents, novel platforms in vaccine design, or as target carriers in drug discovery. Phages also offer solutions for modern cell imaging, biosensor construction or food pathogen detection. Here we present a review of bacteriophage research as a dynamically developing field with promising prospects for further development of medicine and biotechnology.

  3. Bacteriophage Mediated Killing of Staphylococcus aureus In Vitro on Orthopaedic K Wires in Presence of Linezolid Prevents Implant Colonization

    Science.gov (United States)

    Kaur, Sandeep; Harjai, Kusum; Chhibber, Sanjay

    2014-01-01

    Background Infections of bone and joint tissues following arthroplasty surgeries remain a major challenge in orthopaedic settings. Methicillin resistant Staphylococcus aureus (MRSA) is recognised as an established pathogen in such infections. Combination therapy using linezolid and bacteriophage impregnated in biopolymer was investigated in the present study as an alternative strategy to prevent MRSA colonisation on the orthopaedic implant surface. Methodology Coating of stainless steel orthopaedic grade K-wires was achieved using hydroxypropylmethlycellulose (HPMC) mixed with phage alone, linezolid alone and phage and linezolid together. The potential of these agents to inhibit adhesion of S.aureus (MRSA) 43300 on K-wires was assessed. Coated and naked wires were analysed by scanning electron microscopy (SEM) and fluorescent staining. Result Significant reduction in bacterial adhesion was achieved on phage/linezolid wires in comparison to naked as well as HPMC coated wires. However, maximum reduction in bacterial adherence (∼4 log cycles) was observed on the wires coated with phage-linezolid combination. The frequency of emergence of resistant mutants was also negligible in presence of both the agents. Conclusion This study provides evidence to confirm that local delivery system employing linezolid (a potent protein synthesis inhibitor) along with a broad spectrum lytic bacteriophage (capable of self-multiplication) is able to attack the adhered as well as surrounding bacteria present near the implant site. Unlike other antibiotic based therapies, this combination has the potential to significantly restrict the emergence of resistant mutants, thus paving the way for effective treatment of MRSA associated infection of medical implants. PMID:24594764

  4. Molecular studies on bacteriophage endolysins and their potential to control gram-negative bacteria

    OpenAIRE

    Oliveira, Hugo Alexandre Mendes

    2014-01-01

    Thesis for PhD degree in Chemical and Biological Engineeering Bacteriophages are viruses that specifically infect bacterial hosts to reproduce. At the end of the infection cycle, progeny virions are confronted with a rigid cell wall that impedes their release into the environment. Consequently, bacteriophages encode hydrolytic enzymes, called endolysins, to digest the peptidoglycan and cause bacteriolysis. In contrast to their extensively studied counterparts, active against Gram-positi...

  5. STUDIES ON THE BACTERIOPHAGE OF D'HÉRELLE

    Science.gov (United States)

    Hetler, D. M.; Bronfenbrenner, J.

    1928-01-01

    1. During the process of lysis by bacteriophage, there is an appreciable increase in the amount of free amino acid present in the culture. 2. The increase of free amino acid is due to hydrolysis of bacterial protein. PMID:19869482

  6. Bacteriophages: the possible solution to treat infections caused by pathogenic bacteria.

    Science.gov (United States)

    El-Shibiny, Ayman; El-Sahhar, Salma

    2017-11-01

    Since their discovery in 1915, bacteriophages have been used to treat bacterial infections in animals and humans because of their unique ability to infect their specific bacterial hosts without affecting other bacterial populations. The research carried out in this field throughout the 20th century, largely in Georgia, part of USSR and Poland, led to the establishment of phage therapy protocols. However, the discovery of penicillin and sulfonamide antibiotics in the Western World during the 1930s was a setback in the advancement of phage therapy. The misuse of antibiotics has reduced their efficacy in controlling pathogens and has led to an increase in the number of antibiotic-resistant bacteria. As an alternative to antibiotics, bacteriophages have become a topic of interest with the emergence of multidrug-resistant bacteria, which are a threat to public health. Recent studies have indicated that bacteriophages can be used indirectly to detect pathogenic bacteria or directly as biocontrol agents. Moreover, they can be used to develop new molecules for clinical applications, vaccine production, drug design, and in the nanomedicine field via phage display.

  7. Research of pathogenic bacteria and bacteriophages in the residuals of wastewater treatment plants

    International Nuclear Information System (INIS)

    Mathlouthi, Soumaya

    2011-01-01

    The aim of this study is to find the pathogenic bacteria Listeria and Salmonella and to detect of bacterial (fecal coliforms) and viral indicators (bacteriophage) of fecal contamination in the residues of three sewage treatment plants in Greater Tunis: Charguia, Jdaida and Wardia. Three types of samples were analyzed: raw sewage, treated wastewater and sludge. The study showed the presence of pathogenic bacteria in some samples with a frequency of 7 pour cent for Listeria and 21 pour cent for Salmonella. However, none of these organisms has been detected in treated water of Jdaida and Chargia reflecting the efficiency of the purification process in these stations. Furthermore, all samples were positive for the presence of fecal coliforms and bacteriophages with important titles: up to 8.23 log10 (CFU/L) for coliforms and 8.36 log10 (pfu/L) for bacteriophages.

  8. A sperm-agglutinating lectin from seeds of Jack fruit (Artocarpus heterophyllus).

    Science.gov (United States)

    Namjuntra, P; Muanwongyathi, P; Chulavatnatol, M

    1985-04-30

    A lectin specific for N-acetylgalactosamine was isolated from seed extract of Jack fruit (Artocarpus heterophyllus) by ammonium sulfate precipitation, followed by affinity chromatography on a Affigel-galactosamine-agarose column. The lectin possessed agglutinating activities for human and rat sperm as well as human red blood cells. It was found to have Mr = 62,000 consisting of two dissimilar subunits of Mr = 18,000 and 13,000. It also cross-reacted with an antibody against the lectin of Osage Orange (Maclura pomifera).

  9. In vitro characterization and in vivo properties of Salmonellae lytic bacteriophages isolated from free-range layers

    Directory of Open Access Journals (Sweden)

    L Fiorentin

    2004-06-01

    Full Text Available Occurrence of food poisoning related to Salmonella-contaminated eggs and chicken meat has been frequent in humans. Salmonella Enteritidis (SE and Salmonella Typhimurium (ST are included among the most important paratyphoid salmonellae associated with chicken meat and eggs. Elimination of Salmonella at the pre-harvest stage can play a significant role in preventing the introduction of this pathogen into the food chain and consequently in the reduction of food poisoning in humans. Bactericidal bacteriophages may provide a natural, nontoxic, feasible and non-expensive component of the multi-factorial approach for a pre-harvest control of Salmonella in poultry. Five bacteriophages lytic for SE PT4 and ST were obtained from 107 samples of feces of free-range layers in Brazil. All bacteriophages were characterized in vitro and in vivo, showing head and tail morphology and dsDNA as nucleic acids. Results of "in vivo" studies suggested that bacteriophages do not remain in Salmonella-free birds longer than one day, whereas they multiply in Salmonella-infected birds for longer periods. Besides, selection for phage-resistant SE PT4 did not seem to occur in the short term. Isolated bacteriophages will be investigated for their potential for pre-harvest biocontrol of SE PT4 in poultry.

  10. Determination of immune status in dogs against CPV-2 by recombinant protein based latex agglutination test.

    Science.gov (United States)

    Thomas, Jobin; Singh, Mithilesh; Goswami, T K; Glora, Philma; Chakravarti, Soumendu; Chander, Vishal; Upmanyu, Vikramaditya; Verma, Suman; Sharma, Chhavi; Mahendran, K

    2017-09-01

    Canine parvoviral enteritis is a highly contagious viral illness caused by canine parvovirus-2 (CPV-2) which affects puppies of mainly 6-20 weeks of age. Vaccination is pivotal in preventing and controlling CPV-2 infection. Determination of antibody status is a critical determinant for successful vaccination. The hemagglutination inhibition (HI) test is 'gold standard' test for quantification of antibodies specific to CPV-2, although the execution of this test is not feasible under field conditions. The present study was undertaken to develop a point of care testing to determine immune status prior to CPV-2 vaccination or to detect seroconversion in immunized dogs by latex agglutination test (LAT) using recombinant antigen. Truncated portion of VP2 protein (tVP2) of CPV-2 was selected on the basis of antigenic indices, overexpressed the recombinant protein in E. coli system and was subsequently used in development of LAT. A total of 59 serum samples obtained from vaccinated (n = 54) and healthy unvaccinated (n = 5) dogs were tested. The positivity was observed in 85% (46/54) of these dogs with varying agglutination pattern. The overall sensitivity and specificity of latex agglutination test in comparison to HI test was recorded as 90% and 88% respectively with an agreement value of 90% (CI = 95%). Copyright © 2017 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

  11. Effect of HZE particles and space hadrons on bacteriophages

    International Nuclear Information System (INIS)

    Iurov, S.S.; Akoev, I.G.; Leonteva, G.A.

    1983-01-01

    The effects of particle radiation of the type encountered in space flight on bacteriophages are investigated. Survival and mutagenesis were followed in dry film cultures or liquid suspensions of T4Br(+) bacteriophage exposed to high-energy (HZE) particles during orbital flight, to alpha particles and accelerator-generated hardrons in the laboratory, and to high-energy cosmic rays at mountain altitudes. The HZE particles and high-energy hadrons are found to have a greater relative biological efficiency than standard gamma radiation, while exhibiting a highly inhomogeneous spatial structure in the observed biological and genetic effects. In addition, the genetic lesions observed are specific to the type of radiation exposure, consisting primarily of deletions and multiple lesions of low revertability, with mode of action depending on the linear energy transfer. 18 references

  12. Bacteriophages to combat foodborne infections caused by food contamination by bacteria of the Campylobacter genus

    Directory of Open Access Journals (Sweden)

    Magdalena Myga-Nowak

    2016-09-01

    Full Text Available It is estimated that each year more than 2 million people suffer from diarrheal diseases, resulting from the consumption of contaminated meat. Foodborne infections are most frequently caused by small Gram-negative rods Campylobacter. The hosts of these bacteria are mainly birds wherein they are part of the normal intestinal flora. During the commercial slaughter, there is a likelihood of contamination of carcasses by the bacteria found in the intestinal content. In Europe, up to 90% of poultry flocks can be a reservoir of the pathogen. According to the European Food Safety Authority report from 2015, the number of reported and confirmed cases of human campylobacteriosis exceeds 200 thousands per year, and such trend remains at constant level for several years. The occurrence of growing antibiotic resistance in bacteria forces the limitation of antibiotic use in the animal production. Therefore, the European Union allows only using stringent preventive and hygienic treatment on farms. Achieving Campylobacter free chickens using these methods is possible, but difficult to implement and expensive. Utilization of bacterial viruses – bacteriophages, can be a path to provide the hygienic conditions of poultry production and food processing. Formulations applied in the food protection should contain strictly lytic bacteriophages, be non-pyrogenic and retain long lasting biological activity. Currently, on the market there are available commercial bacteriophage preparations for agricultural use, but neither includes phages against Campylobacter. However, papers on the application of bacteriophages against Campylobacter in chickens and poultry products were published in the last few years. In accordance with the estimates, 2-logarithm reduction of Campylobacter in poultry carcases will contribute to the 30-fold reduction in the incidence of campylobacteriosis in humans. Research on bacteriophages against Campylobacter have cognitive and economic

  13. Bacteriophage-resistant mutants in Yersinia pestis: identification of phage receptors and attenuation for mice.

    Directory of Open Access Journals (Sweden)

    Andrey A Filippov

    Full Text Available BACKGROUND: Bacteriophages specific for Yersinia pestis are routinely used for plague diagnostics and could be an alternative to antibiotics in case of drug-resistant plague. A major concern of bacteriophage therapy is the emergence of phage-resistant mutants. The use of phage cocktails can overcome this problem but only if the phages exploit different receptors. Some phage-resistant mutants lose virulence and therefore should not complicate bacteriophage therapy. METHODOLOGY/PRINCIPAL FINDINGS: The purpose of this work was to identify Y. pestis phage receptors using site-directed mutagenesis and trans-complementation and to determine potential attenuation of phage-resistant mutants for mice. Six receptors for eight phages were found in different parts of the lipopolysaccharide (LPS inner and outer core. The receptor for R phage was localized beyond the LPS core. Most spontaneous and defined phage-resistant mutants of Y. pestis were attenuated, showing increase in LD₅₀ and time to death. The loss of different LPS core biosynthesis enzymes resulted in the reduction of Y. pestis virulence and there was a correlation between the degree of core truncation and the impact on virulence. The yrbH and waaA mutants completely lost their virulence. CONCLUSIONS/SIGNIFICANCE: We identified Y. pestis receptors for eight bacteriophages. Nine phages together use at least seven different Y. pestis receptors that makes some of them promising for formulation of plague therapeutic cocktails. Most phage-resistant Y. pestis mutants become attenuated and thus should not pose a serious problem for bacteriophage therapy of plague. LPS is a critical virulence factor of Y. pestis.

  14. THE INVESTIGATION OF BRUCELLA ANTIBODY WITH MILK RING TEST AND AGGLUTINATION TEST IN MILK COLLECTED FROM SAMSUN REGION

    Directory of Open Access Journals (Sweden)

    Goknur TERZI

    2006-06-01

    Full Text Available In this study Brucella antibodies were investigated with agglutination test (Whey-AT and Milk Ring Test (MRT in a total of 100 milk samples as 50 of cow milk and 50 of goat milk collected from center and villages of Samsun. According to MRT Brucella antibodies was positive at 10 samples (20 % of cow milk and 6 samples (12 % of goat milk. In cow milk, 4 (8 % positive, 3 (6 % suspicious and 43 (86 % negative samples; in goat milk 3 (6 % positive, 2 (4 % suspicious and 45 (90 % negative samples were determined according to antibodies titre of serum agglutination test (Whey-AT. [TAF Prev Med Bull 2006; 5(3.000: 196-203

  15. Contractile injection systems of bacteriophages and related systems

    DEFF Research Database (Denmark)

    Taylor, Nicholas M I; van Raaij, Mark J; Leiman, Petr G

    2018-01-01

    Contractile tail bacteriophages, or myobacteriophages, use a sophisticated biomolecular structure to inject their genome into the bacterial host cell. This structure consists of a contractile sheath enveloping a rigid tube that is sharpened by a spike-shaped protein complex at its tip. The spike ...

  16. Bacteriophage Infectivity Against Pseudomonas aeruginosa in Saline Conditions

    KAUST Repository

    Scarascia, Giantommaso; Yap, Scott A.; Kaksonen, Anna H.; Hong, Pei-Ying

    2018-01-01

    at different temperature, pH, and salinity. Bacteriophages showed optimal infectivity at a multiplicity of infection of 10 in saline conditions, and demonstrated lytic abilities over all tested temperature (25, 30, 37, and 45°C) and pH 6–9. Planktonic P

  17. Ability of Bacillus subtilis protoplasts to repair irradiated bacteriophage deoxyribonucleic acid via acquired and natural enzymatic systems

    International Nuclear Information System (INIS)

    Yasbin, R.E.; Andersen, B.J.; Sutherland, B.M.

    1981-01-01

    A novel form of enzyme therapy was achieved by utilizing protoplasts of Bacillus subtilis. Photoreactivating enzyme of Escherichia coli was successfully inserted into the protoplasts of B. subtilis treated with polyethylene glycol. This enzyme was used to photoreactivate ultraviolet-damaged bacteriophage deoxyribonucleic acid (DNA). Furthermore, in polyethylene glycol-treated protoplasts, ultraviolet-irradiated transfecting bacteriophage DNA was shown to be a functional substrate for the host DNA excision repair system. Previous results (R.E. Yasbin, J.D. Fernwalt, and P.I. Fields, J. Bacteriol.; 137: 391-396) showed that ultraviolet-irradiated bacteriophage DNA could not be repaired via the excision repair system of competent cells. Therefore, the processing of bacteriophage DNA by protoplasts and by competent cells must be different. This sensitive protoplast assay can be used to identify and to isolate various types of DNA repair enzymes

  18. RGD peptide-displaying M13 bacteriophage/PLGA nanofibers as cell-adhesive matrices for smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Yong Cheol; Lee, Jong Ho; Jin, Oh Seong; Lee, Eun Ji; Jin, Lin Hua; Kim, Chang Seok; Hong, Suck Won; Han, Dong Wook; Kim, Chun Tae; Oh, Jin Woo [Pusan National University, Busan (Korea, Republic of)

    2015-01-15

    Extracellular matrices (ECMs) are network structures that play an essential role in regulating cellular growth and differentiation. In this study, novel nanofibrous matrices were fabricated by electrospinning M13 bacteriophage and poly(lactic-co-glycolic acid) (PLGA) and were shown to be structurally and functionally similar to natural ECMs. A genetically-engineered M13 bacteriophage was constructed to display Arg-Gly-Asp (RGD) peptides on its surface. The physicochemical properties of RGD peptide-displaying M13 bacteriophage (RGD-M13 phage)/PLGA nanofibers were characterized by using scanning electron microscopy and Fourier-transform infrared spectroscopy. We used immunofluorescence staining to confirm that M13 bacteriophages were homogenously distributed in RGD-M13 phage/PLGA matrices. Furthermore, RGD-M13 phage/PLGA nanofibrous matrices, having excellent biocompatibility, can enhance the behaviors of vascular smooth muscle cells. This result suggests that RGD-M13 phage/PLGA nanofibrous matrices have potentials to serve as tissue engineering scaffolds.

  19. A deep-sea agglutinated foraminifer tube constructed with planktonic foraminifer shells of a single species

    Science.gov (United States)

    Pearson, Paul N.; Expedition 363 Shipboard Scientific Party, IODP

    2018-01-01

    Agglutinated foraminifera are marine protists that show apparently complex behaviour in constructing their shells, involving selecting suitable sedimentary grains from their environment, manipulating them in three dimensions, and cementing them precisely into position. Here we illustrate a striking and previously undescribed example of complex organisation in fragments of a tube-like foraminifer (questionably assigned to Rhabdammina) from 1466 m water depth on the northwest Australian margin. The tube is constructed from well-cemented siliciclastic grains which form a matrix into which hundreds of planktonic foraminifer shells are regularly spaced in apparently helical bands. These shells are of a single species, Turborotalita clarkei, which has been selected to the exclusion of all other bioclasts. The majority of shells are set horizontally in the matrix with the umbilical side upward. This mode of construction, as is the case with other agglutinated tests, seems to require either an extraordinarily selective trial-and-error process at the site of cementation or an active sensory and decision-making system within the cell.

  20. The entomopathogenic fungus Nomuraea rileyi impairs cellular immunity of its host Helicoverpa armigera.

    Science.gov (United States)

    Zhong, Ke; Liu, Zhan-Chi; Wang, Jia-Lin; Liu, Xu-Sheng

    2017-09-01

    In this study, we investigated the effect of the entomopathogenic fungus Nomuraea rileyi on Helicoverpa armigera cellular immune responses. Nomuraea rileyi infection had no effect on total hemocyte count (THC), but impaired hemocyte-mediated phagocytosis, nodulation, and encapsulation responses. Nomuraea rileyi infection led to a significant reduction in hemocyte spreading. An in vitro assay revealed that plasma from N. rileyi infected H. armigera larvae suppressed the spreading ability of hemocytes from naïve larvae. We infer that N. rileyi suppresses the cellular immune response of its host, possibly by secreting exogenous, cytotoxic compounds into the host's hemolymph. © 2017 Wiley Periodicals, Inc.

  1. In vitro evaluation of a novel bacteriophage cocktail as a preventative for bovine coliform mastitis.

    Science.gov (United States)

    Porter, J; Anderson, J; Carter, L; Donjacour, E; Paros, M

    2016-03-01

    The objective of this study was to investigate the potential use of bacteriophage in preventing Escherichia coli mastitis on dairies. A cocktail consisting of 4 distinct bacteriophages was generated by screening against 36 E. coli isolates from dairy cows in Washington State with clinical mastitis. The bacteriophage significantly inhibited growth of 58% of the Washington State isolates and 54% of E. coli mastitis isolates from New York State, suggesting that the cocktail of phages had a relatively broad spectrum of action against relevant strains from 2 distinct geographies. The ability to suppress bacterial growth of these isolates in a liquid growth medium was not affected by the ratio of bacteriophage particles to bacterial cells (multiplicity of infection, MOI). For those E. coli that were completely inhibited by the phage cocktail, an MOI as low as 10 had the same effect as 10 µg/mL of ceftiofur on the growth rate of E. coli over a 12-h period using optical density measurements. A 3.3- to 5.6-log reduction of growth was achieved when E. coli was co-incubated with our phage cocktail in raw milk over a 12-h period at physiologic temperature. A modified gentamicin protection assay using bovine mammary epithelial cells provided a model to test whether bacteriophage could prevent cell attachment and invasion by chronic coliform mastitis strains. Pretreatment of cell cultures with the phage cocktail significantly reduced adhesion and intracellular survival of E. coli compared with controls. When combined with a bismuth-based teat sealant, the phage cocktail was able to inhibit bacterial growth when challenged with 1.6 × 10(3) cfu/mL of a clinical mastitis E. coli strain. In vitro results show bactericidal activity by our phage in raw milk and mammary tissue culture systems. Before a bacteriophage-based dry-cow treatment becomes a potential option for dairies, in vivo studies must be able to demonstrate that a specific dose of bacteriophage can protect cows from

  2. Microscopic agglutination test on captive rattlesnakes : Data on serovars and titers

    Directory of Open Access Journals (Sweden)

    T.C.S. Rodrigues

    2016-06-01

    Full Text Available The microscopic agglutination test (MAT is considered the “golden standard” leptospirosis serodiagnostic test, but there is little information about it as it pertains to snakes. To fill this information gap, we provide data on serovars and titers of fifty-six Crotalus durissus collilineatus sera samples that tested positive by MAT (10.1016/j.actatropica.2016.02.006 (Rodrigues et al., 2016 [5]. These data are presented in a table, along with a description of the methodology used for sample collection and serologic testing.

  3. Identification of novel bacteriophage peptides using a combination of gene sequence LC-MS-MS analysis and BLASTP

    Science.gov (United States)

    Introduction: In an effort to characterize novel bacteriophage with lytic activity against pathogenic E.coli associated with foodborne illness, gene sequencing and mass spectrometry have been used to identify expressed peptides which differentiate isolated bacteriophage from other known phage. Here,...

  4. Rapid detection of methicillin resistance in Staphylococcus aureus isolates by the MRSA-screen latex agglutination test

    NARCIS (Netherlands)

    W.B. van Leeuwen (Willem); C. van Pelt (Cindy); A. Luijendijk (Ad); H.A. Verbrugh (Henri); W.H.F. Goessens (Wil)

    1999-01-01

    textabstractThe slide agglutination test MRSA-Screen (Denka Seiken Co., Niigata, Japan) was compared with the mecA PCR ("gold standard") for the detection of methicillin resistance in Staphylococcus aureus. The MRSA-Screen test detected the penicillin-binding protein 2a

  5. Comparative analysis of the biological and physical properties of Enterococcus faecalis bacteriophage vB_EfaS_GEC-EfS_3 and Streptococcus mitis bacteriophage vB_SmM_GEC-SmitisM_2.

    Science.gov (United States)

    Rigvava, Sophio; Tchgkonia, Irina; Jgenti, Darejan; Dvalidze, Teona; Carpino, James; Goderdzishvili, Marina

    2013-01-01

    Enterococcus faecalis and Streptococcus mitis are common commensal inhabitants of the human gastrointestinal and genitourinary tracts. However, both species can be opportunistic pathogens and cause disease in nosocomial settings. These infections can be difficult to treat because of the frequency of antibiotic resistance among these strains. Bacteriophages are often suggested as an alternative therapeutic agent against these infections. In this study, E. faecalis and S. mitis strains were isolated from female patients with urinary tract infections. Bacteriophages active against these strains were isolated from sewage water from the Mtkvari River. Two phages, designated vB_EfaS_GEC-EfS_3 (Syphoviridae) and vB_SmM_GEC-SmitisM_2 (Myoviridae), were specific for E. faecalis and S. mitis, respectively. Each phage's growth patterns and adsorption rates were quantified. Sensitivity to ultraviolet light and temperature was determined, as was host range and serology. The S. mitis bacteriophage was found to be more resistant to ultraviolet light and exposure to high temperatures than the E. faecalis bacteriophage, despite having a much greater rate of replication. While each phage was able to infect a broad range of strains of the same species as the host species from which they were isolated, they were unable to infect other host species tested.

  6. [Bacteriophages in the battle against multidrug resistant bacteria

    NARCIS (Netherlands)

    Meer, J.W.M. van der; Vandenbroucke-Grauls, C.

    2018-01-01

    Bacteriophages are viruses that infect bacteria. They are highly specific for a bacterial species. The so-called 'lytic phages' can lyse bacteria when they infect them; these phages can be used to treat bacterial infections. Despite a century of experience with phage therapy, the evidence for

  7. Circulating Hemocytes from Larvae of the Japanese Rhinoceros Beetle Allomyrina dichotoma (Linnaeus) (Coleoptera: Scarabaeidae) and the Cellular Immune Response to Microorganisms.

    Science.gov (United States)

    Hwang, Sejung; Bang, Kyeongrin; Lee, Jiae; Cho, Saeyoull

    2015-01-01

    Hemocytes of the last larva of the Japanese rhinoceros beetle A. dichotoma (Linnaeus) (Coleoptera: Scarabaeidae) were classified as granulocytes, plasmatocytes, oenocytoids, spherulocytes, prohemocytes, and adipohemocytes. Among these cell types, only the granulocytes became immunologically activated with obvious morphological changes, displaying large amoeba-like, lobopodia-like, and fan-like structures. In addition, their cytoplasmic granules became larger and greatly increased in number. To explore whether these granules could be immunologically generated as phagosomes, total hemocytes were stained with LysoTracker. Greater than 90% of the granulocytes retained the LysoTracker dye at 4 h post-bacterial infection. In flow cytometry analysis, the red fluorescent signal was highly increased at 4 h post-bacterial infection (60.36%) compared to controls (5.08%), as was confirmed by fluorescent microscopy. After 12 h post-infection, these signals returned to basal levels. The uptake of pathogens by granulocytes rapidly triggered the translocation of the microtubule-associated protein 1 light chain 3 alpha (LC3) to the phagosome, which may result in enhanced pathogen killing.

  8. Circulating Hemocytes from Larvae of the Japanese Rhinoceros Beetle Allomyrina dichotoma (Linnaeus (Coleoptera: Scarabaeidae and the Cellular Immune Response to Microorganisms.

    Directory of Open Access Journals (Sweden)

    Sejung Hwang

    Full Text Available Hemocytes of the last larva of the Japanese rhinoceros beetle A. dichotoma (Linnaeus (Coleoptera: Scarabaeidae were classified as granulocytes, plasmatocytes, oenocytoids, spherulocytes, prohemocytes, and adipohemocytes. Among these cell types, only the granulocytes became immunologically activated with obvious morphological changes, displaying large amoeba-like, lobopodia-like, and fan-like structures. In addition, their cytoplasmic granules became larger and greatly increased in number. To explore whether these granules could be immunologically generated as phagosomes, total hemocytes were stained with LysoTracker. Greater than 90% of the granulocytes retained the LysoTracker dye at 4 h post-bacterial infection. In flow cytometry analysis, the red fluorescent signal was highly increased at 4 h post-bacterial infection (60.36% compared to controls (5.08%, as was confirmed by fluorescent microscopy. After 12 h post-infection, these signals returned to basal levels. The uptake of pathogens by granulocytes rapidly triggered the translocation of the microtubule-associated protein 1 light chain 3 alpha (LC3 to the phagosome, which may result in enhanced pathogen killing.

  9. Insights into bacteriophage application in controlling Vibrio species

    Directory of Open Access Journals (Sweden)

    Vengadesh Letchumanan

    2016-07-01

    Full Text Available Bacterial infections from various organisms including Vibrio sp. pose a serious hazard to humans in many forms from clinical infection to affecting the yield of agriculture and aquaculture via infection of livestock. Vibrio sp. is one of the main foodborne pathogens causing human infection and is also a common cause of losses in the aquaculture industry. Prophylactic and therapeutic usage of antibiotics has become the mainstay of managing this problem, however this in turn led to the emergence of multidrug resistant strains of bacteria in the environment; which has raised awareness of the critical need for alternative non antibiotic based methods of preventing and treating bacterial infections. Bacteriophages - viruses that infect and result in the death of bacteria – are currently of great interest as a highly viable alternative to antibiotics. This article provides an insight into bacteriophage application in controlling Vibrio species as well underlining the advantages and drawbacks of phage therapy.

  10. Isolation and characterization of specific bacteriophage Va1 to Vibrio alginolyticus

    Directory of Open Access Journals (Sweden)

    Carla Fernández Espinel

    2017-04-01

    Full Text Available Vibrio alginolyticus is associated with diseases in aquaculture. The misuse of antibiotics has led to the search for alternatives in the treatment of bacterial diseases, among them the application of bacteriophages that infect and destroy bacteria selectively. In this way, a highly lytic V. alginolyticus bacteriophage, termed Va1, was isolated, with the aim to evaluate its physical chemical parameters. For this purpose, different temperature, pH, chloroform exposure and host range conditions were evaluated. The temperature stability of phage Va1 showed higher titers at 20 and 30 °C decreasing from 40 °C. With respect to pH, the highest titers for the bacteriophage were between 5 and 8, and chloroform exposure reduced viability of the Va1 phage by 25%. The one-step curve determined that the latency period and the burst size were 20 minutes and 192 PFU / infective center respectively. Under the transmission electron microscope, the Va1 phage showed an icosahedral head and a non-contractile tail, belonging to the Podoviridae family. In conclusion, Va1 phage presents potential characteristics for use in phage therapy.

  11. Evidence of translation efficiency adaptation of the coding regions of the bacteriophage lambda.

    Science.gov (United States)

    Goz, Eli; Mioduser, Oriah; Diament, Alon; Tuller, Tamir

    2017-08-01

    Deciphering the way gene expression regulatory aspects are encoded in viral genomes is a challenging mission with ramifications related to all biomedical disciplines. Here, we aimed to understand how the evolution shapes the bacteriophage lambda genes by performing a high resolution analysis of ribosomal profiling data and gene expression related synonymous/silent information encoded in bacteriophage coding regions.We demonstrated evidence of selection for distinct compositions of synonymous codons in early and late viral genes related to the adaptation of translation efficiency to different bacteriophage developmental stages. Specifically, we showed that evolution of viral coding regions is driven, among others, by selection for codons with higher decoding rates; during the initial/progressive stages of infection the decoding rates in early/late genes were found to be superior to those in late/early genes, respectively. Moreover, we argued that selection for translation efficiency could be partially explained by adaptation to Escherichia coli tRNA pool and the fact that it can change during the bacteriophage life cycle.An analysis of additional aspects related to the expression of viral genes, such as mRNA folding and more complex/longer regulatory signals in the coding regions, is also reported. The reported conclusions are likely to be relevant also to additional viruses. © The Author 2017. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

  12. [Detection of anti-Leptospira antibodies in sera of patients in the latex agglutination test].

    Science.gov (United States)

    Volina, E G; Sarukhanova, L E; Iashina, N V; Prokopov, N I; Shkarlat, P E; Barysheva, I V

    2001-01-01

    The results of the preliminary evaluation of the sensitivity and specificity of the newly developed diagnostic test based on the determination of genus-specific antibodies to leptospires in the latex agglutination test, are presented. This test makes it possible to detect anti-Leptospira antibodies of any serogroup. The advantages of the developed test have been determined.

  13. Lytic Infection of Lactococcus lactis by Bacteriophages Tuc2009 and c2 Triggers Alternative Transcriptional Host Responses

    NARCIS (Netherlands)

    Ainsworth, S.; Zomer, A.L.; Mahony, J.; Sinderen, D. van

    2013-01-01

    Here we present an entire temporal transcriptional profile of Lactococcus lactis subsp. cremoris UC509.9 undergoing lytic infection with two distinct bacteriophages, Tuc2009 and c2. Furthermore, corresponding high-resolution whole-phage genome tiling arrays of both bacteriophages were performed

  14. Bacteriophages encode factors required for protection in a symbiotic mutualism.

    Science.gov (United States)

    Oliver, Kerry M; Degnan, Patrick H; Hunter, Martha S; Moran, Nancy A

    2009-08-21

    Bacteriophages are known to carry key virulence factors for pathogenic bacteria, but their roles in symbiotic bacteria are less well understood. The heritable symbiont Hamiltonella defensa protects the aphid Acyrthosiphon pisum from attack by the parasitoid Aphidius ervi by killing developing wasp larvae. In a controlled genetic background, we show that a toxin-encoding bacteriophage is required to produce the protective phenotype. Phage loss occurs repeatedly in laboratory-held H. defensa-infected aphid clonal lines, resulting in increased susceptibility to parasitism in each instance. Our results show that these mobile genetic elements can endow a bacterial symbiont with benefits that extend to the animal host. Thus, phages vector ecologically important traits, such as defense against parasitoids, within and among symbiont and animal host lineages.

  15. Cholera dynamics with Bacteriophage infection: A mathematical study

    International Nuclear Information System (INIS)

    Misra, A.K.; Gupta, Alok; Venturino, Ezio

    2016-01-01

    Highlights: • A mathematical model for the biological control of cholera has been proposed. • The feasibility and stability of all the equilibria have been investigated. • The ODE model is found to exhibit Hopf-bifurcation. • Conditions of global asymptotic stability have been obtained. • The impact of important parameters on cholera spread has been shown. - Abstract: Mathematical modeling of waterborne diseases, such as cholera, including a biological control using Bacteriophage viruses in the aquatic reservoirs is of great relevance in epidemiology. In this paper, our aim is twofold: at first, to understand the cholera dynamics in the region around a water body; secondly, to understand how the spread of Bacteriophage infection in the cholera bacterium V. cholerae controls the disease in the human population. For this purpose, we modify the model proposed by Codeço, for the spread of cholera infection in human population and the one proposed by Beretta and Kuang, for the spread of Bacteriophage infection in the bacteria population [1, 2]. We first discuss the feasibility and local asymptotic stability of all the possible equilibria of the proposed model. Further, in the numerical investigation, we have found that the parameter ϕ, called the phage adsorption rate, plays an important role. There is a critical value, ϕ c , at which the model possess Hopf-bifurcation. For lower values than ϕ c , the equilibrium E * is unstable and periodic solutions are observed, while above ϕ c , the equilibrium E * is locally asymptotically stable, and further shown to be also globally asymptotically stable. We investigate the effect of the various parameters on the dynamics of the infected humans by means of numerical simulations.

  16. Analysis of the complete DNA sequence of the temperate bacteriophage TP901-1: Evolution, structure, and genome organization of lactococcal bacteriophages

    DEFF Research Database (Denmark)

    Brøndsted, Lone; Østergaard, Solvej; Pedersen, Margit

    2001-01-01

    A complete analysis of the entire genome of the temperate lactococcal bacteriophage TP901-1 has been performed and the function of 21 of 56 TP901-1-encoded ORFs has been assigned. This knowledge has been used to propose 10 functional modules each responsible for specific functions during...

  17. Elucidating the pH-Dependent Structural Transition of T7 Bacteriophage Endolysin.

    Science.gov (United States)

    Sharma, Meenakshi; Kumar, Dinesh; Poluri, Krishna Mohan

    2016-08-23

    Bacteriophages are the most abundant and diverse biological entities on earth. Bacteriophage endolysins are unique peptidoglycan hydrolases and have huge potential as effective enzybiotics in various infectious models. T7 bacteriophage endolysin (T7L), also known as N-acetylmuramoyl-l-alanine amidase or T7 lysozyme, is a 17 kDa protein that lyses a range of Gram-negative bacteria by hydrolyzing the amide bond between N-acetylmuramoyl residues and the l-alanine of the peptidoglycan layer. Although the activity profiles of several of the T7 family members have been known for many years, the molecular basis for their pH-dependent differential activity is not clear. In this study, we explored the pH-induced structural, stability, and activity characteristics of T7L by applying a variety of biophysical techniques and protein nuclear magnetic resonance (NMR) spectroscopy. Our studies established a reversible structural transition of T7L below pH 6 and the formation of a partially denatured conformation at pH 3. This low-pH conformation is thermally stable and exposed its hydrophobic pockets. Further, NMR relaxation measurements and structural analysis unraveled that T7L is highly dynamic in its native state and a network of His residues are responsible for the observed pH-dependent conformational dynamics and transitions. As bacteriophage chimeric and engineered endolysins are being developed as novel therapeutics against multiple drug resistance pathogens, we believe that our results are of great help in designing these entities as broadband antimicrobial and/or antibacterial agents.

  18. Identification of expressed genes in cDNA library of hemocytes from the RLO-challenged oyster, Crassostrea ariakensis Gould with special functional implication of three complement-related fragments (CaC1q1, CaC1q2 and CaC3).

    Science.gov (United States)

    Xu, Ting; Xie, Jiasong; Li, Jianming; Luo, Ming; Ye, Shigen; Wu, Xinzhong

    2012-06-01

    A SMARTer™ cDNA library of hemocyte from Rickettsia-like organism (RLO) challenged oyster, Crassostrea ariakensis Gould was constructed. Random clones (400) were selected and single-pass sequenced, resulted in 200 unique sequences containing 96 known genes and 104 unknown genes. The 96 known genes were categorized into 11 groups based on their biological process. Furthermore, we identified and characterized three complement-related fragments (CaC1q1, CaC1q2 and CaC3). Tissue distribution analysis revealed that all of three fragments were ubiquitously expressed in all tissues studied including hemocyte, gills, mantle, digestive glands, gonads and adductor muscle, while the highest level was seen in the hemocyte. Temporal expression profile in the hemocyte monolayers reveled that the mRNA expression levels of three fragments presented huge increase after the RLO incubation at 3 h and 6 h in post-challenge, respectively. And the maximal expression levels at 3 h in post-challenge are about 256, 104 and 64 times higher than the values detected in the control of CaC1q1, CaC1q2 and CaC3, respectively. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Diagnostic value of latex agglutination test in diagnosis of acute bacterial meningitis

    Directory of Open Access Journals (Sweden)

    Syeda Fasiha Mohammadi

    2013-01-01

    Full Text Available Objectives: To know the incidence of bacterial meningitis in children below five years of age. To compare conventional culture and antigen detection methods ( Latex agglutination test. Materials and Methods: 100 CSF samples of clinically suspected meningitis cases in children below 5 years of age were included. The samples were subjected to cell count, Gram stain, culture and LAT. The organisms isolated in the study were characterized according to standard procedures. Results: Of the 100 cases studied, 31 cases were diagnosed as ABM by Gram stain, culture and latex agglutination test as per WHO criteria. The hospital frequency of ABM was 1.7%. 15 (48.38 cases were culture positive. Gram stain was positive in 22(70.96 cases and LAT in 17(54.83 cases. Haemophilus influenzae was the most common causative agent of acute bacterial meningitis followed by S.pneumoniae. Case fatality rate was 45.16%.The sensitivity and specificity of LAT was 66.66% and 87.91% respectively. Conclusion : Bacterial meningitis is a medical emergency and early diagnosis and treatment is life saving and reduces chronic morbidity. LAT was more sensitive compared to conventional Gram stain and Culture technique in identifying the fastidious organisms like H.influenzae, S.pneumoniae and Group B Streptococcus. However, the combination of Gram stain, Culture and LAT proved to be more productive than any of the single tests alone.

  20. Características físicas de dietas para peixes confeccionadas com diferentes aglutinantes Influence of agglutinants on physical stability of fish diets

    Directory of Open Access Journals (Sweden)

    Margarida Maria Barros

    2002-04-01

    Full Text Available Este experimento teve por objetivo avaliar a eficiência de diferentes aglutinantes, a seco e na água, por meio da estabilidade física do pélete. Foram avaliadas duas técnicas de processamento (com ou sem vapor e seis aglutinantes (carboximetilcelulose, polimetilcarbamida, amido de mandioca, alginato de sódio, polivinilpirrolidona, goma guar, através da análise de variância no esquema fatorial (2x6, além de um controle, ao qual não foi acrescido aglutinante. Concluiu-se que o aglutinante melhora significativamente a resistência física do grânulo, independente da técnica de processamento; que o vapor determina grânulos mais estáveis quando em contato com a água e, que o alginato de sódio proporciona grânulos fisicamente mais estáveis, em ambas as técnicas de processamento, enquanto a goma guar a pior.The aim of this paper was to determine the influence of different dry and water agglutinants, through physical stability of pellets. The agglutinants were sodium alginate, guar gum, polymetylcarbamide, polyvinylpirrolidone, carboxymetilcellulose, and cassava starch. The manufacturing processes were two: with and without steam and extrusion. These treatments were evaluated through the variance analysis technique with the factorial scheme 2 x 6 (two processes and six agglutinants, and one control where no extra agglutinants was added. Results showed that, independently of processing technique, the presence the agglutinants improves the physical resistance of the pellets significantly, giving the whole pellets more stability while in contact with water, and that the sodium alginate gives pellets the highest aggregated characteristic, in both processes, while that guar gum gives the lowest.

  1. The effects of ocean acidification on hemocyte of crab species in Alaska from laboratory experiment studies from 2011-07-01 to 2013-07-06 (NODC Accession 0123400)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This archival package contains laboratory experiment data that were collected to examine the effects of ocean acidification on hemocyte of crab species in Alaska....

  2. Bacteriophages: update on application as models for viruses in water

    African Journals Online (AJOL)

    Bacteriophages: update on application as models for viruses in water. ... the resistance of human viruses to water treatment and disinfection processes. ... highly sensitive molecular techniques viruses have been detected in drinking water ...

  3. Molecular cloning and characterization of autophagy-related gene TmATG8 in Listeria-invaded hemocytes of Tenebrio molitor.

    Science.gov (United States)

    Tindwa, Hamisi; Jo, Yong Hun; Patnaik, Bharat Bhusan; Lee, Yong Seok; Kang, Sang Sun; Han, Yeon Soo

    2015-07-01

    Macroautophagy (hereinafter called autophagy) is a highly regulated process used by eukaryotic cells to digest portions of the cytoplasm that remodels and recycles nutrients and disposes of unwanted cytoplasmic constituents. Currently 36 autophagy-related genes (ATG) and their homologs have been characterized in yeast and higher eukaryotes, including insects. In the present study, we identified and functionally characterized the immune function of an ATG8 homolog in a coleopteran insect, Tenebrio molitor (TmATG8). The cDNA of TmATG8 comprises of an ORF of 363 bp that encodes a protein of 120 amino acid residues. TmATG8 transcripts are detected in all the developmental stages analyzed. TmAtg8 protein contains a highly conserved C-terminal glycine residue (Gly116) and shows high amino acid sequence identity (98%) to its Tribolium castaneum homolog, TcAtg8. Loss of function of TmATG8 by RNAi led to a significant increase in the mortality rates of T. molitor larvae against Listeria monocytogenes. Unlike dsEGFP-treated control larvae, TmATG8-silenced larvae failed to turn-on autophagy in hemocytes after injection with L. monocytogenes. These data suggest that TmATG8 play a role in mediating autophagy-based clearance of Listeria in T. molitor. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Bacteriophages and Their Role in Food Safety

    Directory of Open Access Journals (Sweden)

    Sanna M. Sillankorva

    2012-01-01

    Full Text Available The interest for natural antimicrobial compounds has increased due to alterations in consumer positions towards the use of chemical preservatives in foodstuff and food processing surfaces. Bacteriophages fit in the class of natural antimicrobial and their effectiveness in controlling bacterial pathogens in agro-food industry has led to the development of different phage products already approved by USFDA and USDA. The majority of these products are to be used in farm animals or animal products such as carcasses, meats and also in agricultural and horticultural products. Treatment with specific phages in the food industry can prevent the decay of products and the spread of bacterial diseases and ultimately promote safe environments in animal and plant food production, processing, and handling. This is an overview of recent work carried out with phages as tools to promote food safety, starting with a general introduction describing the prevalence of foodborne pathogens and bacteriophages and a more detailed discussion on the use of phage therapy to prevent and treat experimentally induced infections of animals against the most common foodborne pathogens, the use of phages as biocontrol agents in foods, and also their use as biosanitizers of food contact surfaces.

  5. Decreased survival of the λ15 bacteriophage induced by UV-365 nanometers in Escherichia coli

    International Nuclear Information System (INIS)

    Luca, M.E.M. de.

    1989-01-01

    The results of our investigation showed a new effect (not yet described in the current literature) of the UV-365 nm, verified when the bacteria E. coli was irradiated with this wavelenght and then infected with bacteriophage irradiated with short UV (254 nm). In these conditions we observed a decrease in the phage survival. This phenomenon was called Decreased Survival of the Bacteriophage (DSB). We were able to show that DSB was only induced in bacteria irradiated with UV-365 nm, proficient in recombination repair and owning 4-thiouridine in their tRNA. For the induction of DSB it is necessary to promote damage in the bacteriophage through UVA and UVB. It seems that DSB and SOS are antagonistic since DSB is able to suppress the mutation induced by SOS. (author)

  6. 21 CFR 172.785 - Listeria-specific bacteriophage preparation.

    Science.gov (United States)

    2010-04-01

    ... application to meat and poultry products that comply with the ready-to-eat definition in 9 CFR 430.1. Current... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Listeria-specific bacteriophage preparation. 172.785 Section 172.785 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN...

  7. Bacteriophages as Weapons Against Bacterial Biofilms in the Food Industry.

    Science.gov (United States)

    Gutiérrez, Diana; Rodríguez-Rubio, Lorena; Martínez, Beatriz; Rodríguez, Ana; García, Pilar

    2016-01-01

    Microbiological contamination in the food industry is often attributed to the presence of biofilms in processing plants. Bacterial biofilms are complex communities of bacteria attached to a surface and surrounded by an extracellular polymeric material. Their extreme resistance to cleaning and disinfecting processes is related to a unique organization, which implies a differential bacterial growth and gene expression inside the biofilm. The impact of biofilms on health, and the economic consequences, has promoted the development of different approaches to control or remove biofilm formation. Recently, successful results in phage therapy have boosted new research in bacteriophages and phage lytic proteins for biofilm eradication. In this regard, this review examines the environmental factors that determine biofilm development in food-processing equipment. In addition, future perspectives for the use of bacteriophage-derived tools as disinfectants are discussed.

  8. Recombinant Antibodies for the Detection of Bacteriophage MS2 and Ovalbumin

    National Research Council Canada - National Science Library

    O'Connell, Kevin

    2002-01-01

    ...) genes are expressed on the surface of bacteriophage (bacterial virus) particles. We describe here the isolation of additional recombinant antibodies that bind two simulants of biothreat agents...

  9. Lab-on-a-disc agglutination assay for protein detection by optomagnetic readout and optical imaging using nano- and micro-sized magnetic beads

    DEFF Research Database (Denmark)

    Uddin, Rokon; Burger, Robert; Donolato, Marco

    2016-01-01

    of manual steps involved. The detection of the target protein was achieved in two ways: (1) optomagnetic readout using magnetic nanobeads (MNBs); (2) optical imaging using magnetic microbeads (MMBs). The optomagnetic readout of agglutination is based on optical measurement of the dynamics of MNB aggregates...... whereas the imaging method is based on direct visualization and quantification of the average size of MMB aggregates. By enhancing magnetic particle agglutination via application of strong magnetic field pulses, we obtained identical limits of detection of 25 pM with the same sample-to-answer time (15 min...

  10. Leishmaniasis Direct Agglutination Test: Using Pictorials as Training Materials to Reduce Inter-Reader Variability and Improve Accuracy

    NARCIS (Netherlands)

    Adams, Emily R.; Jacquet, Diane; Schoone, Gerard; Gidwani, Kamlesh; Boelaert, Marleen; Cunningham, Jane

    2012-01-01

    Background: The Direct Agglutination Test (DAT) has a high diagnostic accuracy and remains, in some geographical areas, part of the diagnostic algorithm for Visceral Leishmaniasis (VL). However, subjective interpretation of results introduces potential for inter-reader variation. We report an

  11. Laboratory evaluation of a simple and rapid latex agglutination assay for the serodiagnosis of typhoid fever

    NARCIS (Netherlands)

    Abdoel, Theresia H.; Pastoor, Rob; Smits, Henk L.; Hatta, Mochammad

    2007-01-01

    A latex agglutination assay for the serodiagnosis of typhoid fever was evaluated on samples collected from patients with clinical suspicion of typhoid fever in South Sulawesi, Indonesia, where the disease is endemic. The latex assay is very easy to use, gives a rapid result and may be used as a

  12. Interactions of cationic polystyrene nanoparticles with marine bivalve hemocytes in a physiological environment: Role of soluble hemolymph proteins.

    Science.gov (United States)

    Canesi, Laura; Ciacci, Caterina; Fabbri, Rita; Balbi, Teresa; Salis, Annalisa; Damonte, Gianluca; Cortese, Katia; Caratto, Valentina; Monopoli, Marco P; Dawson, Kenneth; Bergami, Elisa; Corsi, Ilaria

    2016-10-01

    The bivalve Mytilus galloprovincialis has proven as a suitable model invertebrate for evaluating the potential impact of nanoparticles (NPs) in the marine environment. In particular, in mussels, the immune system represents a sensitive target for different types of NPs. In environmental conditions, both NP intrinsic properties and those of the receiving medium will affect particle behavior and consequent bioavailability/uptake/toxicity. However, the evaluation of the biological effects of NPs requires additional understanding of how, once within the organism, NPs interact at the molecular level with cells in a physiological environment. In mammalian systems, different NPs associate with serum soluble components, organized into a "protein corona", which affects particle interactions with target cells. However, no information is available so far on the interactions of NPs with biological fluids of aquatic organisms. In this work, the influence of hemolymph serum (HS) on the in vitro effects of amino modified polystyrene NPs (PS-NH2) on Mytilus hemocytes was investigated. Hemocytes were incubated with PS-NH2 suspensions in HS (1, 5 and 50µg/mL) and the results were compared with those obtained in ASW medium. Cell functional parameters (lysosomal membrane stability, oxyradical production, phagocytosis) were evaluated, and morphological changes were investigated by TEM. The activation state of the signalling components involved in Mytilus immune response (p38 MAPK and PKC) was determined. The results show that in the presence of HS, PS-NH2 increased cellular damage and ROS production with respect to ASW medium. The effects were apparently mediated by disregulation of p38 MAPK signalling. The formation of a PS-NH2-protein corona in HS was investigated by centrifugation, and 1D- gel electrophoresis and nano-HPLC-ESI-MS/MS. The results identified the Putative C1q domain containing protein (MgC1q6) as the only component of the PS-NH2 hard protein corona in Mytilus

  13. The direct agglutination test as an alternative method for the diagnosis of canine and human visceral leishmaniasis

    NARCIS (Netherlands)

    Terán-Angel, Guillermo; Schallig, Henk; Zerpa, Olga; Rodríguez, Vestalia; Ulrich, Marian; Cabrera, Maira

    2007-01-01

    Visceral leishmaniasis is the most severe clinical form of leishmaniasis and is often fatal without proper treatment. Therefore, early and accurate diagnosis is important, but often difficult in endemic areas. The aim was to evaluate a direct agglutination test as a potential visceral leishmaniasis

  14. Selective Deactivation of M13 Bacteriophage in E. Coli using Femtosecond Laser Pulses

    CSIR Research Space (South Africa)

    Molukanele, P

    2010-09-01

    Full Text Available Deactivation of M13 Bacteriophage in E. Coli using Femtosecond Laser Pulses P. Molukanele 1, 3, A. Du Plessis 1, T. Roberts 1, L. Botha 1, M. Khati 2,3, W. Campos 2, 3 1CSIR National Laser Centre, Femtosecond Science group, Pretoria, South Africa 2CSIR... that is about 1 ?m long and 5-6 nm in diameter. Its host Escherichia coli (E.coli), is approximately 2-6 ?m long and 1-1.5 ?m in diameter, see figure 1 below. Figure 1: Schematic representations of M13 bacteriophage and its host E.coli...

  15. Immuno compatibility of Bacteriophages as Nano medicines

    International Nuclear Information System (INIS)

    Kaur, T.; Nafissi, N.; Wasfi, O.; Sheldon, K.; Wettig, Sh.; Slavcev, R.

    2012-01-01

    Bacteriophage-based medical research provides the opportunity to develop targeted nano medicines with heightened efficiency and safety profiles. Filamentous phages also can and have been formulated as targeted drug-delivery nano medicines, and phage may also serve as promising alternatives/complements to antibiotics. Over the past decade the use of phage for both the prophylaxis and the treatment of bacterial infection, has gained special significance in view of a dramatic rise in the prevalence of antibiotic resistance bacterial strains. Two potential medical applications of phages are the treatment of bacterial infections and their use as immunizing agents in diagnosis and monitoring patients with immunodeficiencies. Recently, phages have been employed as gene-delivery vectors (phage nano medicine), for nearly half a century as tools in genetic research, for about two decades as tools for the discovery of specific target-binding proteins and peptides, and for almost a decade as tools for vaccine development. As phage applications to human therapeutic development grow at an exponential rate, it will become essential to evaluate host immune responses to initial and repetitive challenges by therapeutic phage in order to develop phage therapies that offer suitable utility. This paper examines and discusses phage nano medicine applications and the immunomodulatory effects of bacteriophage exposure and treatment modalities.

  16. A bacteriophages journey through the human body.

    Science.gov (United States)

    Barr, Jeremy J

    2017-09-01

    The human body is colonized by a diverse collective of microorganisms, including bacteria, fungi, protozoa and viruses. The smallest entity of this microbial conglomerate are the bacterial viruses. Bacteriophages, or phages for short, exert significant selective pressure on their bacterial hosts, undoubtedly influencing the human microbiome and its impact on our health and well-being. Phages colonize all niches of the body, including the skin, oral cavity, lungs, gut, and urinary tract. As such our bodies are frequently and continuously exposed to diverse collections of phages. Despite the prevalence of phages throughout our bodies, the extent of their interactions with human cells, organs, and immune system is still largely unknown. Phages physically interact with our mucosal surfaces, are capable of bypassing epithelial cell layers, disseminate throughout the body and may manipulate our immune system. Here, I establish the novel concept of an "intra-body phageome," which encompasses the collection of phages residing within the classically "sterile" regions of the body. This review will take a phage-centric view of the microbiota, human body, and immune system with the ultimate goal of inspiring a greater appreciation for both the indirect and direct interactions between bacteriophages and their mammalian hosts. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  17. Optimizing Propagation of Staphylococcus aureus Infecting Bacteriophage vB_SauM-phiIPLA-RODI on Staphylococcus xylosus Using Response Surface Methodology

    OpenAIRE

    Eva González-Menéndez; Francisco Noé Arroyo-López; Beatriz Martínez; Pilar García; Antonio Garrido-Fernández; Ana Rodríguez

    2018-01-01

    The use of bacteriophages for killing pathogenic bacteria is a feasible alternative to antibiotics and disinfectants. To obtain the large quantities of phages required for this application, large-scale production of bacteriophages must be optimized. This study aims to define conditions that maximize the phage yield of the virulent and polyvalent staphylococcal bacteriophage vB_SauM-phiIPLA-RODI in broth culture, using the food-grade species Staphylococcus xylosus as the host strain to reduce ...

  18. Co-therapy using lytic bacteriophage and linezolid: effective treatment in eliminating methicillin resistant Staphylococcus aureus (MRSA from diabetic foot infections.

    Directory of Open Access Journals (Sweden)

    Sanjay Chhibber

    Full Text Available BACKGROUND: Staphylococcus aureus remains the predominant pathogen in diabetic foot infections and prevalence of methicillin resistant S.aureus (MRSA strains further complicates the situation. The incidence of MRSA in infected foot ulcers is 15-30% and there is an alarming trend for its increase in many countries. Diabetes acts as an immunosuppressive state decreasing the overall immune functioning of body and to worsen the situation, wounds inflicted with drug resistant strains represent a morbid combination in diabetic patients. Foot infections caused by MRSA are associated with an increased risk of amputations, increased hospital stay, increased expenses and higher infection-related mortality. Hence, newer, safer and effective treatment strategies are required for treating MRSA mediated diabetic foot infections. The present study focuses on the use of lytic bacteriophage in combination with linezolid as an effective treatment strategy against foot infection in diabetic population. METHODOLOGY: Acute hindpaw infection with S.aureus ATCC 43300 was established in alloxan induced diabetic BALB/c mice. Therapeutic efficacy of a well characterized broad host range lytic bacteriophage, MR-10 was evaluated alone as well as in combination with linezolid in resolving the course of hindpaw foot infection in diabetic mice. The process of wound healing was also investigated. RESULTS AND CONCLUSIONS: A single administration of phage exhibited efficacy similar to linezolid in resolving the course of hindpaw infection in diabetic animals. However, combination therapy using both the agents was much more effective in arresting the entire infection process (bacterial load, lesion score, foot myeloperoxidase activity and histopathological analysis. The entire process of tissue healing was also hastened. Use of combined agents has been known to decrease the frequency of emergence of resistant mutants, hence this approach can serve as an effective strategy in

  19. Biocontrol of Pectobacterium carotovorum subsp. carotovorum using bacteriophage PP1.

    Science.gov (United States)

    Lim, Jeong-A; Jee, Samnyu; Lee, Dong Hwan; Roh, Eunjung; Jung, Kyusuk; Oh, Changsik; Heu, Sunggi

    2013-08-01

    Pectobacterium carotovorum subsp. carotovorum (formerly Erwinia carotovora subsp. carotovora) is a plant pathogen that causes soft rot and stem rot diseases in several crops, including Chinese cabbage, potato, and tomato. To control this bacterium, we isolated a bacteriophage, PP1, with lytic activity against P. carotovorum subsp. carotovorum. Transmission electron microscopy revealed that the PP1 phage belongs to the Podoviridae family of the order Caudovirales, which exhibit icosahedral heads and short non-contractile tails. PP1 phage showed high specificity for P. carotovorum subsp. carotovorum, and several bacteria belonging to different species and phyla were resistant to PP1. This phage showed rapid and strong lytic activity against its host bacteria in liquid medium and was stable over a broad range of pH values. Disease caused by P. carotovorum subsp. carotovorum was significantly reduced by PP1 treatment. Overall, PP1 bacteriophage effectively controls P. carotovorum subsp. carotovorum.

  20. Characterization of newly isolated lytic bacteriophages active against Acinetobacter baumannii.

    Directory of Open Access Journals (Sweden)

    Maia Merabishvili

    Full Text Available Based on genotyping and host range, two newly isolated lytic bacteriophages, myovirus vB_AbaM_Acibel004 and podovirus vB_AbaP_Acibel007, active against Acinetobacter baumannii clinical strains, were selected from a new phage library for further characterization. The complete genomes of the two phages were analyzed. Both phages are characterized by broad host range and essential features of potential therapeutic phages, such as short latent period (27 and 21 min, respectively, high burst size (125 and 145, respectively, stability of activity in liquid culture and low frequency of occurrence of phage-resistant mutant bacterial cells. Genomic analysis showed that while Acibel004 represents a novel bacteriophage with resemblance to some unclassified Pseudomonas aeruginosa phages, Acibel007 belongs to the well-characterized genus of the Phikmvlikevirus. The newly isolated phages can serve as potential candidates for phage cocktails to control A. baumannii infections.

  1. The isolation and characterization of Stenotrophomonas maltophilia T4-like bacteriophage DLP6.

    Directory of Open Access Journals (Sweden)

    Danielle L Peters

    Full Text Available Increasing isolation of the extremely antibiotic resistant bacterium Stenotrophomonas maltophilia has caused alarm worldwide due to the limited treatment options available. A potential treatment option for fighting this bacterium is 'phage therapy', the clinical application of bacteriophages to selectively kill bacteria. Bacteriophage DLP6 (vB_SmoM-DLP6 was isolated from a soil sample using clinical isolate S. maltophilia strain D1571 as host. Host range analysis of phage DLP6 against 27 clinical S. maltophilia isolates shows successful infection and lysis in 13 of the 27 isolates tested. Transmission electron microscopy of DLP6 indicates that it is a member of the Myoviridae family. Complete genome sequencing and analysis of DLP6 reveals its richly recombined evolutionary history, featuring a core of both T4-like and cyanophage genes, which suggests that it is a member of the T4-superfamily. Unlike other T4-superfamily phages however, DLP6 features a transposase and ends with 229 bp direct terminal repeats. The isolation of this bacteriophage is an exciting discovery due to the divergent nature of DLP6 in relation to the T4-superfamily of phages.

  2. Use of UV-irradiated bacteriophage T6 to kill extracellular bacteria in tissue culture infectivity assays

    International Nuclear Information System (INIS)

    Shaw, D.R.; Maurelli, A.T.; Goguen, J.D.; Straley, S.C.; Curtiss, R. III

    1983-01-01

    The authors have utilized 'lysis from without' mediated by UV-inactivated bacteriophage T6 to eliminate extracellular bacteria in experiments measuring the internalization, intracellular survival and replication of Yersinia pestis within mouse peritoneal macrophages and of Shigella flexneri within a human intestinal epithelial cell line. The technique described has the following characteristics: (a) bacterial killing is complete within 15 min at 37 0 C, with a >10 3 -fold reduction in colony-forming units (CFU); (b) bacteria within cultured mammalian cells are protected from killing by UV-inactivated T6; (c) the mammalian cells are not observably affected by exposure to UV-inactivated T6. This technique has several advantages over the use of antibiotics to eliminate extracellular bacteria and is potentially widely applicable in studies of the interactions between pathogenic bacteria and host phagocytic cells as well as other target tissues. (Auth.)

  3. Complete genome analysis of two new bacteriophages isolated from impetigo strains of Staphylococcus aureus.

    Science.gov (United States)

    Botka, Tibor; Růžičková, Vladislava; Konečná, Hana; Pantůček, Roman; Rychlík, Ivan; Zdráhal, Zbyněk; Petráš, Petr; Doškař, Jiří

    2015-08-01

    Exfoliative toxin A (ETA)-coding temperate bacteriophages are leading contributors to the toxic phenotype of impetigo strains of Staphylococcus aureus. Two distinct eta gene-positive bacteriophages isolated from S. aureus strains which recently caused massive outbreaks of pemphigus neonatorum in Czech maternity hospitals were characterized. The phages, designated ϕB166 and ϕB236, were able to transfer the eta gene into a prophageless S. aureus strain which afterwards converted into an ETA producer. Complete phage genome sequences were determined, and a comparative analysis of five designed genomic regions revealed major variances between them. They differed in the genome size, number of open reading frames, genome architecture, and virion protein patterns. Their high mutual sequence similarity was detected only in the terminal regions of the genome. When compared with the so far described eta phage genomes, noticeable differences were found. Thus, both phages represent two new lineages of as yet not characterized bacteriophages of the Siphoviridae family having impact on pathogenicity of impetigo strains of S. aureus.

  4. UV ability to destroy poliovirus end FRNA specific bacteriophages

    Energy Technology Data Exchange (ETDEWEB)

    Baron, J.; Joret, J.C.; Lesavre, J.; Perrot, J.Y.

    1996-01-01

    In France, the use of ultraviolet radiation to disinfect secondary effluents is only in its initial stage. The aim of this study was to examine the ability of UV to destroy Poliovirus Type 1 and FRNA specific bacteriophages (laboratory MS2 phages and indigenous phages). Concentrated viral solutions were mixed with secondary effluents artificially enriched with suspended solids and then irradiated at various UV dose in a collimated beam. Bacteriological analysis of Escherichia coli and enterococci were performed at the same time. UV were very efficient to kill Poliovirus : Inactivation of 3 and 5 log units were observed respectively at UV doses of 20 and 40 mW/cm{sup 2}. The Poliovirus disinfection rate was almost the same than Escherichia coli. Enterococci were more resistant than E. coli. Inactivation of MS2 bacteriophages was significantly correlated to UV dose following the relationship MS2 Inactivation = 0.047{sup *} Dose + 0,396. At UV dose of 20 mWs/cm{sup 2}, MS2 phages were 2.3 times more resistant to UV than Poliovirus, i.e. they need UV dose 2,3 times greater to be disinfected at the same level. A review of the literature has also shown that viruses more resistant to UV treatment have never been reported. All this would tend to confirm the interest of this group of virus as indicators of the disinfection efficiency of UV, which could indicate, on site, the inactivation of pathogenic viruses. Inactivation rates obtained for FRNA phages proved the good virucidal activity of UV. The inactivation of indigenous FRNA bacteriophages was not correlated with E. coli inactivation. On the other hand, it was correlated with enterococci inactivation. (Author). 23 refs., 7 figs., 4 tabs.

  5. Sero-epidemiological assessment and diagnosis of visceral leishmaniasis in an endemic locality using Fast Agglutination Screening Test (FAST)

    NARCIS (Netherlands)

    Hailu, A.; Kroon, C. C. M.; Schoone, G. J.; Berhe, N.; Schallig, H. D. F. H.; Kager, P. A.

    2002-01-01

    The Fast Agglutination Screening Test (FAST) was employed on sera obtained from an endemic area of visceral leishmaniasis in southwestern Ethiopia, in February 2000. The study involved (i) active case detection among 1575 residents of two villages; and (ii) passive case detection in an outpatient

  6. Bacteriophages of Leuconostoc, Oenococcus, and Weissella

    DEFF Research Database (Denmark)

    Kot, Witold; Neve, Horst; Heller, Knut J

    2014-01-01

    Leuconostoc (Ln.), Weissella, and Oenococcus form a group of related genera of lactic acid bacteria, which once all shared the name Leuconostoc. They are associated with plants, fermented vegetable products, raw milk, dairy products, meat, and fish. Most of industrially relevant Leuconostoc strains...... can be classified as either Ln. mesenteroides or Ln. pseudomesenteroides. They are important flavor producers in dairy fermentations and they initiate nearly all vegetable fermentations. Therefore, bacteriophages attacking Leuconostoc strains may negatively influence the production process....... Bacteriophages attacking Leuconostoc strains were first reported in 1946. Since then, the majority of described Leuconostoc phages was isolated from either dairy products or fermented vegetable products. Both lytic and temperate phages of Leuconostoc were reported. Most of Leuconostoc phages examined using...

  7. The Baseplate of Lactobacillus delbrueckii Bacteriophage Ld17 Harbors a Glycerophosphodiesterase.

    Science.gov (United States)

    Cornelissen, Anneleen; Sadovskaya, Irina; Vinogradov, Evgeny; Blangy, Stéphanie; Spinelli, Silvia; Casey, Eoghan; Mahony, Jennifer; Noben, Jean-Paul; Dal Bello, Fabio; Cambillau, Christian; van Sinderen, Douwe

    2016-08-05

    Glycerophosphodiester phosphodiesterases (GDPDs; EC 3.1.4.46) typically hydrolyze glycerophosphodiesters to sn-glycerol 3-phosphate (Gro3P) and their corresponding alcohol during patho/physiological processes in bacteria and eukaryotes. GDPD(-like) domains were identified in the structural particle of bacterial viruses (bacteriophages) specifically infecting Gram-positive bacteria. The GDPD of phage 17 (Ld17; GDPDLd17), representative of the group b Lactobacillus delbrueckii subsp. bulgaricus (Ldb)-infecting bacteriophages, was shown to hydrolyze, besides the simple glycerophosphodiester, two complex surface-associated carbohydrates of the Ldb17 cell envelope: the Gro3P decoration of the major surface polysaccharide d-galactan and the oligo(glycerol phosphate) backbone of the partially glycosylated cell wall teichoic acid, a minor Ldb17 cell envelope component. Degradation of cell wall teichoic acid occurs according to an exolytic mechanism, and Gro3P substitution is presumed to be inhibitory for GDPDLd17 activity. The presence of the GDPDLd17 homotrimer in the viral baseplate structure involved in phage-host interaction together with the dependence of native GDPD activity, adsorption, and efficiency of plating of Ca(2+) ions supports a role for GDPDLd17 activity during phage adsorption and/or phage genome injection. In contrast to GDPDLd17, we could not identify any enzymatic activity for the GDPD-like domain in the neck passage structure of phage 340, a 936-type Lactococcus lactis subsp. lactis bacteriophage. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. [Diagnosis of human brucellosis. Role of pH in the seroagglutination test and influence of pH on the agglutinating activity of IgM, IgG and IgA antibodies].

    Science.gov (United States)

    Rubio Vallejo, Manuel; del Pozo, José L; Del Pozo León, José Luis; Hernández-Molina, Juan Manuel; Dorronsoro Ibero, Inés; Marrodán Ciordia, Teresa; Díaz García, Ramón

    2002-04-01

    To evaluate the role of pH in the seroagglutination test (SAT)and Rose Bengal (RB) test, and to determine the influence of pH on the agglutinating activity of IgM, IgG and IgA antibodies. The SAT was performed at pH 7.2 or pH 5.0 in standard microtiter-type polystyrene plates using Ring Test antigen or the Brucella suspension (BRUCAPT) provided in the Brucellacapt kits. Specific antibodies against native hapten were determined by radial immunodiffusion. Additionally, IgG, IgA and IgM fractions were separated from 8 sera by absorption chromatography and their agglutinating capacity was studied at pH 7.2 and 5.0. We studied 72 sera from patients with clinical brucellosis taken at the time of hospitalization, 16 from persons in contact with infected animals, and 16 from healthy donors. SAT results at pH 5.0 correlated with those obtained with the Rose Bengal test. Four Rose Bengal-positive sera were found to be SAT-negative at pH 7.2 and SAT-positive at pH 5.0. SAT performed at pH 5.0 with BRUCAPT antigen yielded higher titers than tests performed at pH 7.2 or 5.0 with Ring Test antigen (p IgA fractions were SAT-negative at pH 7.2 and SAT-positive at pH 5.0; the other 5 agglutinated at both pH conditions and were DTT-sensitive. All IgA fractions but one were positive by Rose Bengal. Agglutinating activity of the IgM fraction was not affected by pH. The SAT performed with the buffer and antigen suspension included in the Brucellacapt kit (pH 5.0) is highly useful for detecting agglutinating and non-agglutinating antibodies at pH 7.2.

  9. Biodiversity of bacteriophages: morphological and biological properties of a large group of phages isolated from urban sewage

    OpenAIRE

    Agata Jurczak-Kurek; Tomasz Gąsior; Bożena Nejman-Faleńczyk; Sylwia Bloch; Aleksandra Dydecka; Gracja Topka; Agnieszka Necel; Magdalena Jakubowska-Deredas; Magdalena Narajczyk; Malwina Richert; Agata Mieszkowska; Borys Wróbel; Grzegorz Węgrzyn; Alicja Węgrzyn

    2016-01-01

    A large scale analysis presented in this article focuses on biological and physiological variety of bacteriophages. A collection of 83 bacteriophages, isolated from urban sewage and able to propagate in cells of different bacterial hosts, has been obtained (60 infecting Escherichia coli, 10 infecting Pseudomonas aeruginosa, 4 infecting Salmonella enterica, 3 infecting Staphylococcus sciuri, and 6 infecting Enterococcus faecalis). High biological diversity of the collection is indicated by its...

  10. 76 FR 66187 - Bacteriophage of Clavibacter Michiganensis Subspecies Michiganensis; Exemption From the...

    Science.gov (United States)

    2011-10-26

    ... history of bacteriophage laboratory and pesticidal usage, adverse reports in the literature have not been... cheese factory in Argentina. Journal of Dairy Science 89:3791-3799. 19. Guillaumes J, Houdeau G, Germain...

  11. Arthropod Innate Immune Systems and Vector-Borne Diseases.

    Science.gov (United States)

    Baxter, Richard H G; Contet, Alicia; Krueger, Kathryn

    2017-02-21

    Arthropods, especially ticks and mosquitoes, are the vectors for a number of parasitic and viral human diseases, including malaria, sleeping sickness, Dengue, and Zika, yet arthropods show tremendous individual variation in their capacity to transmit disease. A key factor in this capacity is the group of genetically encoded immune factors that counteract infection by the pathogen. Arthropod-specific pattern recognition receptors and protease cascades detect and respond to infection. Proteins such as antimicrobial peptides, thioester-containing proteins, and transglutaminases effect responses such as lysis, phagocytosis, melanization, and agglutination. Effector responses are initiated by damage signals such as reactive oxygen species signaling from epithelial cells and recognized by cell surface receptors on hemocytes. Antiviral immunity is primarily mediated by siRNA pathways but coupled with interferon-like signaling, antimicrobial peptides, and thioester-containing proteins. Molecular mechanisms of immunity are closely linked to related traits of longevity and fertility, and arthropods have the capacity for innate immunological memory. Advances in understanding vector immunity can be leveraged to develop novel control strategies for reducing the rate of transmission of both ancient and emerging threats to global health.

  12. Comparative transcriptomic analysis of shrimp hemocytes in response to acute hepatopancreas necrosis disease (AHPND) causing Vibrio parahemolyticus infection.

    Science.gov (United States)

    Zheng, Zhihong; Wang, Fan; Aweya, Jude Juventus; Li, Ruiwei; Yao, Defu; Zhong, Mingqi; Li, Shengkang; Zhang, Yueling

    2018-03-01

    The recent emergence of acute hepatopancreas necrosis disease (AHPND) in shrimps has posed a major challenge in the shrimp aquaculture industry. The Pir toxin proteins carried by some strains of Vibrio parahaemolyticus are believed to play essential roles in the pathogenesis of AHPND. However, few studies have so far explored how the host immune system responds to these bacteria. In this study, AHPND V. parahaemolyticus (with Pir) and non-AHPND V. parahaemolyticus (without Pir) were injected into two groups of shrimps, and the hemocytes collected for comparative transcriptomic analyses. A total of 1064 differentially expressed genes (DEGs) were identified, of which 910 were up-regulated and 154 were down-regulated. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that many DEGs were involved in a number of biological processes such as cellular process, metabolic process and single-organism process in the AHPND V. parahaemolyticus injected group than the non-AHPND V. parahaemolyticus injected group. Among these, major metabolic processes such as carbohydrate metabolism, lipid metabolism and amino acid metabolism were further identified as the major responsive gene groups. We observed that genes involved in cell growth and anti-apoptosis including src, iap2, cas2, cytochrome P450, gst and cytochromecoxidase were strongly activated in the AHPND V. parahaemolyticus group than in the non-AHPND V. parahaemolyticus group. Collectively, our results unveiled that shrimp hemocytes respond to AHPND related strain of Vibrio parahaemolyticus infection at the transcriptional level, which is useful in furthering our understanding of AHPND. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Interactions of the cell-wall glycopolymers of lactic acid bacteria with their bacteriophages

    Directory of Open Access Journals (Sweden)

    Marie-Pierre eChapot-Chartier

    2014-05-01

    Full Text Available Lactic acid bacteria (LAB are Gram positive bacteria widely used in the production of fermented food in particular cheese and yoghurts. Bacteriophage infections during fermentation processes have been for many years a major industrial concern and have stimulated numerous research efforts. Better understanding of the molecular mechanisms of bacteriophage interactions with their host bacteria is required for the development of efficient strategies to fight against infections. The bacterial cell wall plays key roles in these interactions. First, bacteriophages must adsorb at the bacterial surface through specific interactions with receptors that are cell wall components. At next step, phages must overcome the barrier constituted by cell wall peptidoglycan to inject DNA inside bacterial cell. Also at the end of the infection cycle, phages synthesize endolysins able to hydrolyze peptidoglycan and lyse bacterial cells to release phage progeny. In the last decade, concomitant development of genomics and structural analysis of cell wall components allowed considerable advances in the knowledge of their structure and function in several model LAB. Here, we describe the present knowledge on the structure of the cell wall glycopolymers of the best characterized LAB emphasizing their structural variations and we present the available data regarding their role in bacteria-phage specific interactions at the different steps of the infection cycle.

  14. Oxidative stress and cytotoxicity elicited lipid peroxidation in hemocytes of Bombyx mori larva infested with dipteran parasitoid, Exorista bombycis.

    Science.gov (United States)

    Pooja, Makwana; Pradeep, Appukuttan Nair R; Hungund, Shambhavi P; Sagar, Chandrashekhar; Ponnuvel, Kangayam M; Awasthi, Arvind K; Trivedy, Kanika

    2017-12-20

    Parasitization of silkworm, Bombyx mori by invasive larva of dipteran parasitoid Exorista bombycis caused upto 20% revenue loss in sericulture. The parasitism was successful by suppressing host immune system however mechanism of immune suppression induced by E. bombycis is unknown which is unravelled here. The infestation induced cytotoxic symptoms in host hemocytes, such as vacuolated cytoplasm, porous plasma membrane, indented nuclei with condensed chromatin and dilated RER. One of the markers of necrosis is cell permeabilization, which can be measured as released lactate dehydrogenase (LDH). LDH level showed significantly (Pmori.

  15. Streptophage-mediated control of off-flavour taint producing streptomycetes isolated from barramundi ponds

    Directory of Open Access Journals (Sweden)

    Jodi Anne Jonns

    2017-06-01

    Following the determination of the streptophage susceptibility of the isolates one of the most odourous streptomycete species (USC-14510 was selected to be tested further using different pond simulations resembling real-life applications. Geosmin was tested as the indicator of off-flavour taint production and as it has been previously reported that the cyanobacteria-actinomycete interactions occurring in ponds result in even greater levels of geosmin and 2-methylisoborneol, the geosmin levels for the isolate in the presence of cyanobacteria and streptophages were also tested. Findings indicated that the highly odourous Streptomyces species (USC-14510 once infected with streptophages, can lose its capacity to produce off-flavour taints. Pond simulation studies also revealed geosmin production was significantly reduced when streptophages were introduced into the pond water where streptomycete species were grown. The bacteriophage control method developed in the presented study might again confirm significant potential for the bacteriophage-mediated remediation strategy to be adapted by the aquaculture industry.

  16. The effect of Beauveria bassiana infection on cell mediated and humoral immune response in house fly, Musca domestica L.

    Science.gov (United States)

    Mishra, Sapna; Kumar, Peeyush; Malik, Anushree

    2015-10-01

    Entomopathogenic fungi that manifest infections by overcoming insect's immune response could be a successful control agent for the house fly, Musca domestica L. which is a major domestic, medical, and veterinary pest. In this study, the immune response of house fly to Beauveria bassiana infection was investigated to reveal fundamental aspects of house fly hemocyte biology, such as hemocyte numbers and size, which is poorly understood. The total hemocyte counts (THCs) in B. bassiana-infected house fly showed an initial increase (from 6 to 9 h), followed by subsequent decrease (9 to 12 h) with increase in time of infection. The THCs was slightly greater in infected flies than the non-infected ones. Insight into relative hemocyte counts depicted a significant increase in prohemocyte (PR) and decrease in granulocyte (GR) in infected house flies compared to non-infected ones. The relative cell area of hemocyte cells showed a noticeable increase in PR and intermediate cells (ICs), while a considerable reduction was observed for plasmatocyte (PL) and GR. The considerable variation in relative cell number and cell area in the B. bassiana-infected house flies indicated stress development during infection. The present study highlights changes occurring during B. bassiana invasion to house fly leading to establishment of infection along with facilitation in understanding of basic hemocyte biology. The results of the study is expected to help in better understanding of house fly immune response during fungal infection, so as to assist production of more efficient mycoinsecticides for house fly control using B. bassiana.

  17. Neuraminidase-mediated haemagglutination of recent human influenza A(H3N2) viruses is determined by arginine 150 flanking the neuraminidase catalytic site.

    Science.gov (United States)

    Mögling, Ramona; Richard, Mathilde J; Vliet, Stefan van der; Beek, Ruud van; Schrauwen, Eefje J A; Spronken, Monique I; Rimmelzwaan, Guus F; Fouchier, Ron A M

    2017-06-01

    Over the last decade, an increasing proportion of circulating human influenza A(H3N2) viruses exhibited haemagglutination activity that was sensitive to neuraminidase inhibitors. This change in haemagglutination as compared to older circulating A(H3N2) viruses prompted an investigation of the underlying molecular basis. Recent human influenza A(H3N2) viruses were found to agglutinate turkey erythrocytes in a manner that could be blocked with either oseltamivir or neuraminidase-specific antisera, indicating that agglutination was driven by neuraminidase, with a low or negligible contribution of haemagglutinin. Using representative virus recombinants it was shown that the haemagglutinin of a recent A(H3N2) virus indeed had decreased activity to agglutinate turkey erythrocytes, while its neuraminidase displayed increased haemagglutinating activity. Viruses with chimeric and mutant neuraminidases were used to identify the amino acid substitution histidine to arginine at position 150 flanking the neuraminidase catalytic site as the determinant of this neuraminidase-mediated haemagglutination. An analysis of publicly available neuraminidase gene sequences showed that viruses with histidine at position 150 were rapidly replaced by viruses with arginine at this position between 2005 and 2008, in agreement with the phenotypic data. As a consequence of neuraminidase-mediated haemagglutination of recent A(H3N2) viruses and poor haemagglutination via haemagglutinin, haemagglutination inhibition assays with A(H3N2) antisera are no longer useful to characterize the antigenic properties of the haemagglutinin of these viruses for vaccine strain selection purposes. Continuous monitoring of the evolution of these viruses and potential consequences for vaccine strain selection remains important.

  18. Computational determination of the effects of virulent Escherichia coli and salmonella bacteriophages on human gut.

    Science.gov (United States)

    Mostafa, Marwa Mostafa; Nassef, Mohammad; Badr, Amr

    2016-10-01

    Salmonella and Escherichia coli are different types of bacteria that cause food poisoning in humans. In the elderly, infants and people with chronic conditions, it is very dangerous if Salmonella or E. coli gets into the bloodstream and then they must be treated by phage therapy. Treating Salmonella and E. coli by phage therapy affects the gut flora. This research paper presents a system for detecting the effects of virulent E. coli and Salmonella bacteriophages on human gut. A method based on Domain-Domain Interactions (DDIs) model is implemented in the proposed system to determine the interactions between the proteins of human gut bacteria and the proteins of bacteriophages that infect virulent E. coli and Salmonella. The system helps gastroenterologists to realize the effect of injecting bacteriophages that infect virulent E. coli and Salmonella on the human gut. By testing the system over Enterobacteria phage 933W, Enterobacteria phage VT2-Sa and Enterobacteria phage P22, it resulted in four interactions between the proteins of the bacteriophages that infect E. coli O157:H7, E. coli O104:H4 and Salmonella typhimurium and the proteins of human gut bacterium strains. Several effects were detected such as: antibacterial activity against a number of bacterial species in human gut, regulation of cellular differentiation and organogenesis during gut, lung, and heart development, ammonia assimilation in bacteria, yeasts, and plants, energizing defense system and its function in the detoxification of lipopolysaccharide, and in the prevention of bacterial translocation in human gut. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  19. Methods for Isolation, Purification, and Propagation of Bacteriophages of Campylobacter jejuni

    DEFF Research Database (Denmark)

    Gencay, Yilmaz Emre; Birk, Tina; Sørensen, Martine Camilla Holst

    2017-01-01

    Here, we describe the methods for isolation, purification, and propagation of Campylobacter jejuni bacteriophages from samples expected to contain high number of phages such as chicken feces. The overall steps are (1) liberation of phages from the sample material; (2) observation of plaque-formin...

  20. Molecular characterization of podoviral bacteriophages virulent for Clostridium perfringens and their comparison with members of the Picovirinae.

    Directory of Open Access Journals (Sweden)

    Nikolay V Volozhantsev

    Full Text Available Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium responsible for human food-borne disease as well as non-food-borne human, animal and poultry diseases. Because bacteriophages or their gene products could be applied to control bacterial diseases in a species-specific manner, they are potential important alternatives to antibiotics. Consequently, poultry intestinal material, soil, sewage and poultry processing drainage water were screened for virulent bacteriophages that lysed C. perfringens. Two bacteriophages, designated ΦCPV4 and ΦZP2, were isolated in the Moscow Region of the Russian Federation while another closely related virus, named ΦCP7R, was isolated in the southeastern USA. The viruses were identified as members of the order Caudovirales in the family Podoviridae with short, non-contractile tails of the C1 morphotype. The genomes of the three bacteriophages were 17.972, 18.078 and 18.397 kbp respectively; encoding twenty-six to twenty-eight ORF's with inverted terminal repeats and an average GC content of 34.6%. Structural proteins identified by mass spectrometry in the purified ΦCP7R virion included a pre-neck/appendage with putative lyase activity, major head, tail, connector/upper collar, lower collar and a structural protein with putative lysozyme-peptidase activity. All three podoviral bacteriophage genomes encoded a predicted N-acetylmuramoyl-L-alanine amidase and a putative stage V sporulation protein. Each putative amidase contained a predicted bacterial SH3 domain at the C-terminal end of the protein, presumably involved with binding the C. perfringens cell wall. The predicted DNA polymerase type B protein sequences were closely related to other members of the Podoviridae including Bacillus phage Φ29. Whole-genome comparisons supported this relationship, but also indicated that the Russian and USA viruses may be unique members of the sub-family Picovirinae.

  1. Isolation and Characterization of a Bacteriophage Preying an Antifungal Bacterium

    Directory of Open Access Journals (Sweden)

    Aryan Rahimi-Midani

    2016-12-01

    Full Text Available Several Bacillus species were isolated from rice field soils, and 16S rRNA gene sequence analysis showed that Bacillus cereus was the most abundant. A strain named BC1 showed antifungal activity against Rhizoctonia solani. Bacteriophages infecting strain BC1 were isolated from the same soil sample. The isolated phage PK16 had an icosahedral head of 100 ± 5 nm and tail of 200 ± 5 nm, indicating that it belonged to the family Myoviridae. Analysis of the complete linear dsDNA genome revealed a 158,127-bp genome with G + C content of 39.9% comprising 235 open reading frames as well as 19 tRNA genes (including 1 pseudogene. Blastp analysis showed that the proteins encoded by the PK16 genome had the closest hits to proteins of seven different bacteriophages. A neighbor-joining phylogenetic tree based on the major capsid protein showed a robust clustering of phage PK16 with phage JBP901 and BCP8-2 isolated from Korean fermented food.

  2. Bacteriophage prehistory: Is or is not Hankin, 1896, a phage reference?

    Science.gov (United States)

    Abedon, Stephen T; Thomas-Abedon, Cameron; Thomas, Anne; Mazure, Hubert

    2011-05-01

    We identified 30 actual or presumptive "bacteriophage" references dating between the years 1895 and 1917 and have further explored one of the oldest: Hankin's 1896 study of a bactericidal action associated with the waters of the Ganges and Jumna rivers in India. As Hankin's work took place approximately 20 years prior to the actual discovery of bacteriophages, no claims were made as to a possible phage nature of the phenomenon. Here we suggest that it may be imprudent to assume nevertheless that it represents an early observation of phagemediated bactericidal activity. Our principal argument is that the antibacterial aspect of these river waters was able to retain full potency following "heating" for one-half hour in hermetically sealed tubes, where heating in "open" tubes resulted in loss of antibacterial activity. We also suggest that environmental phage counts would have had to have been unusually high-greater than 10(6)/ml impacting a single host strain-to achieve the rates of bacterial loss that Hankin observed.

  3. Bioactive extracts of red seaweeds Pterocladiella capillacea and Osmundaria obtusiloba (Floridophyceae: Rhodophyta) with antioxidant and bacterial agglutination potential.

    Science.gov (United States)

    de Alencar, Daniel Barroso; de Carvalho, Fátima Cristiane Teles; Rebouças, Rosa Helena; Dos Santos, Daniel Rodrigues; Dos Santos Pires-Cavalcante, Kelma Maria; de Lima, Rebeca Larangeira; Baracho, Bárbara Mendes; Bezerra, Rayssa Mendes; Viana, Francisco Arnaldo; Dos Fernandes Vieira, Regine Helena Silva; Sampaio, Alexandre Holanda; de Sousa, Oscarina Viana; Saker-Sampaio, Silvana

    2016-04-01

    To evaluate the antioxidant, antibacterial and bacterial cell agglutination activities of the hexane (Hex) and 70% ethanol (70% EtOH) extracts of two species of red seaweeds Pterocladiella capillacea (P. capillacea) and Osmundaria obtusiloba. In vitro antioxidant activity was determined by DPPH radical scavenging assay, ferric-reducing antioxidant power assay, ferrous ion chelating assay, β-carotene bleaching assay and total phenolic content quantification. Antimicrobial activity was tested using the method of disc diffusion on Mueller-Hinton medium. The ability of algal extracts to agglutinate bacterial cells was also tested. The 70% EtOH extract of the two algae showed the highest values of total phenolic content compared to the Hex extract. The results of DPPH for both extracts (Hex, 70% EtOH) of Osmundaria obtusiloba (43.46% and 99.47%) were higher than those of P. capillacea (33.04% and 40.81%) at a concentration of 1000 μg/mL. As for the ferrous ion chelating, there was an opposite behavior, extracts of P. capillacea had a higher activity. The extracts showed a low ferric-reducing antioxidant power, with optical density ranging from 0.054 to 0.180. Antioxidant activities of all extracts evaluated for β-carotene bleaching were above 40%. There was no antibacterial activity against bacterial strains tested. However, the extracts of both species were able to agglutinate bacterial Gram positive cells of Staphylococcus aureus and Gram negative cells of Escherichia coli, multidrug-resistant Salmonella and Vibrio harveyi. This is the first report of the interaction between these algal extracts, rich in natural compounds with antioxidant potential, and Gram positive and Gram negative bacterial cells. Copyright © 2016 Hainan Medical College. Production and hosting by Elsevier B.V. All rights reserved.

  4. Environmental Bacteriophages of the Emerging Enterobacterial Phytopathogen, Dickeya solani, Show Genomic Conservation and Capacity for Horizontal Gene Transfer between Their Bacterial Hosts

    Directory of Open Access Journals (Sweden)

    Andrew Day

    2017-08-01

    Full Text Available Dickeya solani is an economically important phytopathogen widespread in mainland Europe that can reduce potato crop yields by 25%. There are no effective environmentally-acceptable chemical systems available for diseases caused by Dickeya. Bacteriophages have been suggested for use in biocontrol of this pathogen in the field, and limited field trials have been conducted. To date only a small number of bacteriophages capable of infecting D. solani have been isolated and characterized, and so there is a need to expand the repertoire of phages that may have potential utility in phage therapy strategies. Here we describe 67 bacteriophages from environmental sources, the majority of which are members of the viral family Myoviridae. Full genomic sequencing of two isolates revealed a high degree of DNA identity with D. solani bacteriophages isolated in Europe in the past 5 years, suggesting a wide ecological distribution of this phage family. Transduction experiments showed that the majority of the new environmental bacteriophages are capable of facilitating efficient horizontal gene transfer. The possible risk of unintentional transfer of virulence or antibiotic resistance genes between hosts susceptible to transducing phages cautions against their environmental use for biocontrol, until specific phages are fully tested for transduction capabilities.

  5. Making antigen of trypanosoma evansi and its examination by catt (card agglutination test) method

    International Nuclear Information System (INIS)

    Arifin, M; Irtisam; Estikoma Dyah; Yulia Ernawati; Boky, J.T

    1998-01-01

    An experiment was carried out for making antigen (Ag) of T. evansi by using experimental animals rats and guinea pigs for for developing parasites, and in cattle for making immune serum and normal serum. The parasites were irradiated by gamma rays ( 60 Co) at a dose of 300 Gy. The determination of antigen was done by using Card Agglutination Test (CATT) to the field samples serum of cattle. The results obtained showed that antigen was good enough potently and stable at minus 70 0 C storage during five months. (author)

  6. Latex agglutination using the periplasmic proteins antigen of Brucella melitensis is a successful, rapid, and specific serodiagnostic test for ovine brucellosis.

    Science.gov (United States)

    Ismael, Alaa Bassuny; Swelum, Ayman Abdel-Aziz; Mostafa, Salama A-H; Alhumiany, Abdel-Rahman A

    2016-09-01

    Brucellosis, especially caused by Brucella melitensis, is considered the most-widespread zoonosis in the world, particularly in developing countries. This study was planned to develop an accurate test for diagnosis of ovine brucellosis using a specific hot saline extracted soluble Brucella melitensis periplasmic proteins (SBPPs). The efficacy of the latex agglutination test (LAT) using SBPPs compared to the Rose Bengal test (RBT), buffered plate agglutination test (BPAT), serum agglutination test (SAT), and an indirect enzyme-linked immunosorbent assay (i-ELISA) was evaluated in the field diagnosis of ovine brucellosis. The test performance was evaluated by estimating sensitivity (Se), specificity (Sp), positive predictive value (PPV), negative predictive value (NPV), disease prevalence (DP), positive likelihood ratio (PLR), and negative likelihood ratio (NLR) using test agreement and bacteriological culture in 1777 samples. The false-positive result was significantly (P ⩽0.05) lower in LAT than RBT, BPAT, SAT, and i-ELISA. With reference to test agreement, the Se, Sp, PPV, and PLR were highest (P ⩽0.05) in LAT 99.33%, 99.88%, 98.68%, and 827.25%, respectively. With reference to bacteriological culture, the LAT and i-ELISA tests showed a significant difference in Se with SAT. However, no significant difference in specificity was detected. The DP was 8.44% in the five tests. In conclusion, LAT using SBPPs of B. melitensis could be a suitable serodiagnostic field test for ovine brucellosis, with high sensitivity and specificity. © The Author(s) 2016.

  7. Four new species of deep water agglutinated foraminifera from the Oligocene-Miocene of the Congo Fan (offshore Angola)

    OpenAIRE

    Kender, S.; Kaminski, M. A.; Jones, R. W.

    2006-01-01

    Four new species of deep-water agglutinated benthic foraminifera are described from the Oligocene and Miocene of the Congo Fan, offshore Angola. Scherochorella congoensis n.sp., Paratrochamminoides goroyskiformis n.sp., Haplophragmoides nauticus n.sp. and Portatrochammina profunda n.sp. all occur in deep-sea turbiditic shales and sands from the distal section of the Congo Fan.

  8. Initiation and termination of the bacteriophage phi X174 rolling circle DNA replication in vivo: packaging of plasmid single-stranded DNA into bacteriophage phi X174 coats

    NARCIS (Netherlands)

    van der Ende, A.; Teertstra, R.; Weisbeek, P. J.

    1982-01-01

    The bacteriophage phi X174 viral (+) origin when inserted in a plasmid can interact in vivo with the A protein produced by infecting phi X174 phages. A consequence of this interaction is packaging of single-stranded plasmid DNA into preformed phage coats resulting in infective particles (1). This

  9. Effectiveness of lytic bacteriophages in reducing E. coli O157:H7 populations introduced through cross-contamination on fresh cut lettuce

    Science.gov (United States)

    Previous research has shown that lytic bacteriophages (phages) can kill E. coli O157:H7 on produce surfaces. The role of lytic bacteriophages in preventing cross contamination of produce has not been evaluated. A cocktail of three lytic phages specific for E. coli O157:H7 (EcoShield) at 10^8 PFU/m...

  10. T4-related bacteriophage LIMEstone isolates for the control of soft rot on potato caused by 'Dickeya solani'.

    Directory of Open Access Journals (Sweden)

    Evelien M Adriaenssens

    Full Text Available The bacterium 'Dickeya solani', an aggressive biovar 3 variant of Dickeya dianthicola, causes rotting and blackleg in potato. To control this pathogen using bacteriophage therapy, we isolated and characterized two closely related and specific bacteriophages, vB_DsoM_LIMEstone1 and vB_DsoM_LIMEstone2. The LIMEstone phages have a T4-related genome organization and share DNA similarity with Salmonella phage ViI. Microbiological and molecular characterization of the phages deemed them suitable and promising for use in phage therapy. The phages reduced disease incidence and severity on potato tubers in laboratory assays. In addition, in a field trial of potato tubers, when infected with 'Dickeya solani', the experimental phage treatment resulted in a higher yield. These results form the basis for the development of a bacteriophage-based biocontrol of potato plants and tubers as an alternative for the use of antibiotics.

  11. Key Players in the Genetic Switch of Bacteriophage TP901-1

    DEFF Research Database (Denmark)

    Alsing, Anne; Pedersen, Margit; Sneppen, Kim

    2011-01-01

    the bistable genetic switch of bacteriophage TP901-1 through experiments and statistical mechanical modeling. We examine the activity of the lysogenic promoter Pr at different concentrations of the phage repressor, CI, and compare the effect of CI on Pr in the presence or absence of the phage-encoded MOR...

  12. Interaction of packaging motor with the polymerase complex of dsRNA bacteriophage

    International Nuclear Information System (INIS)

    Lisal, Jiri; Kainov, Denis E.; Lam, TuKiet T.; Emmett, Mark R.; Wei Hui; Gottlieb, Paul; Marshall, Alan G.; Tuma, Roman

    2006-01-01

    Many viruses employ molecular motors to package their genomes into preformed empty capsids (procapsids). In dsRNA bacteriophages the packaging motor is a hexameric ATPase P4, which is an integral part of the multisubunit procapsid. Structural and biochemical studies revealed a plausible RNA-translocation mechanism for the isolated hexamer. However, little is known about the structure and regulation of the hexamer within the procapsid. Here we use hydrogen-deuterium exchange and mass spectrometry to delineate the interactions of the P4 hexamer with the bacteriophage phi12 procapsid. P4 associates with the procapsid via its C-terminal face. The interactions also stabilize subunit interfaces within the hexamer. The conformation of the virus-bound hexamer is more stable than the hexamer in solution, which is prone to spontaneous ring openings. We propose that the stabilization within the viral capsid increases the packaging processivity and confers selectivity during RNA loading

  13. Bacteriophage amplification assay for detection of Listeria spp. using virucidal laser treatment

    Directory of Open Access Journals (Sweden)

    I.C. Oliveira

    2012-09-01

    Full Text Available A protocol for the bacteriophage amplification technique was developed for quantitative detection of viable Listeria monocytogenes cells using the A511 listeriophage with plaque formation as the end-point assay. Laser and toluidine blue O (TBO were employed as selective virucidal treatment for destruction of exogenous bacteriophage. Laser and TBO can bring a total reduction in titer phage (ca. 10(8 pfu/mL without affecting the viability of L. monocytogenes cells. Artificially inoculated skimmed milk revealed mean populations of the bacteria as low as between 13 cfu/mL (1.11 log cfu/mL, after a 10-h assay duration. Virucidal laser treatment demonstrated better protection of Listeria cells than the other agents previously tested. The protocol was faster and easier to perform than standard procedures. This protocol constitutes an alternative for rapid, sensitive and quantitative detection of L. monocytogenes.

  14. Montmorillonite-induced Bacteriophage φ6 Disassembly

    Science.gov (United States)

    Trusiak, A.; Gottlieb, P.; Katz, A.; Alimova, A.; Steiner, J. C.; Block, K. A.

    2012-12-01

    It is estimated that there are 1031 virus particles on Earth making viruses an order of magnitude more prevalent in number than prokaryotes with the vast majority of viruses being bacteriophages. Clays are a major component of soils and aquatic sediments and can react with RNA, proteins and bacterial biofilms. The clays in soils serve as an important moderator between phage and their host bacteria, helping to preserve the evolutionary balance. Studies on the effects of clays on viral infectivity have given somewhat contradictory results; possibly a consequence of clay-virus interactions being dependent on the unique structure of particular viruses. In this work, the interaction between montmorillonite and the bacteriophage φ6 is investigated. φ6 is a member of the cystovirus family that infects Pseudomonas syringe, a common plant pathogen. As a member of the cystovirus family with an enveloped structure, φ6 serves as a model for reoviruses, a human pathogen. Experiments were conducted with φ6 suspended in dilute, purified homoionic commercial-grade montmorillonite over a range of virus:clay ratios. At a 1:100000 virus:clay ratio, the clay reduced viral infectivity by 99%. The minimum clay to virus ratio which results in a measurable reduction of P. syringae infection is 1:1. Electron microscopy demonstrates that mixed suspensions of smectite and virus co-aggregate to form flocs encompassing virions within the smectite. Both free viral particles as well as those imbedded in the flocs are seen in the micrographs to be missing the envelope- leaving only the nucleocapsid (NC) intact; indicating that smectite inactivates the virus by envelope disassembly. These results have strong implications in the evolution of both the φ6 virus and its P. syringae host cells. TEM of aggregate showing several disassembled NCs.

  15. Bacteriophages of Leuconostoc, Oenococcus and Weissella

    Directory of Open Access Journals (Sweden)

    Witold P. Kot

    2014-04-01

    Full Text Available Leuconostoc (Ln., Weissella and Oenococcus form a group of related genera of lactic acid bacteria, which once all shared the name Leuconostoc. They are associated with plants, fermented vegetable products, raw milk, dairy products, meat and fish. Most of industrially relevant Leuconostoc strains can be classified as either Ln. mesenteroides or Ln. pseudomesenteroides. They are important flavor producers in dairy fermentations and they initiate nearly all vegetable fermentations. Therefore bacteriophages attacking Leuconostoc strains may negatively influence the production process. Bacteriophages attacking Leuconostoc strains were first reported in 1946. Since then, the majority of described Leuconostoc phages was isolated from either dairy products or fermented vegetable products. Both lytic and temperate phages of Leuconostoc were reported. Most of Leuconostoc phages examined using electron microscopy belong to the Siphoviridae family and differ in morphological details. Hybridization and comparative genomic studies of Leuconostoc phages suggest that they can be divided into several groups, however overall diversity of Leuconostoc phages is much lower as compared to e.g. lactococcal phages. Several fully sequenced genomes of Leuconostoc phages have been deposited in public databases. Lytic phages of Leuconostoc can be divided into two host species-specific groups with similarly organized genomes that shared very low nucleotide similarity. Phages of dairy Leuconostoc have rather limited host-ranges. The receptor binding proteins of two lytic Ln. pseudomesenteroides phages have been identified. Molecular tools for detection of dairy Leuconostoc phages have been developed. The rather limited data on phages of Oenococcus and Weissella show that i lysogeny seems to be abundant in Oenococcus strains, and ii several phages infecting Weissella cibaria are also able to productively infect strains of other Weissella species and even strains of the genus

  16. Bacteriophages show promise as antimicrobial agents.

    Science.gov (United States)

    Alisky, J; Iczkowski, K; Rapoport, A; Troitsky, N

    1998-01-01

    The emergence of antibiotic-resistant bacteria has prompted interest in alternatives to conventional drugs. One possible option is to use bacteriophages (phage) as antimicrobial agents. We have conducted a literature review of all Medline citations from 1966-1996 that dealt with the therapeutic use of phage. There were 27 papers from Poland, the Soviet Union, Britain and the U.S.A. The Polish and Soviets administered phage orally, topically or systemically to treat a wide variety of antibiotic-resistant pathogens in both adults and children. Infections included suppurative wound infections, gastroenteritis, sepsis, osteomyelitis, dermatitis, empyemas and pneumonia; pathogens included Staphylococcus, Streptococcus, Klebsiella, Escherichia, Proteus, Pseudomonas, Shigella and Salmonella spp. Overall, the Polish and Soviets reported success rates of 80-95% for phage therapy, with rare, reversible gastrointestinal or allergic side effects. However, efficacy of phage was determined almost exclusively by qualitative clinical assessment of patients, and details of dosages and clinical criteria were very sketchy. There were also six British reports describing controlled trials of phage in animal models (mice, guinea pigs and livestock), measuring survival rates and other objective criteria. All of the British studies raised phage against specific pathogens then used to create experimental infections. Demonstrable efficacy against Escherichia, Acinetobacter, Pseudomonas and Staphylococcus spp. was noted in these model systems. Two U.S. papers dealt with improving the bioavailability of phage. Phage is sequestered in the spleen and removed from circulation. This can be overcome by serial passage of phage through mice to isolate mutants that resist sequestration. In conclusion, bacteriophages may show promise for treating antibiotic resistant pathogens. To facilitate further progress, directions for future research are discussed and a directory of authors from the reviewed

  17. Integration of agglutination assay for protein detection in microfluidic disc using Blu-ray optical pickup unit and optical fluid scanning

    DEFF Research Database (Denmark)

    Uddin, Rokon; Burger, Robert; Donolato, Marco

    2015-01-01

    We present a novel strategy for thrombin detection by combining a magnetic bead based agglutination assay and low-cost microfluidic disc. The detection method is based on an optomagnetic readout system implemented using a Blu-ray optical pickup unit (OPU) as main optoelectronic component. The ass...

  18. Bacteriophage lambda: The path from biology to theranostic agent.

    Science.gov (United States)

    Catalano, Carlos E

    2018-03-13

    Viral particles provide an attractive platform for the engineering of semisynthetic therapeutic nanoparticles. They can be modified both genetically and chemically in a defined manner to alter their surface characteristics, for targeting specific cell types, to improve their pharmacokinetic features and to attenuate (or enhance) their antigenicity. These advantages derive from a detailed understanding of virus biology, gleaned from decades of fundamental genetic, biochemical, and structural studies that have provided mechanistic insight into virus assembly pathways. In particular, bacteriophages offer significant advantages as nanoparticle platforms and several have been adapted toward the design and engineering of "designer" nanoparticles for therapeutic and diagnostic (theranostic) applications. The present review focuses on one such virus, bacteriophage lambda; I discuss the biology of lambda, the tools developed to faithfully recapitulate the lambda assembly reactions in vitro and the observations that have led to cooptation of the lambda system for nanoparticle design. This discussion illustrates how a fundamental understanding of virus assembly has allowed the rational design and construction of semisynthetic nanoparticles as potential theranostic agents and illustrates the concept of benchtop to bedside translational research. This article is categorized under: Biology-Inspired Nanomaterials> Protein and Virus-Based Structures Biology-Inspired Nanomaterials> Nucleic Acid-Based Structures. © 2018 Wiley Periodicals, Inc.

  19. Deletion mutations of bacteriophage

    International Nuclear Information System (INIS)

    Ryo, Yeikou

    1975-01-01

    Resolution of mutation mechanism with structural changes of DNA was discussed through the studies using bacteriophage lambda. One of deletion mutations inductions of phage lambda is the irradiation of ultraviolet ray. It is not clear if the inductions are caused by errors in reparation of ultraviolet-induced damage or by the activation of int gene. Because the effective site of int gene lies within the regions unnecessary for existing, it is considered that int gene is connected to deletion mutations induction. A certain system using prophage complementarity enables to detect deletion mutations at essential hereditary sites and to solve the relations of deletion mutations with other recombination system, DNA reproduction and repairment system. Duplication and multiplication of hereditary elements were discussed. If lambda deletion mutations of the system, which can control recombination, reproduction and repairment of added DNA, are constructed, mutations mechanism with great changes of DNA structure can be solved by phage lambda. (Ichikawa, K.)

  20. Efficient engineering of a bacteriophage genome using the type I-E CRISPR-Cas system.

    Science.gov (United States)

    Kiro, Ruth; Shitrit, Dror; Qimron, Udi

    2014-01-01

    The clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated (Cas) system has recently been used to engineer genomes of various organisms, but surprisingly, not those of bacteriophages (phages). Here we present a method to genetically engineer the Escherichia coli phage T7 using the type I-E CRISPR-Cas system. T7 phage genome is edited by homologous recombination with a DNA sequence flanked by sequences homologous to the desired location. Non-edited genomes are targeted by the CRISPR-Cas system, thus enabling isolation of the desired recombinant phages. This method broadens CRISPR Cas-based editing to phages and uses a CRISPR-Cas type other than type II. The method may be adjusted to genetically engineer any bacteriophage genome.

  1. A quorum-sensing-induced bacteriophage defense mechanism

    DEFF Research Database (Denmark)

    Høyland-Kroghsbo, Nina Molin; Mærkedahl, Rasmus Baadsgaard; Svenningsen, Sine

    2013-01-01

    of uninfected survivor cells after a potent attack by virulent phages. Notably, this mechanism may apply to a broader range of phages, as AHLs also reduce the risk of ¿ phage infection through a different receptor. IMPORTANCE To enable the successful manipulation of bacterial populations, a comprehensive...... sensing plays an important role in determining the susceptibility of E. coli to infection by bacteriophages ¿ and ¿. On the basis of our findings in the classical Escherichia coli-¿ model system, we suggest that quorum sensing may serve as a general strategy to protect bacteria specifically under...

  2. Hyperparasitism by the bacteriophage (Caudovirales) infecting Candidatus Xenohaliotis californiensis (Rickettsiales-like prokaryote) parasite of wild abalone Haliotis fulgens and Haliotis corrugata from the Peninsula of Baja California, Mexico.

    Science.gov (United States)

    Cruz-Flores, Roberto; Cáceres-Martínez, Jorge; Muñoz-Flores, Monserrat; Vásquez-Yeomans, Rebeca; Hernández Rodriguez, Mónica; Ángel Del Río-Portilla, Miguel; Rocha-Olivares, Axayácatl; Castro-Longoria, Ernestina

    2016-10-01

    Candidatus Xenohaliotis californiensis (CXc) is a Rickettsiales-like prokaryote that is considered the causal agent of Withering Syndrome (WS), a chronic disease of abalone, from the west coast of North America and it is listed by the International Organization for Animal Health (OIE) as a reportable agent due to its pathogenicity. This bacterium in red abalone Haliotis rufescens, black abalone Haliotis cracherodii, and yellow abalone Haliotis corrugata from California, US and Baja California, Mexico has been found to be infected by a bacteriophage. To date, there is no information on the epizootiology of CXc and its bacteriophage in natural populations of abalone; furthermore, it is unknown if the bacteriophage was also present in CXc infecting blue abalone Haliotis fulgens. The objective of this study was to determine the distribution, prevalence and intensity of CXc, as well as to determine the distribution and prevalence of the bacteriophage and to study interactions between host sex and hyperparasitism in blue abalone and yellow abalone. Tissue samples were obtained from seven localities where the commercial capture of wild abalone is carried out. Samplings were conducted throughout the 2012-2013 capture seasons and a total of 182 blue abalone and 170 yellow abalone were obtained. The prevalence and intensity of CXc and the prevalence of the bacteriophage were determined by histology. The identity of CXc was confirmed by PCR, product sequence analysis and in situ hybridization while the identity of the bacteriophage was corroborated by TEM. The prevalence of CXc infected and uninfected by the bacteriophage was 80% in blue abalone and 62% in yellow abalone. Low infection intensities were found in 86% of blue abalone and 82% of yellow abalone. Infection intensity was significantly higher in undifferentiated yellow abalone. The bacteriophage in CXc showed a prevalence of 22% and 31% in blue abalone and yellow abalone respectively. These results show that CXc and

  3. Decontamination of materials contaminated with Francisella philomiragia or MS2 bacteriophage using PES-Solid, a solid source of peracetic acid.

    Science.gov (United States)

    Buhr, T L; Young, A A; Johnson, C A; Minter, Z A; Wells, C M

    2014-08-01

    The aim of the study was to develop test methods and evaluate survival of Francisella philomiragia cells and MS2 bacteriophage after exposure to PES-Solid (a solid source of peracetic acid) formulations with or without surfactants. Francisella philomiragia cells (≥7·6 log10 CFU) or MS2 bacteriophage (≥6·8 log10 PFU) were deposited on seven different test materials and treated with three different PES-Solid formulations, three different preneutralized samples and filter controls at room temperature for 15 min. There were 0-1·3 log10 CFU (6 log10 CFU/PFU F. philomiragia cells and/or MS2 bacteriophage on different materials. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

  4. Solid phase radioimmunoassay for quantitation of IgM rheumatoid factor (RF). Comparison with agglutination techniques and radioimmunoprecipitation polyethylene glycol assay (RIPEGA)

    Energy Technology Data Exchange (ETDEWEB)

    Herrmann, D.; Jaeger, L.; Hein, G.; Henzgen, M.; Fiebig, H.; Schlenvoigt, G.; Vogelsang, H. (Friedrich-Schiller-Universitaet, Jena (German Democratic Republic). Bereich Medizin; Karl-Marx-Universitaet, Leipzig (German Democratic Republic). Sektion Biowissenschaften)

    1985-01-01

    A solid-phase radioimmunoassay capable of detecting nanogram quantities of human IgM rheumatoid factor using a monoclonal anti-..mu..-chain antibody is described. Human IgG did not interfere with the detection of IgM RF by this method. The small nonspecific binding of nonRF IgM to the human IgG coated tubes utilized in the assay must be corrected for by assaying samples in parallel bovine serum albumin coated control tubes only in cases of deviation of IgM from normal range. 69 coded and randomly arranged sera from patients with rheumatoid arthritis (RA), nonrheumatic joint diseases and healthy adult control subjects were investigated by this method, agglutination techniques as well as RIPEGA. A good correlation between solid-phase radioimmunoassay and agglutination techniques was found. Patients with seropositive RA had significantly higher concentrations of IgM RF than seronegative RA patients or control subjects.

  5. Shrimp hemocyte homeostasis-associated protein (PmHHAP) interacts with WSSV134 to control apoptosis in white spot syndrome virus infection.

    Science.gov (United States)

    Apitanyasai, Kantamas; Amparyup, Piti; Charoensapsri, Walaiporn; Sangsuriya, Pakkakul; Tassanakajon, Anchalee

    2018-05-01

    Hemocyte homeostasis-associated protein (PmHHAP) was first identified as a viral-responsive gene, due to a high upregulation in transcription following white spot syndrome virus (WSSV) infection. Functional studies using RNA interference have suggested that PmHHAP is involved in hemocyte homeostasis by controlling apoptosis during WSSV infection. In this study, the role of PmHHAP in host-viral interactions was further investigated. Yeast two-hybrid assay and co-immunoprecipitation revealed that PmHHAP binds to an anti-apoptosis protein, WSSV134. The viral protein WSSV134 is a late protein of WSSV, expressed 24 h post infection (hpi). Gene silencing of WSSV134 in WSSV-infected shrimp resulted in a reduction of the expression level of the viral replication marker genes VP28, wsv477, and ie-1, which suggests that WSSV134 is likely involved in viral propagation. However, co-silencing of PmHHAP and WSSV134 counteracted the effects on WSSV infection, which implies the importance of the host-pathogen interaction between PmHHAP and WSSV134 in WSSV infection. In addition, caspase 3/7 activity was noticeably induced in the PmHHAP and WSSV134 co-silenced shrimp upon WSSV infection. Moreover, PmHHAP and WSSV134 inhibited caspase-induced activation of PmCasp in vitro in a non-competitive manner. Taken together, these results suggest that PmHHAP and WSSV134 play a role in the host-pathogen interaction and work concordantly to control apoptosis in WSSV infection. Copyright © 2018 Elsevier Ltd. All rights reserved.

  6. Ammolagena clavata (Jones and Parker, 1860), an agglutinated benthic foraminiferal species - first report from the Recent sediments, Arabian Sea, Indian Ocean region

    Digital Repository Service at National Institute of Oceanography (India)

    Nigam, R.; Mazumder, A.; Saraswat, R.

    The rare presence of the agglutinated foraminiferal species Ammolagena clavata is presented for the first time from the Recent sediments of the Indian Ocean region. This species has previously been reported in Recent sediments from all other oceans...

  7. Phenotypic, fermentation characterization, and resistance mechanism analysis of bacteriophage-resistant mutants of Lactobacillus delbrueckii ssp. bulgaricus isolated from traditional Chinese dairy products.

    Science.gov (United States)

    Deng, Kaibo; Fang, Wei; Zheng, Baodong; Miao, Song; Huo, Guicheng

    2018-03-01

    Bacteriophage infection is a large factor in dairy industrial production failure on the basis of pure inoculation fermentation, and developing good commercial starter cultures from wild dairy products and improving the environmental vigor of starter cultures by enhancing their phage resistance are still the most effective solutions. Here we used a spontaneous isolation method to obtain bacteriophage-resistant mutants of Lactobacillus delbrueckii ssp. bulgaricus strains that are used in traditional Chinese fermented dairy products. We analyzed their phenotypes, fermentation characteristics, and resistance mechanisms. The results showed that bacteriophage-insensitive mutants (BIM) BIM8 and BIM12 had high bacteriophage resistance while exhibiting fermentation and coagulation attributes that were as satisfying as those of their respective parent strains KLDS1.1016 and KLDS1.1028. According to the attachment receptor detection, mutants BIM8 and BIM12 exhibited reduced absorption to bacteriophage phiLdb compared with their respective bacteriophage-sensitive parent strains because of changes to the polysaccharides or teichoic acids connected to their peptidoglycan layer. Additionally, genes, including HSDR, HSDM, and HSDS, encoding 3 subunits of a type I restriction-modification system were identified in their respective parent strains. We also discovered that HSDR and HSDM were highly conserved but that HSDS was variable because it is responsible for the DNA specificity of the complex. The late lysis that occurred only in strain KLDS1.1016 and not in strain KLDS1.1028 suggests that the former and its mutant BIM8 also may have an activatable restriction-modification mechanism. We conclude that the L. bulgaricus BIM8 and BIM12 mutants have great potential in the dairy industry as starter cultures, and their phage-resistance mechanism was effective mainly due to the adsorption interference and restriction-modification system. Copyright © 2018 American Dairy Science

  8. Structural changes of bacteriophage phi29 upon DNA packaging and release.

    Science.gov (United States)

    Xiang, Ye; Morais, Marc C; Battisti, Anthony J; Grimes, Shelley; Jardine, Paul J; Anderson, Dwight L; Rossmann, Michael G

    2006-11-01

    Cryo-electron microscopy three-dimensional reconstructions have been made of mature and of emptied bacteriophage phi29 particles without making symmetry assumptions. Comparisons of these structures with each other and with the phi29 prohead indicate how conformational changes might initiate successive steps of assembly and infection. The 12 adsorption capable 'appendages' were found to have a structure homologous to the bacteriophage P22 tailspikes. Two of the appendages are extended radially outwards, away from the long axis of the virus, whereas the others are around and parallel to the phage axis. The appendage orientations are correlated with the symmetry-mismatched positions of the five-fold related head fibers, suggesting a mechanism for partial cell wall digestion upon rotation of the head about the tail when initiating infection. The narrow end of the head-tail connector is expanded in the mature virus. Gene product 3, bound to the 5' ends of the genome, appears to be positioned within the expanded connector, which may potentiate the release of DNA-packaging machine components, creating a binding site for attachment of the tail.

  9. The Protein Interaction Network of Bacteriophage Lambda with Its Host, Escherichia coli

    Science.gov (United States)

    Blasche, Sonja; Wuchty, Stefan; Rajagopala, Seesandra V.

    2013-01-01

    Although most of the 73 open reading frames (ORFs) in bacteriophage λ have been investigated intensively, the function of many genes in host-phage interactions remains poorly understood. Using yeast two-hybrid screens of all lambda ORFs for interactions with its host Escherichia coli, we determined a raw data set of 631 host-phage interactions resulting in a set of 62 high-confidence interactions after multiple rounds of retesting. These links suggest novel regulatory interactions between the E. coli transcriptional network and lambda proteins. Targeted host proteins and genes required for lambda infection are enriched among highly connected proteins, suggesting that bacteriophages resemble interaction patterns of human viruses. Lambda tail proteins interact with both bacterial fimbrial proteins and E. coli proteins homologous to other phage proteins. Lambda appears to dramatically differ from other phages, such as T7, because of its unusually large number of modified and processed proteins, which reduces the number of host-virus interactions detectable by yeast two-hybrid screens. PMID:24049175

  10. M13 Bacteriophage/Silver Nanowire Surface-Enhanced Raman Scattering Sensor for Sensitive and Selective Pesticide Detection.

    Science.gov (United States)

    Koh, Eun Hye; Mun, ChaeWon; Kim, ChunTae; Park, Sung-Gyu; Choi, Eun Jung; Kim, Sun Ho; Dang, Jaejeung; Choo, Jaebum; Oh, Jin-Woo; Kim, Dong-Ho; Jung, Ho Sang

    2018-03-28

    A surface-enhanced Raman scattering (SERS) sensor comprising silver nanowires (AgNWs) and genetically engineered M13 bacteriophages expressing a tryptophan-histidine-tryptophan (WHW) peptide sequence (BPWHW) was fabricated by simple mixing of BPWHW and AgNW solutions, followed by vacuum filtration onto a glass-fiber filter paper (GFFP) membrane. The AgNWs stacked on the GFFP formed a high density of SERS-active hot spots at the points of nanowire intersections, and the surface-coated BPWHW functioned as a bioreceptor for selective pesticide detection. The BPWHW-functionalized AgNW (BPWHW/AgNW) sensor was characterized by scanning electron microscopy, confocal scanning fluorescence microscopy, atomic force microscopy, and Fourier transform infrared spectroscopy. The Raman signal enhancement and the selective pesticide SERS detection properties of the BPWHW/AgNW sensor were investigated in the presence of control substrates such as wild-type M13 bacteriophage-decorated AgNWs (BPWT/AgNW) and undecorated AgNWs (AgNW). The BPWHW/AgNW sensor exhibited a significantly higher capture capability for pesticides, especially paraquat (PQ), than the control SERS substrates, and it also showed a relatively higher selectivity for PQ than for other bipyridylium pesticides such as diquat and difenzoquat. Furthermore, as a field application test, PQ was detected on the surface of PQ-pretreated apple peels, and the results demonstrated the feasibility of using a paper-based SERS substrate for on-site residual pesticide detection. The developed M13 bacteriophage-functionalized AgNW SERS sensor might be applicable for the detection of various pesticides and chemicals through modification of the M13 bacteriophage surface peptide sequence.

  11. Isolation and characterization of Enterobacteriaceae species infesting post-harvest strawberries and their biological control using bacteriophages.

    Science.gov (United States)

    Kurtböke, D Ipek; Palk, A; Marker, A; Neuman, C; Moss, L; Streeter, K; Katouli, M

    2016-10-01

    Strawberry is a significantly consumed fruit worldwide, mostly without being subjected to disinfection processes. During the harvest and transfer from farm to consumers as well as where organic farming practises have been employed, the surface of the fruit may become contaminated by pathogenic bacteria. Post-harvest strawberry fruits in punnets available for public consumption were thus screened for the presence of enteric bacteria in the Sunshine Coast region of Queensland, Australia. Some of the tested samples (13 %) were found to carry such bacteria and even in greater numbers if organic amendments were used (69 %). The bacteria were found to belong in the genera of Escherichia, Enterobacter, Raoultella, Klebsiella, Pantoea, Shigella, Citrobacter and Cronobacter within the family Enterobacteriaceae. Some of the isolates were found to adhere to Caco-2 cells representing human gut epithelium as well as carrying virulence and toxin genes. Resistance mostly against sulphafurazole, cefoxitin, ampicillin and nitrofurantoin was found among 14 different antimicrobial agents tested including 100 % resistance to cefoxitin and ampicillin in the genus Pantoea. In the second phase of the study, bacteriophages were isolated against the isolates and were subsequently applied to post-harvest fruits. A significant (P ≤ 0.001) reduction in the number of enteric bacteria was observed when a high-titre polyvalent bacteriophage suspension (×10(12) PFU/mL) was applied to the fruit surface. Bacteriophages also decreased the adhesion of the Escherichia coli isolates to Caco-2 cells. Findings might indicate that biological control using bacteriophages might be of significant value for the industry targeting to reduce pathogenic loads of bacteria on the fruit.

  12. Ex vivo and in vivo evaluation of microemulsion based transdermal delivery of E. coli specific T4 bacteriophage: A rationale approach to treat bacterial infection.

    Science.gov (United States)

    Rastogi, Vaibhav; Yadav, Pragya; Verma, Anurag; Pandit, Jayanta K

    2017-09-30

    This study is focused on the development and evaluation of transdermal delivery of E. coli-specific T4 bacteriophages both ex-vivo and in-vivo using microemulsion as delivery carrier in eradicating the infection caused by E. coli. Microemulsions were prepared by mixing selected oil, surfactants and aqueous phase containing bacteriophages. The formulations were subjected to physicochemical characterization, ex-vivo and in-vivo permeation, stability studies, histological and immunofluorescence examination. The colloidal system exhibits a uniform size distribution, of finite size (150-320nm). Transmission electron microscopy revealed the encapsulation of bacteriophage in the aqueous globule. Ex-vivo permeation across skin was successfully achieved as 6×10 6 PFU/mL and 6.7×10 6 PFU/mL of T4 permeated from ME 6% and 10%, respectively. ME 6% was found to be thermodynamically stable and in-vivo permeation resulted in 5.49×10 5 PFU/mL of bacteriophages in the blood of the E. coli challenged rats, while 2.48×10 5 PFU/mL was detected in germ free rats, at the end of the study. Infected rats that were treated with bacteriophage were survived while significant mortality was observed in others. Histological and IL-6 immunofluorescence examination of the tissues revealed the efficacy/safety of the therapy. The microemulsion-based transdermal delivery of bacteriophage could be a promising approach to treat the infections caused by antibiotic-resistant bacteria. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. An influenza A virus agglutination test using antibody-like polymers.

    Science.gov (United States)

    Sukjee, Wannisa; Thitithanyanont, Arunee; Wiboon-Ut, Suwimon; Lieberzeit, Peter A; Paul Gleeson, M; Navakul, Krongkaew; Sangma, Chak

    2017-10-01

    Antibodies are commonly used in diagnostic routines to identify pathogens. The testing protocols are relatively simple, requiring a certain amount of a specific antibody to detect its corresponding pathogen. Antibody functionality can be mimicked by synthesizing molecularly imprinted polymers (MIPs), i.e. polymers that can selectively recognize a given template structure. Thus, MIPs are sometimes termed 'plastic antibody (PA)'. In this study, we have synthesized new granular MIPs using influenza A virus templates by precipitation polymerization. The selective binding of influenza A to the MIP particles was assessed and subsequently contrasted with other viruses. The affinities of influenza A virus towards the MIP was estimated based on an agglutination test by measuring the amount of influenza subtypes absorbed onto the MIPs. The MIPs produced using the H1N1 template showed specific reactivity to H1N1 while those produced using H5N1 and H3N2 templates showed cross-reactivity.

  14. Use of commercial extenders and alternatives to prevent sperm agglutination for cryopreservation of brown bear semen.

    Science.gov (United States)

    Gomes-Alves, S; Alvarez, M; Nicolas, M; Lopez-Urueña, E; Martínez-Rodríguez, C; Borragan, S; de Paz, P; Anel, L

    2014-08-01

    The objective of this study was to evaluate different bovine and canine commercial semen extenders for cryopreservation of brown bear ejaculates and the effect of semen collection directly into extender on sperm agglutination. Semen samples were obtained by electroejaculation from 13 adult males. In experiment 1, eleven ejaculates from eight bears were used to evaluate Bioxcell and Andromed as extenders, whereas in experiment 2, nine ejaculates from six bears were used to evaluate Triladyl canine, CaniPro, and Extender 2 as extenders. An extender specifically developed for brown bears (Test-Tris-fructose-egg yolk-glycerol, TTF-ULE/bear) served as a control extender in both experiments. After thawing, total and progressive sperm motility and sperm viability were greater (P bear and Andromed extenders than for Bioxcell in experiment 1 and greater (P bear extender than for Triladyl Canine, CaniPro, and Extender 2 in experiment 2. In experiment 3, addition of handling extender (TTF-H) to the semen collection tube for eight ejaculates from seven bears resulted in less (P bear is the most suitable extender for brown bear semen cryopreservation, but comparable results can be obtained with the commercial extender Andromed. In addition, collection of ejaculates directly in TTF-H extender decreases sperm agglutination in fresh samples. Copyright © 2014 Elsevier Inc. All rights reserved.

  15. A highly abundant bacteriophage discovered in the unknown sequences of human faecal metagenomes

    NARCIS (Netherlands)

    Dutilh, Bas E; Cassman, Noriko; McNair, Katelyn; Sanchez, Savannah E; Silva, Genivaldo G Z; Boling, Lance; Barr, Jeremy J; Speth, Daan R; Seguritan, Victor; Aziz, Ramy K; Felts, Ben; Dinsdale, Elizabeth A; Mokili, John L; Edwards, Robert A

    2014-01-01

    Metagenomics, or sequencing of the genetic material from a complete microbial community, is a promising tool to discover novel microbes and viruses. Viral metagenomes typically contain many unknown sequences. Here we describe the discovery of a previously unidentified bacteriophage present in the

  16. Re-initiation repair in bacteriophage T4

    International Nuclear Information System (INIS)

    Cupido, M.

    1981-01-01

    Irradiation of bacteriophage T4 with ultraviolet light induces the formation of pyrimidine dimers in its DNA. These dimers hamper replication of DNA and, to a lesser extent, transcription of DNA after its infection of bacteria. A number of pathways enable phage T4 to multiply dimer-containing DNA. One of these pathways has been named replication repair and is described in this thesis. The properties of two phage strains, unable to perform replication repair, have been studied to obtain a picture of the repair process. The mutations in these strains that affect replication repair have been located on the genomic map of T4. (Auth.)

  17. The inactivating and mutagenic effect of hydroxylamine on bacteriophage φX174

    NARCIS (Netherlands)

    Pol, J.H. van de; Arkel, G.A. van

    1965-01-01

    The inactivation of bacteriophage ΦXI74 by the mutagenic agents nitrous acid and ultraviolet irradiation proceeds according to a single-hit kinetics. However, treatment of purified ΦXI74 by hydroxylamine (HA) at pH 6 and 25° results in an inactivation that is not strictly exponential. The

  18. Activity of Bacteriophages in Removing Biofilms of Pseudomonas aeruginosa Isolates from Chronic Rhinosinusitis Patients

    NARCIS (Netherlands)

    Fong, Stephanie A.; Drilling, Amanda; Morales, Sandra; Cornet, Marjolein E.; Woodworth, Bradford A.; Fokkens, Wytske J.; Psaltis, Alkis J.; Vreugde, Sarah; Wormald, Peter-John

    2017-01-01

    Introduction:Pseudomonas aeruginosa infections are prevalent amongst chronic rhinosinusitis (CRS) sufferers. Many P. aeruginosa strains form biofilms, leading to treatment failure. Lytic bacteriophages (phages) are viruses that infect, replicate within, and lyse bacteria, causing bacterial death.

  19. Discriminating the hemolytic risk of blood type A plasmas using the complement hemolysis using human erythrocytes (CHUHE) assay.

    Science.gov (United States)

    Cunnion, Kenji M; Hair, Pamela S; Krishna, Neel K; Sass, Megan A; Enos, Clinton W; Whitley, Pamela H; Maes, Lanne Y; Goldberg, Corinne L

    2017-03-01

    The agglutination-based cross-matching method is sensitive for antibody binding to red blood cells but is only partially predictive of complement-mediated hemolysis, which is important in many acute hemolytic transfusion reactions. Here, we describe complement hemolysis using human erythrocytes (CHUHE) assays that directly evaluate complement-mediated hemolysis between individual serum-plasma and red blood cell combinations. The CHUHE assay is used to evaluate correlations between agglutination titers and complement-mediated hemolysis as well as the hemolytic potential of plasma from type A blood donors. Plasma or serum from each type A blood donor was incubated with AB or B red blood cells in the CHUHE assay and measured for free hemoglobin release. CHUHE assays for serum or plasma demonstrate a wide, dynamic range and high sensitivity for complement-mediated hemolysis for individual serum/plasma and red blood cell combinations. CHUHE results suggest that agglutination assays alone are only moderately predictive of complement-mediated hemolysis. CHUHE results also suggest that plasma from particular type A blood donors produce minimal complement-mediated hemolysis, whereas plasma from other type A blood donors produce moderate to high-level complement-mediated hemolysis, depending on the red blood cell donor. The current results indicate that the CHUHE assay can be used to assess complement-mediated hemolysis for plasma or serum from a type A blood donor, providing additional risk discrimination over agglutination titers alone. © 2016 AABB.

  20. Comparison of Brucella immunoglobulin M and G flow assays with serum agglutination and 2-mercaptoethanol tests in the diagnosis of brucellosis

    NARCIS (Netherlands)

    Zeytinoğlu, Ayşin; Turhan, Ajda; Altuğlu, Imre; Bilgiç, Altinay; Abdoel, Theresia H.; Smits, Henk L.

    2006-01-01

    The diagnostic value of Brucella IgM/IgG flow assays was evaluated in comparison with serum agglutination and 2-mercaptoethanol tests by testing a selection of serum samples submitted to the laboratory because of clinical suspicion of brucellosis. All 39 admission and 11 follow-up samples that

  1. On a grain of sand - a microhabitat for the opportunistic agglutinated foraminifera Hemisphaerammina apta n. sp., from the early Eocene Arctic Ocean

    Science.gov (United States)

    McNeil, David H.; Neville, Lisa A.

    2018-02-01

    Hemisphaerammina apta n. sp. is an attached monothalamous agglutinated foraminifera discovered in shelf sediments of the early Eocene Arctic Ocean. It is a simple yet distinctive component of the endemic agglutinated foraminiferal assemblage that colonized the Arctic Ocean after the microfaunal turnover caused by the Paleocene-Eocene Thermal Maximum. Associated foraminifera are characterized by a high percentage of monothalamous species (up to 60 %) and are entirely agglutinated indicating a brackish (mesohaline) early Eocene Arctic Ocean. Hemisphaerammina apta occurs exclusively as individuals attached to fine detrital grains (0.2 to 1.8 mm) of sediment. It is a small species (0.06 to 0.2 mm in diameter), fine-grained, with a low hemispherical profile, no floor across the attachment area, no substantive marginal flange, no internal structures, and no aperture. Lacking an aperture, it apparently propagated and fed through minute (micrometre-sized) interstitial pores in the test wall. Attachment surfaces vary from concave to convex and rough to smooth. Grains for attachment are diverse in shape and type but are predominantly of quartz and chert. The presence of H. apta in the early Eocene was an opportunistic response to an environment with an active hydrological system (storm events). Attachment to grains of sand would provide a more stable base on a sea floor winnowed by storm-generated currents. Active transport is indicated by the relative abundance of reworked foraminifera mixed with in situ species. Contemporaneous reworking and colonization by H. apta is suggested by its attachment to a reworked specimen of Cretaceous foraminifera.

  2. Prevalence of agglutinating antibodies to Sarcocystis neurona in skunks (Mephitis Mephitis), raccoons (Procyon lotor), and opossums (Didelphis Virginiana) from Connecticut.

    Science.gov (United States)

    Mitchell, Sheila M; Richardson, Dennis J; Cheadle, M Andy; Zajac, Anne M; Lindsay, David S

    2002-10-01

    Equine protozoal myeloencephalitis is the most important protozoan disease of horses in North America and is usually caused by Sarcocystis neurona. Natural cases of encephalitis caused by S. neurona have been reported in skunks (Mephitis mephitis) and raccoons (Procyon lotor). Opossums (Didelphis spp.) are the only known definitive host. Sera from 24 striped skunks, 12 raccoons, and 7 opossums (D. virginiana) from Connecticut were examined for agglutinating antibodies to S. neurona using the S. neurona agglutination test (SAT) employing formalin-fixed merozoites as antigen. The SAT was validated for skunk sera using pre- and postinfection serum samples from 2 experimentally infected skunks. Of the 24 (46%) skunks 11 were positive, and all 12 raccoons were positive for S. neurona antibodies. None of the 7 opossums was positive for antibodies to S. neurona. These results suggest that exposure to sporocysts of S. neurona by intermediate hosts is high in Connecticut. The absence of antibodies in opossums collected from the same areas is most likely because of the absence of systemic infection in the definitive host.

  3. Comparison of the virucidal efficacy of peracetic acid, potassium monopersulphate and sodium hypochlorite on bacteriophages P001 and MS2.

    Science.gov (United States)

    Morin, T; Martin, H; Soumet, C; Fresnel, R; Lamaudière, S; Le Sauvage, A L; Deleurme, K; Maris, P

    2015-09-01

    The phagicidal activity of peroxy products against the virulent bacteriophage P001 infecting lactic acid bacteria and bacteriophage MS2 used as a surrogate of enteric viruses (EVs) was evaluated and compared to sodium hypochlorite using the EN 13610 European suspension test and a surface test developed in our laboratories. Infectivity tests were adapted and/or developed to determine the activity of disinfectants against reference P001 phage of Lactoccocus lactis and F-specific RNA phage MS2 of Escherichia coli in conditions simulating practical use. Similar concentrations of sodium hypochlorite were phagicidal against both bacteriophages, either at 0·05-0·125% of active chlorine using the suspension test or at 0·12-0·5% using the surface test. For Potassium monopersulphate (MPS), phagicidal concentrations varied from 0·006 to 0·012% whatever the type of test and phages. However, for peracetic acid products (PAP) used in suspension, concentrations 55 times higher were necessary against MS2 (0·271%) than against P001 (0·005%). With the surface test, 0·089-0·178% concentrations of PAP were effective against MS2, but these concentrations were 16-32 times greater than needed against P001. Sodium hypochlorite and MPS had similar phagicidal activities against P001 and MS2, but PAP did not. This is the first comparative study to investigate through suspension and surface tests the difference in resistance to peroxy compounds between a reference bacteriophage (P001) used to evaluate phagicidal concentrations in European standards and a surrogate of EVs (MS2). Results underline the importance of validation tests on pertinent surrogates of viruses or bacteriophages to adjust the concentration of disinfectants for use in the food and water industries. © 2015 The Society for Applied Microbiology.

  4. Bacteriophage T7 structure according to the data of small-angle X-ray scattering

    Energy Technology Data Exchange (ETDEWEB)

    Rol' bin, Yu A; Svergun, D I; Feigin, L A; Gashpar, Sh; Ronto, D [AN SSSR, Moscow. Inst. Kristallografii

    1980-01-01

    An attempt is made to obtain complete data on the form, sizes, weight and hydration of the T7 bacteriophage cultivated on E.coli cells and the peculiarities of phage DNA structure using the method of small-angle scattering.

  5. Rapid and sensitive detection of Yersinia pestis using amplification of plague diagnostic bacteriophages monitored by real-time PCR.

    Directory of Open Access Journals (Sweden)

    Kirill V Sergueev

    Full Text Available BACKGROUND: Yersinia pestis, the agent of plague, has caused many millions of human deaths and still poses a serious threat to global public health. Timely and reliable detection of such a dangerous pathogen is of critical importance. Lysis by specific bacteriophages remains an essential method of Y. pestis detection and plague diagnostics. METHODOLOGY/PRINCIPAL FINDINGS: The objective of this work was to develop an alternative to conventional phage lysis tests--a rapid and highly sensitive method of indirect detection of live Y. pestis cells based on quantitative real-time PCR (qPCR monitoring of amplification of reporter Y. pestis-specific bacteriophages. Plague diagnostic phages phiA1122 and L-413C were shown to be highly effective diagnostic tools for the detection and identification of Y. pestis by using qPCR with primers specific for phage DNA. The template DNA extraction step that usually precedes qPCR was omitted. phiA1122-specific qPCR enabled the detection of an initial bacterial concentration of 10(3 CFU/ml (equivalent to as few as one Y. pestis cell per 1-microl sample in four hours. L-413C-mediated detection of Y. pestis was less sensitive (up to 100 bacteria per sample but more specific, and thus we propose parallel qPCR for the two phages as a rapid and reliable method of Y. pestis identification. Importantly, phiA1122 propagated in simulated clinical blood specimens containing EDTA and its titer rise was detected by both a standard plating test and qPCR. CONCLUSIONS/SIGNIFICANCE: Thus, we developed a novel assay for detection and identification of Y. pestis using amplification of specific phages monitored by qPCR. The method is simple, rapid, highly sensitive, and specific and allows the detection of only live bacteria.

  6. Effective inhibition of lytic development of bacteriophages λ, P1 and T4 by starvation of their host, Escherichia coli

    Directory of Open Access Journals (Sweden)

    Węgrzyn Alicja

    2007-02-01

    Full Text Available Abstract Background Bacteriophage infections of bacterial cultures cause serious problems in genetic engineering and biotechnology. They are dangerous not only because of direct effects on the currently infected cultures, i.e. their devastation, but also due to a high probability of spreading the phage progeny throughout a whole laboratory or plant, which causes a real danger for further cultivations. Therefore, a simple method for quick inhibition of phage development after detection of bacterial culture infection should be very useful. Results Here, we demonstrate that depletion of a carbon source from the culture medium, which provokes starvation of bacterial cells, results in rapid inhibition of lytic development of three Escherichia coli phages, λ, P1 and T4. Since the effect was similar for three different phages, it seems that it may be a general phenomenon. Moreover, similar effects were observed in flask cultures and in chemostats. Conclusion Bacteriophage lytic development can be inhibited efficiently by carbon source limitation in bacterial cultures. Thus, if bacteriophage contamination is detected, starvation procedures may be recommended to alleviate deleterious effects of phage infection on the culture. We believe that this strategy, in combination with the use of automated and sensitive bacteriophage biosensors, may be employed in the fermentation laboratory practice to control phage outbreaks in bioprocesses more effectively.

  7. Role of bacteriophages in STEC infections: new implications for the design of prophylactic and treatment approaches [v2; ref status: indexed, http://f1000r.es/437

    Directory of Open Access Journals (Sweden)

    Jaime H. Amorim

    2014-08-01

    Full Text Available Shiga toxin (Stx is considered the main virulence factor in Shiga toxin-producing Escherichia coli (STEC infections. Previously we reported the expression of biologically active Stx by eukaryotic cells in vitro and in vivo following transfection with plasmids encoding Stx under control of the native bacterial promoter1,2. Since stx genes are present in the genome of lysogenic bacteriophages, here we evaluated the relevance of bacteriophages during STEC infection. We used the non-pathogenic E. coli C600 strain carrying a lysogenic 933W mutant bacteriophage in which the stx operon was replaced by a gene encoding the green fluorescent protein (GFP. Tracking GFP expression using an In Vivo Imaging System (IVIS, we detected fluorescence in liver, kidney, and intestine of mice infected with the recombinant E. coli strain after treatment with ciprofloxacin, which induces the lytic replication and release of bacteriophages. In addition, we showed that chitosan, a linear polysaccharide composed of d-glucosamine residues and with a number of commercial and biomedical uses, had strong anti-bacteriophage effects, as demonstrated at in vitro and in vivo conditions. These findings bring promising perspectives for the prevention and treatment of haemolytic uremic syndrome (HUS cases.

  8. Bacteriophages use hypermodified nucleosides to evade host's defence systems

    DEFF Research Database (Denmark)

    Kot, Witold; Olsen, Nikoline S.; Carstens, Alexander Byth

    developed several strategies to evade these defence mechanisms. Ultimately, this led to the oldest and still running arms race - microorganisms vs. their molecular parasites. We here describe a remarkable new strategy used by the recently isolated Escherichia coli phage CAjan belonging to...... to investigate this mechanism in detail we have used several methods including direct plaque sequencing, restriction endonuclease analysis and CRISPR-Cas genome editing. Through generation of specific mutants, we were able to introduce a restriction sensitive phenotype in the CAjan bacteriophage providing new...

  9. Different expression patterns of genes from the exo-xis region of bacteriophage λ and Shiga toxin-converting bacteriophage Ф24B following infection or prophage induction in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Sylwia Bloch

    Full Text Available Lambdoid bacteriophages serve as useful models in microbiological and molecular studies on basic biological process. Moreover, this family of viruses plays an important role in pathogenesis of enterohemorrhagic Escherichia coli (EHEC strains, as they are carriers of genes coding for Shiga toxins. Efficient expression of these genes requires lambdoid prophage induction and multiplication of the phage genome. Therefore, understanding the mechanisms regulating these processes appears essential for both basic knowledge and potential anti-EHEC applications. The exo-xis region, present in genomes of lambdoid bacteriophages, contains highly conserved genes of largely unknown functions. Recent report indicated that the Ea8.5 protein, encoded in this region, contains a newly discovered fused homeodomain/zinc-finger fold, suggesting its plausible regulatory role. Moreover, subsequent studies demonstrated that overexpression of the exo-xis region from a multicopy plasmid resulted in impaired lysogenization of E. coli and more effective induction of λ and Ф24B prophages. In this report, we demonstrate that after prophage induction, the increase in phage DNA content in the host cells is more efficient in E. coli bearing additional copies of the exo-xis region, while survival rate of such bacteria is lower, which corroborated previous observations. Importantly, by using quantitative real-time reverse transcription PCR, we have determined patterns of expressions of particular genes from this region. Unexpectedly, in both phages λ and Ф24B, these patterns were significantly different not only between conditions of the host cells infection by bacteriophages and prophage induction, but also between induction of prophages with various agents (mitomycin C and hydrogen peroxide. This may shed a new light on our understanding of regulation of lambdoid phage development, depending on the mode of lytic cycle initiation.

  10. The cell-mediated immunity of Drosophila melanogaster: hemocyte lineages, immune compartments, microanatomy and regulation.

    Science.gov (United States)

    Honti, Viktor; Csordás, Gábor; Kurucz, Éva; Márkus, Róbert; Andó, István

    2014-01-01

    In the animal kingdom, innate immunity is the first line of defense against invading pathogens. The dangers of microbial and parasitic attacks are countered by similar mechanisms, involving the prototypes of the cell-mediated immune responses, the phagocytosis and encapsulation. Work on Drosophila has played an important role in promoting an understanding of the basic mechanisms of phylogenetically conserved modules of innate immunity. The aim of this review is to survey the developments in the identification and functional definition of immune cell types and the immunological compartments of Drosophila melanogaster. We focus on the molecular and developmental aspects of the blood cell types and compartments, as well as the dynamics of blood cell development and the immune response. Further advances in the characterization of the innate immune mechanisms in Drosophila will provide basic clues to the understanding of the importance of the evolutionary conserved mechanisms of innate immune defenses in the animal kingdom. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. LEPINOCONUS CHIOCCHINII GEN. N., N. SP., A CONICAL AGGLUTINATED FORAMINIFERA FROM THE UPPER CRETACEOUS OF ITALY

    Directory of Open Access Journals (Sweden)

    ERZIKA CRUZ-ABAD

    2017-04-01

    Full Text Available A new conical agglutinated foraminifer, Lepinoconus chiocchinii gen n., n. sp. from the lower Campanian shallow-water platform deposits of the Lepini Mountains (central Apennines, Italy, is described. It has a pseudo-keriothecal wall structure, uniserial arrangement of the adult chambers and multiple apertures. The exoskeleton is constituted by beams (main and intercalary continuous from one chamber to the next, while the endoskeleton bears pillars. The new taxon is included in the Coskinolinidae family. Lepinoconus chiocchinii gen. n., n. sp. is known from southern Italy, Greece and Albania.

  12. Effects of Sample Impurities on the Analysis of MS2 Bacteriophage by Small-Angle Neutron Scattering

    National Research Council Canada - National Science Library

    Elashvili, Ilya; Wick, Charles H; Kuzmanovic, Deborah A; Krueger, Susan; O'Connell, Catherine

    2005-01-01

    .... The impact of small molecular weight impurities of the resolution of structural data obtained by SANS of the bacteriophage MS2 distorts the resolution and sharpness of contrast variation peaks...

  13. Targeting Antibacterial Agents by Using Drug-Carrying Filamentous Bacteriophages

    Science.gov (United States)

    Yacoby, Iftach; Shamis, Marina; Bar, Hagit; Shabat, Doron; Benhar, Itai

    2006-01-01

    Bacteriophages have been used for more than a century for (unconventional) therapy of bacterial infections, for half a century as tools in genetic research, for 2 decades as tools for discovery of specific target-binding proteins, and for nearly a decade as tools for vaccination or as gene delivery vehicles. Here we present a novel application of filamentous bacteriophages (phages) as targeted drug carriers for the eradication of (pathogenic) bacteria. The phages are genetically modified to display a targeting moiety on their surface and are used to deliver a large payload of a cytotoxic drug to the target bacteria. The drug is linked to the phages by means of chemical conjugation through a labile linker subject to controlled release. In the conjugated state, the drug is in fact a prodrug devoid of cytotoxic activity and is activated following its dissociation from the phage at the target site in a temporally and spatially controlled manner. Our model target was Staphylococcus aureus, and the model drug was the antibiotic chloramphenicol. We demonstrated the potential of using filamentous phages as universal drug carriers for targetable cells involved in disease. Our approach replaces the selectivity of the drug itself with target selectivity borne by the targeting moiety, which may allow the reintroduction of nonspecific drugs that have thus far been excluded from antibacterial use (because of toxicity or low selectivity). Reintroduction of such drugs into the arsenal of useful tools may help to combat emerging bacterial antibiotic resistance. PMID:16723570

  14. Bacteriophage: laboratorial diagnosis and phage therapy Bacteriofagos: diagnóstico laboratorial e terapia fágica

    Directory of Open Access Journals (Sweden)

    Joas L. Da Silva

    2009-09-01

    Full Text Available Bacteriophages have been researched as a new alternative to antibiotics. These viruses inject their genetic material into bacteria and use their host machinery to multiply themselves. The research of bacteriophages in Brazil will certainly provide low-cost treatment of multidrug resistant bacteria, new microbiological diagnosis and advantages for the Brazilian food industry.Bacteriófagos têm sido pesquisados como uma alternativa ao uso de antibióticos. Estes vírus infectam as bactérias e utilizam a maquinaria celular para multiplicar o próprio material genético. O estudo de bacteriófagos no Brasil levará ao desenvolvimento de tratamentos de baixo custo, novos testes diagnósticos e vantagens para a industria alimentícia.

  15. Regions of incompatibility in single-stranded DNA bacteriophages phi X174 and G4

    NARCIS (Netherlands)

    van der Avoort, H. G.; van der Ende, A.; van Arkel, G. A.; Weisbeek, P. J.

    1984-01-01

    The intracellular presence of a recombinant plasmid containing the intercistronic region between the genes H and A of bacteriophage phi X174 strongly inhibits the conversion of infecting single-stranded phi X DNA to parental replicative-form DNA. Also, transfection with single-stranded or

  16. The membrane-bound form of gene 9 minor coat protein of bacteriophage M13

    NARCIS (Netherlands)

    Houbiers, M.C.

    2002-01-01

    Bacteriophage M13 is a virus that infects the bacteria Escherichia coli ( E. coli ), a single cell organism that resides in our intestines. It consists of the cytoplasm (contents) and a double membrane that keeps the

  17. Lectin-induced agglutination method of urinary exosomes isolation followed by mi-RNA analysis: Application for prostate cancer diagnostic.

    Science.gov (United States)

    Samsonov, Roman; Shtam, Tatiana; Burdakov, Vladimir; Glotov, Andrey; Tsyrlina, Evgenia; Berstein, Lev; Nosov, Alexander; Evtushenko, Vladimir; Filatov, Michael; Malek, Anastasia

    2016-01-01

    Prostate cancer is the most common cancer in men. Prostate-specific antigen has, however, insufficient diagnostic specificity. Novel complementary diagnostic approaches are greatly needed. MiRNAs are small regulatory RNAs which play an important role in tumorogenesis and are being investigated as a cancer biomarker. In addition to their intracellular regulatory functions, miRNAs are secreted into the extracellular space and can be found in various body fluids, including urine. The stability of extracellular miRNAs is defined by association with proteins, lipoprotein particles, and membrane vesicles. Among the known forms of miRNA packaging, tumour-derived exosome-enclosed miRNAs is thought to reflect the vital activity of cancer cells. The assessment of the exosomal fraction of urinary miRNA may present a new and highly specific method for prostate cancer diagnostics; however, this is challenged by the absence of reliable and inexpensive methods for isolation of exosomes. Prostate cancer (PC) cell lines and urine samples collected from 35 PC patients and 35 healthy donors were used in the study. Lectins, phytohemagglutinin, and concanavalin A were used to induce agglutination of exosomes. The efficiency of isolation process was evaluated by AFM and DLS assays. The protein content of isolated exosomes was analysed by western blotting. Exosomal RNA was assayed by automated electrophoresis and expression level of selected miRNAs was evaluated by RT-qPCR. The diagnostic potency of the urinary exosomal miRNA assessment was estimated by the ROC method. The formation of multi-vesicular agglutinates in urine can be induced by incubation with lectin at a final concentration of 2 mg/ml. These agglutinates contain urinary exosomes and may be pelleted by centrifugation with a relatively low G-force. The analysis of PC-related miRNA in urinary exosomes revealed significant up-regulation of miR-574-3p, miR-141-5p, and miR-21-5p associated with PC. Lectin-induced aggregation is a

  18. A resolvase-like protein is requered for the site-specific integration of the temperate lactococcal bacteriophage TP901-1

    DEFF Research Database (Denmark)

    Christiansen, Bettina; Brøndsted, Lone; Vogensen, Finn K.

    1996-01-01

    upstream of attP. The N-terminal 150 to 1180 amino acids of Orf1 showed 38 to 44% similarity to the resolvase group of site-specific integrases, while no similarity to know proteins was found in the C-terminal end. Bacteriophage 'TP901-1 therefore contains a unique integration system that does not resemble...... the Int class of site-specific integrases usually found in temperate bacteriophages. The constructed integration vector, pBC170, integrates into the chromosomal attachment site very efficiently and forms stable transformants with a frequency corresponding to 20% of the transformation efficiency....

  19. RNA secondary structures of the bacteriophage phi6 packaging regions.

    OpenAIRE

    Pirttimaa, M J; Bamford, D H

    2000-01-01

    Bacteriophage phi6 genome consists of three segments of double-stranded RNA. During maturation, single-stranded copies of these segments are packaged into preformed polymerase complex particles. Only phi6 RNA is packaged, and each particle contains only one copy of each segment. An in vitro packaging and replication assay has been developed for phi6, and the packaging signals (pac sites) have been mapped to the 5' ends of the RNA segments. In this study, we propose secondary structure models ...

  20. Bacteriophage lambda: early pioneer and still relevant

    Science.gov (United States)

    Casjens, Sherwood R.; Hendrix, Roger W.

    2015-01-01

    Molecular genetic research on bacteriophage lambda carried out during its golden age from the mid 1950's to mid 1980's was critically important in the attainment of our current understanding of the sophisticated and complex mechanisms by which the expression of genes is controlled, of DNA virus assembly and of the molecular nature of lysogeny. The development of molecular cloning techniques, ironically instigated largely by phage lambda researchers, allowed many phage workers to switch their efforts to other biological systems. Nonetheless, since that time the ongoing study of lambda and its relatives have continued to give important new insights. In this review we give some relevant early history and describe recent developments in understanding the molecular biology of lambda's life cycle. PMID:25742714

  1. ELISA Cut-off Point for the Diagnosis of Human Brucellosis; a Comparison with Serum Agglutination Test

    Directory of Open Access Journals (Sweden)

    Anahita Sanaei Dashti

    2012-03-01

    Full Text Available Background: Brucellosis is a world-wide disease, which has a diverse clinical manifestation, and its diagnosis has to be proven by laboratory data. Serum agglutination test (SAT is the most-widely used test for diagnosing brucellosis. The enzyme linked immunosorbent assay (ELISA can also determine specific antibody classes against brucella. It is a sensitive, simple and rapid test, which could be an acceptable alternative to SAT with fewer limitations, however, like any other new test it should be further evaluated and standardized for various populations. This study was planned to determine an optimal cut-off point, for ELISA which would offer maximum sensitivity and specificity for the test when compared to SAT.Methods: Four hundred and seven patients with fever and other compatible symptoms of brucellosis were enrolled in the study. Serum agglutination test, 2-Mercaptoethanol test, and ELISA were performed on their sera. Results: The cut-off point of 53 IU/ml of ELISA-IgG yielded the maximal sensitivity and specificity comparing to the other levels of ELISA-IgG, and was considered the best cut off-point of ELISA-IgG to diagnose acute brucellosis. At this cut-off, the sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, and negative likelihood ratio were 84.09%, 85.38%, 62.20, 94.90, 5.75, 0.18, respectively.Conclusion: The best cut-off point of ELISA-IgG is 53 IU/ml, which yields the maximal sensitivity and specificity to diagnose acute brucellosis.

  2. Misidentification of Vibrio cholerae O155 isolated from imported shrimp as O serogroup O139 due to cross-agglutination with commercial O139 antisera

    DEFF Research Database (Denmark)

    Dalsgaard, A.; Mazur, J.; Dalsgaard, Inger

    2002-01-01

    . The strain contained two plasmids, in contrast to other O139 strains, which normally do not contain plasmids. The characteristics of the strain led to further agglutination testing with other antisera that are not commercially available, and the strain was found to agglutinate O155 antiserum in repeated...... was isolated from imported raw frozen shrimp. The toxigenicity of the strain was analyzed, and the results of a polymerase chain reaction showed that the V. cholerae strain did not contain the virulence genes ctx, tcp9, and zot, which are normally found in V. cholerae O1 and O139. The strain was resistant...... to colistin and spectinomycin. The high susceptibility of the strain to antimicrobial agents was confirmed by the lack of an SXT element, a self-transmissible, chromosomal genetic element that is normally present in O139 strains and encodes resistance to sulfonamides, trimethoprim, and streptomycin...

  3. Problem-Solving Test: RNA and Protein Synthesis in Bacteriophage-Infected "E. coli" Cells

    Science.gov (United States)

    Szeberenyi, Jozsef

    2008-01-01

    The classic experiment presented in this problem-solving test was designed to identify the template molecules of translation by analyzing the synthesis of phage proteins in "Escherichia coli" cells infected with bacteriophage T4. The work described in this test led to one of the most seminal discoveries of early molecular biology: it dealt a…

  4. Genetic and mechanistic evaluation for the mixed-field agglutination in B3 blood type with IVS3+5G>A ABO gene mutation.

    Directory of Open Access Journals (Sweden)

    Ding-Ping Chen

    Full Text Available BACKGROUND: The ABO blood type B(3 is the most common B subtype in the Chinese population with a frequency of 1/900. Although IVS3+5G>A (rs55852701 mutation of B gene has been shown to associate with the development of B(3 blood type, genetic and mechanistic evaluation for the unique mixed-field agglutination phenotype has not yet been completely addressed. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we analyzed 16 cases of confirmed B(3 individuals and found that IVS3+5G>A attributes to all cases of B(3. RT-PCR analyses revealed the presence of at least 7 types of aberrant B(3 splicing transcripts with most of the transcripts causing early termination and producing non-functional protein during translation. The splicing transcript without exon 3 that was predicted to generate functional B(3 glycosyltransferase lacking 19 amino acids at the N-terminal segment constituted only 0.9% of the splicing transcripts. Expression of the B(3 cDNA with exon 3 deletion in the K562 erythroleukemia cells revealed that the B(3 glycosyltransferase had only 40% of B(1 activity in converting H antigen to B antigen. Notably, the typical mixed-field agglutination of B(3-RBCs can be mimicked by adding anti-B antibody to the K562-B(3 cells. CONCLUSIONS/SIGNIFICANCE: This study thereby demonstrates that both aberrant splicing of B transcripts and the reduced B(3 glycosyltransferase activity contribute to weak B expression and the mixed-field agglutination of B(3, adding to the complexity for the regulatory mechanisms of ABO gene expression.

  5. The Effectiveness of Bacteriophages against Methicillin-Resistant Staphylococcus aureus ST398 Nasal Colonization in Pigs

    NARCIS (Netherlands)

    Verstappen, Koen M; Tulinski, Pawel; Duim, Birgitta; Fluit, Ad C; Carney, Jennifer; van Nes, Arie; Wagenaar, Jaap A

    2016-01-01

    UNLABELLED: Methicillin-resistant Staphylococcus aureus (MRSA) is an important colonizer in animals and an opportunistic pathogen in humans. In humans, MRSA can cause infections that might be difficult to treat because of antimicrobial resistance. The use of bacteriophages has been suggested as a

  6. Bacteriophage populations

    International Nuclear Information System (INIS)

    Klieve, A.V.; Gilbert, R.A.

    2005-01-01

    Bacteriophages are ubiquitous to the rumen ecosystem; they have a role in nitrogen metabolism through bacterial lysis in the rumen, they may help to regulate bacterial population densities, be an agent for genetic exchange and be of use in biocontrol of bacterial populations through phage therapy. In Chapter 2.1, classical methodologies to enable the isolation, enumeration, storage and morphological characterization of phages were presented. In addition to these classic procedures, molecular biological techniques have resulted in a range of methodologies to investigate the type, topology and size of phage nucleic acids, to fingerprint individual phage strains and to create a profile of ruminal phage populations. Different phage families possess all the currently identified combinations of double-stranded or single-stranded RNA or DNA and may also possess unusual bases such as 5-hydroxymethylcytosine (found in T-even phage) or 5- hydroxymethyluracil and uracil in place of thymidine. In all morphological groups of phage except the filamentous phages, the nucleic acid is contained within a head or polyhedral structure, predominantly composed of protein. Filamentous phages have their nucleic acid contained inside the helical filament, occupying much of its length. Many of the procedures used with phage nucleic acids and double-stranded (ds) DNA, in particular, are not specific to ruminal phages but are the same as in other areas where nucleic acids are investigated and are covered elsewhere in the literature and this chapter. Most applications with rumen phages are similar to those reported for phages of non-ruminal bacteria and are covered in general texts such as Maniatis et al. In this chapter, we will concentrate on aspects of methodology as they relate to ruminal phages

  7. Gammasphaerolipovirus, a newly proposed bacteriophage genus, unifies viruses of halophilic archaea and thermophilic bacteria within the novel family Sphaerolipoviridae.

    Science.gov (United States)

    Pawlowski, Alice; Rissanen, Ilona; Bamford, Jaana K H; Krupovic, Mart; Jalasvuori, Matti

    2014-06-01

    A new family of viruses named Sphaerolipoviridae has been proposed recently. It comprises icosahedral, tailless haloarchaeal viruses with an internal lipid membrane located between the protein capsid and the dsDNA genome. The proposed family Sphaerolipoviridae was divided into two genera: Alphasphaerolipovirus, including Haloarcula hispanica viruses SH1, PH1 and HHIV-2, and Betasphaerolipovirus, including Natrinema virus SNJ1. Here, we propose to expand the family Sphaerolipoviridae to include a group of bacteriophages infecting extreme thermophilic Thermus thermophilus and sharing a number of structural and genomic properties with archaeal sphaerolipoviruses. This new group comprises two members, lytic phage P23-77 and temperate phage IN93, as well as putative members P23-72 and P23-65H. In addition, several related proviruses have been discovered as integrated elements in bacterial genomes of the families Thermus and Meiothermus. Morphology of the virus particles and the overall capsid architecture of these bacteriophages resembles that of archaeal members of the Sphaerolipoviridae, including an unusual capsid arrangement in a T = 28 dextro lattice. Alpha- and betasphaerolipoviruses share with P23-77-like bacteriophages a conserved block of core genes that encode a putative genome-packaging ATPase and the two major capsid proteins (MCPs). The recently determined X-ray structure of the small and large MCPs of P23-77 revealed a single beta-barrel (jelly-roll) fold that is superimposable with the cryo-EM density maps of the SH1 capsomers. Given the common features of these viruses, we propose to include the so far unclassified P23-77-like bacteriophages into a new genus, "Gammasphaerolipovirus", within the family Sphaerolipoviridae.

  8. The evaluation of hollow-fiber ultrafiltration and celite concentration of enteroviruses, adenoviruses and bacteriophage from different water matrices

    Data.gov (United States)

    U.S. Environmental Protection Agency — The data to support the evaluation of hollow-fiber ultrafiltration and celite concentration of enteroviruses, adenoviruses and bacteriophage from different water...

  9. Rapid detection of methicillin resistance in coagulase-negative staphylococci by a penicillin-binding protein 2a-specific latex agglutination test.

    Science.gov (United States)

    Horstkotte, M A; Knobloch, J K; Rohde, H; Mack, D

    2001-10-01

    The detection of PBP 2a by the MRSA-Screen latex agglutination test with 201 clinical coagulase-negative staphylococci had an initial sensitivity of 98% and a high degree of specificity for Staphylococcus epidermidis strains compared to PCR for mecA. Determination of oxacillin MICs evaluated according to the new breakpoint (0.5 microg/ml) of the National Committee for Clinical Laboratory Standards exhibited an extremely low specificity for this population.

  10. Structural studies demonstrating a bacteriophage-like replication cycle of the eukaryote-infecting Paramecium bursaria chlorella virus-1.

    Directory of Open Access Journals (Sweden)

    Elad Milrot

    2017-08-01

    Full Text Available A fundamental stage in viral infection is the internalization of viral genomes in host cells. Although extensively studied, the mechanisms and factors responsible for the genome internalization process remain poorly understood. Here we report our observations, derived from diverse imaging methods on genome internalization of the large dsDNA Paramecium bursaria chlorella virus-1 (PBCV-1. Our studies reveal that early infection stages of this eukaryotic-infecting virus occurs by a bacteriophage-like pathway, whereby PBCV-1 generates a hole in the host cell wall and ejects its dsDNA genome in a linear, base-pair-by-base-pair process, through a membrane tunnel generated by the fusion of the virus internal membrane with the host membrane. Furthermore, our results imply that PBCV-1 DNA condensation that occurs shortly after infection probably plays a role in genome internalization, as hypothesized for the infection of some bacteriophages. The subsequent perforation of the host photosynthetic membranes presumably enables trafficking of viral genomes towards host nuclei. Previous studies established that at late infection stages PBCV-1 generates cytoplasmic organelles, termed viral factories, where viral assembly takes place, a feature characteristic of many large dsDNA viruses that infect eukaryotic organisms. PBCV-1 thus appears to combine a bacteriophage-like mechanism during early infection stages with a eukaryotic-like infection pathway in its late replication cycle.

  11. STUDIES ON THE BACTERIOPHAGE OF D'HERELLE : IX. EVIDENCE OF HYDROLYSIS OF BACTERIAL PROTEIN DURING LYSIS.

    Science.gov (United States)

    Hetler, D M; Bronfenbrenner, J

    1928-07-31

    1. During the process of lysis by bacteriophage, there is an appreciable increase in the amount of free amino acid present in the culture. 2. The increase of free amino acid is due to hydrolysis of bacterial protein.

  12. The effectiveness of bacteriophages against methicillin-resistant Staphylococcus aureus ST398 nasal colonization in pigs

    NARCIS (Netherlands)

    Verstappen, Koen M.; Tulinski, Pawel; Duim, Birgitta; Fluit, Ad C.; Carney, Jennifer; Nes, Van Arie; Wagenaar, Jaap A.

    2016-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) is an important colonizer in animals and an opportunistic pathogen in humans. In humans, MRSA can cause infections that might be difficult to treat because of antimicrobial resistance. The use of bacteriophages has been suggested as a potential

  13. Reverse-Transcriptase PCR Detection of Leptospira: Absence of Agreement with Single-Specimen Microscopic Agglutination Testing.

    Science.gov (United States)

    Waggoner, Jesse J; Balassiano, Ilana; Mohamed-Hadley, Alisha; Vital-Brazil, Juliana Magalhães; Sahoo, Malaya K; Pinsky, Benjamin A

    2015-01-01

    Reference diagnostic tests for leptospirosis include nucleic acid amplification tests, bacterial culture, and microscopic agglutination testing (MAT) of acute and convalescent serum. However, clinical laboratories often do not receive paired specimens. In the current study, we tested serum samples using a highly sensitive real-time nucleic acid amplification test for Leptospira and compared results to MAT performed on the same specimens. 478 serum samples from suspected leptospirosis cases in Rio de Janeiro were tested using a real-time RT-PCR for the diagnosis of leptospirosis, malaria and dengue (the Lepto-MD assay). The Lepto-MD assay detects all species of Leptospira (saprophytic, intermediate, and pathogenic), and in the current study, we demonstrate that this assay amplifies both Leptospira RNA and DNA. Dengue virus RNA was identified in 10 patients, and no cases of malaria were detected. A total of 65 samples (13.6%) were positive for Leptospira: 35 samples (7.3%) in the Lepto-MD assay, 33 samples (6.9%) by MAT, and 3 samples tested positive by both (kappa statistic 0.02). Poor agreement between methods was consistent regardless of the titer used to define positive MAT results or the day of disease at sample collection. Leptospira nucleic acids were detected in the Lepto-MD assay as late as day 22, and cycle threshold values did not differ based on the day of disease. When Lepto-MD assay results were added to the MAT results for all patients in 2008 (n=818), the number of detected leptospirosis cases increased by 30.4%, from 102 (12.5%) to 133 (16.3%). This study demonstrates a lack of agreement between nucleic acid detection of Leptospira and single-specimen MAT, which may result from the clearance of bacteremia coinciding with the appearance of agglutinating antibodies. A combined testing strategy for acute leptospirosis, including molecular and serologic testing, appears necessary to maximize case detection.

  14. Neutron irradiation of bacteriophage λ

    International Nuclear Information System (INIS)

    Bozin, D.; Milosevic, M. . E-mail address of corresponding author: bozinde@vin.bg.ac.yu

    2005-01-01

    Double strand breaks (DSB) are the most dangerous lesions in DNA caused by irradiation, but many other lesions, usually called mutations, have not been clearly identified. These lesions, like DSB, can be the source of serious chromosomal damages and finally - cell death. Growing interest in heavy particles for radiotherapy and radioprotection encourages the search of the molecular basis of their action. In this respect, we chose bacteriophage λ1390 as the model system for the study of consequences of neutron irradiation. This derivative of λ phage possesses an unique ability to reversibly reorganize their genome in response to various selective pressures. The phages were irradiated with 13 Gy of mixed neutrons (7.5 Gy from fast and 5.6 Gy from thermal neutrons) and phages genomes were tested to DSB and mutations. Additionally, the stability of λ capsid proteins were tested. After all tests, we can conclude that, under our conditions, low flux of neutrons does not induce neither DNA strand break or DNA mutation nor the stability of λ capsid proteins. (author)

  15. Bacteriophage Distributions and Temporal Variability in the Ocean’s Interior

    Directory of Open Access Journals (Sweden)

    Elaine Luo

    2017-11-01

    Full Text Available Bacteriophages are numerically the most abundant DNA-containing entities in the oligotrophic ocean, yet how specific phage populations vary over time and space remains to be fully explored. Here, we conducted a metagenomic time-series survey of double-stranded DNA phages throughout the water column in the North Pacific Subtropical Gyre, encompassing 1.5 years from depths of 25 to 1,000 m. Viral gene sequences were identified in assembled metagenomic samples, yielding an estimated 172,385 different viral gene families. Viral marker gene distributions suggested that lysogeny was more prevalent at mesopelagic depths than in surface waters, consistent with prior prophage induction studies using mitomycin C. A total of 129 ALOHA viral genomes and genome fragments from 20 to 108 kbp were selected for further study, which represented the most abundant phages in the water column. Phage genotypes displayed discrete population structures. Most phages persisted throughout the time-series and displayed a strong depth structure that mirrored the stratified depth distributions of co-occurring bacterial taxa in the water column. Mesopelagic phages were distinct from surface water phages with respect to diversity, gene content, putative life histories, and temporal persistence, reflecting depth-dependent differences in host genomic architectures and phage reproductive strategies. The spatiotemporal distributions of the most abundant open-ocean bacteriophages that we report here provide new insight into viral temporal persistence, life history, and virus-host-environment interactions throughout the open-ocean water column.

  16. Analysis of Bacillus subtilis sporulation with spore-converting bacteriophage PMB12.

    OpenAIRE

    Kinney, D M; Bramucci, M G

    1981-01-01

    Previous observations concerning the ability of the spore-converting bacteriophage PMB12 to cause sporulation in certain sporulation-deficient mutants of Bacillus subtilis 168 were extended to include a spoOK mutant and a mutant temperature sensitive for sporulation due to a ribosomal mutation. Mutants of PMB12 that were unable to induce sporulation in the spoOK mutant were isolated to determine whether PMB12-encoded products had to affect the sporulation-specific functions of both the transc...

  17. Framing the Future with Bacteriophages in Agriculture.

    Science.gov (United States)

    Svircev, Antonet; Roach, Dwayne; Castle, Alan

    2018-04-25

    The ability of agriculture to continually provide food to a growing world population is of crucial importance. Bacterial diseases of plants and animals have continually reduced production since the advent of crop cultivation and animal husbandry practices. Antibiotics have been used extensively to mitigate these losses. The rise of antimicrobial resistant (AMR) bacteria, however, together with consumers’ calls for antibiotic-free products, presents problems that threaten sustainable agriculture. Bacteriophages (phages) are proposed as bacterial population control alternatives to antibiotics. Their unique properties make them highly promising but challenging antimicrobials. The use of phages in agriculture also presents a number of unique challenges. This mini-review summarizes recent development and perspectives of phages used as antimicrobial agents in plant and animal agriculture at the farm level. The main pathogens and their adjoining phage therapies are discussed.

  18. High temperature and bacteriophages can indirectly select for bacterial pathogenicity in environmental reservoirs.

    Directory of Open Access Journals (Sweden)

    Ville-Petri Friman

    2011-03-01

    Full Text Available The coincidental evolution hypothesis predicts that traits connected to bacterial pathogenicity could be indirectly selected outside the host as a correlated response to abiotic environmental conditions or different biotic species interactions. To investigate this, an opportunistic bacterial pathogen, Serratia marcescens, was cultured in the absence and presence of the lytic bacteriophage PPV (Podoviridae at 25°C and 37°C for four weeks (N = 5. At the end, we measured changes in bacterial phage-resistance and potential virulence traits, and determined the pathogenicity of all bacterial selection lines in the Parasemia plantaginis insect model in vivo. Selection at 37°C increased bacterial motility and pathogenicity but only in the absence of phages. Exposure to phages increased the phage-resistance of bacteria, and this was costly in terms of decreased maximum population size in the absence of phages. However, this small-magnitude growth cost was not greater with bacteria that had evolved in high temperature regime, and no trade-off was found between phage-resistance and growth rate. As a result, phages constrained the evolution of a temperature-mediated increase in bacterial pathogenicity presumably by preferably infecting the highly motile and virulent bacteria. In more general perspective, our results suggest that the traits connected to bacterial pathogenicity could be indirectly selected as a correlated response by abiotic and biotic factors in environmental reservoirs.

  19. Evaluation of Penicillin Binding Protein 2a Latex Agglutination Assay for Identification of Methicillin-Resistant Staphylococcus aureus Directly from Blood Cultures

    OpenAIRE

    Chapin, Kimberle C.; Musgnug, Michael C.

    2004-01-01

    The penicillin binding protein 2a (PBP2a) latex agglutination test using a blood culture pellet was compared to the oxacillin screen agar method using isolated colonies. For blood cultures positive for Staphylococcus aureus (n = 70), the direct PBP2a test was 18% sensitive and 100% specific. The PBP2a test shows poor sensitivity when used directly with positive blood cultures.

  20. Crystallization and preliminary crystallographic characterization of the origin-binding domain of the bacteriophage λ O replication initiator

    International Nuclear Information System (INIS)

    Struble, E. B.; Gittis, A. G.; Bianchet, M. A.; McMacken, R.

    2007-01-01

    Crystallization and preliminary diffraction data of the N-terminal 19–139 fragment of the origin-binding domain of bacteriophage λ O replication initiator are reported. The bacteriophage λ O protein binds to the λ replication origin (oriλ) and serves as the primary replication initiator for the viral genome. The binding energy derived from the binding of O to oriλ is thought to help drive DNA opening to facilitate initiation of DNA replication. Detailed understanding of this process is severely limited by the lack of high-resolution structures of O protein or of any lambdoid phage-encoded paralogs either with or without DNA. The production of crystals of the origin-binding domain of λ O that diffract to 2.5 Å is reported. Anomalous dispersion methods will be used to solve this structure

  1. Reactivity of various leishmanial antigens in a direct agglutination test and their value in differentiating post-kala azar dermal leishmaniasis from leprosy and other skin conditions

    NARCIS (Netherlands)

    El Harith, A.; Chowdhury, S.; Al-Masum, A.; Semião-Santos, S.; Das, P. K.; Akhter, S.; Vetter, J. C.; Haq, I.

    1996-01-01

    A direct agglutination test (DAT) for the detection of post-kala azar dermal leishmaniasis (PKDL) was evaluated in conditions that simulate the disease clinically or immunologically. A reference strain of Leishmania donovani (LEM 1399), and antigen preparations from Leishmania isolates from

  2. Protozoan Predation of Escherichia coli O157:H7 Is Unaffected by the Carriage of Shiga Toxin-Encoding Bacteriophages.

    Directory of Open Access Journals (Sweden)

    Carrie E Schmidt

    Full Text Available Escherichia coli O157:H7 is a food-borne bacterium that causes hemorrhagic diarrhea and hemolytic uremic syndrome in humans. While cattle are a known source of E. coli O157:H7 exposure resulting in human infection, environmental reservoirs may also be important sources of infection for both cattle and humans. Bacteriophage-encoded Shiga toxins (Stx carried by E. coli O157:H7 may provide a selective advantage for survival of these bacteria in the environment, possibly through their toxic effects on grazing protozoa. To determine Stx effects on protozoan grazing, we co-cultured Paramecium caudatum, a common ciliate protozoon in cattle water sources, with multiple strains of Shiga-toxigenic E. coli O157:H7 and non-Shiga toxigenic cattle commensal E. coli. Over three days at ambient laboratory temperature, P. caudatum consistently reduced both E. coli O157:H7 and non-Shiga toxigenic E. coli populations by 1-3 log cfu. Furthermore, a wild-type strain of Shiga-toxigenic E. coli O157:H7 (EDL933 and isogenic mutants lacking the A subunit of Stx 2a, the entire Stx 2a-encoding bacteriophage, and/or the entire Stx 1-encoding bacteriophage were grazed with similar efficacy by both P. caudatum and Tetrahymena pyriformis (another ciliate protozoon. Therefore, our data provided no evidence of a protective effect of either Stx or the products of other bacteriophage genes on protozoan predation of E. coli. Further research is necessary to determine if the grazing activity of naturally-occurring protozoa in cattle water troughs can serve to decrease cattle exposure to E. coli O157:H7 and other Shiga-toxigenic E. coli.

  3. Rapid Detection of Methicillin Resistance in Coagulase-Negative Staphylococci by a Penicillin-Binding Protein 2a-Specific Latex Agglutination Test

    OpenAIRE

    Horstkotte, Matthias A.; Knobloch, Johannes K.-M.; Rohde, Holger; Mack, Dietrich

    2001-01-01

    The detection of PBP 2a by the MRSA-Screen latex agglutination test with 201 clinical coagulase-negative staphylococci had an initial sensitivity of 98% and a high degree of specificity for Staphylococcus epidermidis strains compared to PCR for mecA. Determination of oxacillin MICs evaluated according to the new breakpoint (0.5 μg/ml) of the National Committee for Clinical Laboratory Standards exhibited an extremely low specificity for this population.

  4. Phage “delay” towards enhancing bacterial escape from biofilms: a more comprehensive way of viewing resistance to bacteriophages

    Directory of Open Access Journals (Sweden)

    Stephen T. Abedon

    2017-03-01

    Full Text Available In exploring bacterial resistance to bacteriophages, emphasis typically is placed on those mechanisms which completely prevent phage replication. Such resistance can be detected as extensive reductions in phage ability to form plaques, that is, reduced efficiency of plating. Mechanisms include restriction-modification systems, CRISPR/Cas systems, and abortive infection systems. Alternatively, phages may be reduced in their “vigor” when infecting certain bacterial hosts, that is, with phages displaying smaller burst sizes or extended latent periods rather than being outright inactivated. It is well known, as well, that most phages poorly infect bacteria that are less metabolically active. Extracellular polymers such as biofilm matrix material also may at least slow phage penetration to bacterial surfaces. Here I suggest that such “less-robust” mechanisms of resistance to bacteriophages could serve bacteria by slowing phage propagation within bacterial biofilms, that is, delaying phage impact on multiple bacteria rather than necessarily outright preventing such impact. Related bacteria, ones that are relatively near to infected bacteria, e.g., roughly 10+ µm away, consequently may be able to escape from biofilms with greater likelihood via standard dissemination-initiating mechanisms including erosion from biofilm surfaces or seeding dispersal/central hollowing. That is, given localized areas of phage infection, so long as phage spread can be reduced in rate from initial points of contact with susceptible bacteria, then bacterial survival may be enhanced due to bacteria metaphorically “running away” to more phage-free locations. Delay mechanisms—to the extent that they are less specific in terms of what phages are targeted—collectively could represent broader bacterial strategies of phage resistance versus outright phage killing, the latter especially as require specific, evolved molecular recognition of phage presence. The

  5. Functional requirements for bacteriophage growth: gene essentiality and expression in mycobacteriophage Giles.

    Science.gov (United States)

    Dedrick, Rebekah M; Marinelli, Laura J; Newton, Gerald L; Pogliano, Kit; Pogliano, Joseph; Hatfull, Graham F

    2013-05-01

    Bacteriophages represent a majority of all life forms, and the vast, dynamic population with early origins is reflected in their enormous genetic diversity. A large number of bacteriophage genomes have been sequenced. They are replete with novel genes without known relatives. We know little about their functions, which genes are required for lytic growth, and how they are expressed. Furthermore, the diversity is such that even genes with required functions - such as virion proteins and repressors - cannot always be recognized. Here we describe a functional genomic dissection of mycobacteriophage Giles, in which the virion proteins are identified, genes required for lytic growth are determined, the repressor is identified, and the transcription patterns determined. We find that although all of the predicted phage genes are expressed either in lysogeny or in lytic growth, 45% of the predicted genes are non-essential for lytic growth. We also describe genes required for DNA replication, show that recombination is required for lytic growth, and that Giles encodes a novel repressor. RNAseq analysis reveals abundant expression of a small non-coding RNA in a lysogen and in late lytic growth, although it is non-essential for lytic growth and does not alter lysogeny. © 2013 Blackwell Publishing Ltd.

  6. Spectroscopic analysis of the oligosaccharides produced by bacteriophage-borne enzyme action on Klebsiella K36 polysaccharide

    Energy Technology Data Exchange (ETDEWEB)

    Ravenscroft, N; Jackson, G E; Joao, H; Stephen, A M

    1988-06-01

    Mass spectral analysis of the permethylated oligossacharides obtained by bacteriophage degradation of Klebsiella K36 polysaccharide has enabled the sequence of sugar residues to be determined. 2D N.m.r. studies confirmed the inter-sugar linkages and established the anomeric configurations.

  7. DPS - a rapid method for genome sequencing of DNA-containing bacteriophages directly from a single plaque

    DEFF Research Database (Denmark)

    Kot, Witold Piotr; Vogensen, Finn Kvist; Sørensen, Søren Johannes

    2014-01-01

    Bacteriophages (phages) coexist with bacteria in all environments and influence microbial diversity, evolution and industrial production processes. As a result of this major impact of phages on microbes, tools that allow rapid characterization of phages are needed. Today, one of the most powerful...

  8. [A STUDY OF THE ISOLATED BACTERIOPHAGE ΦAB-SP7 ADSORPTION ON THE CELL SURFACE OF THE AZOSPIRILLUM BRASILENSE SP7].

    Science.gov (United States)

    Guliy, O I; Karavaeva, O A; Velikov, V A; Sokolov, O I; Pavily, S A; Larionova, O S; Burov, A M; Ignatov, O V

    2016-01-01

    The bacteriophage ΦAb-Sp7 was isolated from the cells of the Azospirillum brasilense Sp7. The morphology, size of the gram-negative colonies, and range of lytic activity against other strains and species of the genus Azospirillum was tested. The isolated phage DNA was examined using electrophoretic and restriction analysis, and the size of the genome were established. The electron microscopy. resuIts show that the phage (capsid) has a strand-like form. The electron microscopy study of the bacteriophage ΦAb-Sp7 adsorption on the A. brasilense Sp7 bacterial surface was performed.

  9. Plasmid integration in a wide range of bacteria mediated by the integrase of Lactobacillus delbrueckii bacteriophage mv4.

    Science.gov (United States)

    Auvray, F; Coddeville, M; Ritzenthaler, P; Dupont, L

    1997-01-01

    Bacteriophage mv4 is a temperate phage infecting Lactobacillus delbrueckii subsp. bulgaricus. During lysogenization, the phage integrates its genome into the host chromosome at the 3' end of a tRNA(Ser) gene through a site-specific recombination process (L. Dupont et al., J. Bacteriol., 177:586-595, 1995). A nonreplicative vector (pMC1) based on the mv4 integrative elements (attP site and integrase-coding int gene) is able to integrate into the chromosome of a wide range of bacterial hosts, including Lactobacillus plantarum, Lactobacillus casei (two strains), Lactococcus lactis subsp. cremoris, Enterococcus faecalis, and Streptococcus pneumoniae. Integrative recombination of pMC1 into the chromosomes of all of these species is dependent on the int gene product and occurs specifically at the pMC1 attP site. The isolation and sequencing of pMC1 integration sites from these bacteria showed that in lactobacilli, pMC1 integrated into the conserved tRNA(Ser) gene. In the other bacterial species where this tRNA gene is less or not conserved; secondary integration sites either in potential protein-coding regions or in intergenic DNA were used. A consensus sequence was deduced from the analysis of the different integration sites. The comparison of these sequences demonstrated the flexibility of the integrase for the bacterial integration site and suggested the importance of the trinucleotide CCT at the 5' end of the core in the strand exchange reaction. PMID:9068626

  10. Bacteriophages : an underestimated role in human and animal health ?

    Directory of Open Access Journals (Sweden)

    Marianne eDe Paepe

    2014-03-01

    Full Text Available Metagenomic approaches applied to viruses have highlighted their prevalence in almost all microbial ecosystems investigated. In all ecosystems, notably those associated with humans or animals, the viral fraction is dominated by bacteriophages. Whether they contribute to dysbiosis, i.e. the departure from microbiota composition in symbiosis at equilibrium and entry into a state favoring human or animal disease is unknown at present. This review summarizes what has been learnt on phages associated with human and animal microbiota, and focuses on examples illustrating the several ways by which phages may contribute to a shift to pathogenesis, either by modifying population equilibrium, by horizontal transfer, or by modulating immunity.

  11. Scavenger Receptor C Mediates Phagocytosis of White Spot Syndrome Virus and Restricts Virus Proliferation in Shrimp

    Science.gov (United States)

    Yang, Ming-Chong; Shi, Xiu-Zhen; Yang, Hui-Ting; Sun, Jie-Jie; Xu, Ling; Wang, Xian-Wei; Zhao, Xiao-Fan

    2016-01-01

    Scavenger receptors are an important class of pattern recognition receptors that play several important roles in host defense against pathogens. The class C scavenger receptors (SRCs) have only been identified in a few invertebrates, and their role in the immune response against viruses is seldom studied. In this study, we firstly identified an SRC from kuruma shrimp, Marsupenaeus japonicus, designated MjSRC, which was significantly upregulated after white spot syndrome virus (WSSV) challenge at the mRNA and protein levels in hemocytes. The quantity of WSSV increased in shrimp after knockdown of MjSRC, compared with the controls. Furthermore, overexpression of MjSRC led to enhanced WSSV elimination via phagocytosis by hemocytes. Pull-down and co-immunoprecipitation assays demonstrated the interaction between MjSRC and the WSSV envelope protein. Electron microscopy observation indicated that the colloidal gold-labeled extracellular domain of MjSRC was located on the outer surface of WSSV. MjSRC formed a trimer and was internalized into the cytoplasm after WSSV challenge, and the internalization was strongly inhibited after knockdown of Mjβ-arrestin2. Further studies found that Mjβ-arrestin2 interacted with the intracellular domain of MjSRC and induced the internalization of WSSV in a clathrin-dependent manner. WSSV were co-localized with lysosomes in hemocytes and the WSSV quantity in shrimp increased after injection of lysosome inhibitor, chloroquine. Collectively, this study demonstrated that MjSRC recognized WSSV via its extracellular domain and invoked hemocyte phagocytosis to restrict WSSV systemic infection. This is the first study to report an SRC as a pattern recognition receptor promoting phagocytosis of a virus. PMID:28027319

  12. Comparison of the cystine-tryptic digest agar-carbohydrate co-agglutination and BACTEC Neisseria differentiation methods for identification of Neisseria gonorrhoeae in the clinical laboratory

    International Nuclear Information System (INIS)

    Morello, J.A.; Beheshti, S.; Bohnhoff, M.

    1980-01-01

    The author evaluated CTA-carbohydrate, BACTEC and co-agglutination systems to determine their accuracy for identifying N. gonorrhoeae strains and for distinguishing them from other Neisseria species. BACTEC is a radiometric assay based on the measurement of liberated radiolabelled CO 2 from metabolised carbohydrates which have been tagged with 14 C. (Auth.)

  13. The use of the rapid osmotic fragility test as an additional test to diagnose canine immune-mediated haemolytic anaemia

    DEFF Research Database (Denmark)

    Paes, Geert; Paepe, Dominique; Meyer, Evelyne

    2013-01-01

    Background: Diagnosing canine immune-mediated haemolytic anaemia (IMHA) is often challenging because all currently available tests have their limitations. Dogs with IMHA often have an increased erythrocyte osmotic fragility (OF), a characteristic that is sometimes used in the diagnosis of IMHA...... hyperlipemic dogs (group 3), 10 dogs with lymphoma (group 4), 8 dogs with an infection (group 5) and 13 healthy dogs (group 6) were included. In all dogs, blood smear examination, in-saline auto-agglutination test, Coombs' test, COFT and ROFT were performed. In the COFT, OF5, OF50 and OF90 were defined...

  14. Binding of Mn-deoxyribonucleoside Triphosphates to the Active Site of the DNA Polymerase of Bacteriophage T7

    Energy Technology Data Exchange (ETDEWEB)

    B Akabayov; C Richardson

    2011-12-31

    Divalent metal ions are crucial as cofactors for a variety of intracellular enzymatic activities. Mg{sup 2+}, as an example, mediates binding of deoxyribonucleoside 5'-triphosphates followed by their hydrolysis in the active site of DNA polymerase. It is difficult to study the binding of Mg{sup 2+} to an active site because Mg{sup 2+} is spectroscopically silent and Mg{sup 2+} binds with low affinity to the active site of an enzyme. Therefore, we substituted Mg{sup 2+} with Mn{sup 2+}:Mn{sup 2+} that is not only visible spectroscopically but also provides full activity of the DNA polymerase of bacteriophage T7. In order to demonstrate that the majority of Mn{sup 2+} is bound to the enzyme, we have applied site-directed titration analysis of T7 DNA polymerase using X-ray near edge spectroscopy. Here we show how X-ray near edge spectroscopy can be used to distinguish between signal originating from Mn{sup 2+} that is free in solution and Mn{sup 2+} bound to the active site of T7 DNA polymerase. This method can be applied to other enzymes that use divalent metal ions as a cofactor.

  15. Isolation and Characterization of a Virulent Bacteriophage AB1 of Acinetobacter baumannii

    Directory of Open Access Journals (Sweden)

    Jia Shiru

    2010-04-01

    Full Text Available Abstract Background Acinetobacter baumannii is an emerging nosocomial pathogen worldwide with increasing prevalence of multi-drug and pan-drug resistance. A. baumannii exists widely in natural environment, especially in health care settings, and has been shown difficult to be eradicated. Bacteriophages are often considered alternative agent for controlling bacterial infection and contamination. In this study, we described the isolation and characterization of one virulent bacteriophage AB1 capable of specifically infecting A. baumannii. Results A virulent bacteriophage AB1, specific for infecting a clinical strain A. baumannii KD311, was first isolated from marine sediment sample. Restriction analysis indicated that phage AB1 was a dsDNA virus with an approximate genome size of 45.2 kb to 46.9 kb. Transmission electron microscopy showed that phage AB1 had an icosahedral head with a non-contractile tail and collar or whisker structures, and might be tentatively classified as a member of the Siphoviridae family. Proteomic pattern of phage AB1, generated by SDS-PAGE using purified phage particles, revealed five major bands and six minor bands with molecular weight ranging from 14 to 80 kilo-dalton. Also determined was the adsorption rate of phage AB1 to the host bacterium, which was significantly enhanced by addition of 10 mM CaCl2. In a single step growth test, phage AB1 was shown having a latent period of 18 minutes and a burst size of 409. Moreover, pH and thermal stability of phage AB1 were also investigated. At the optimal pH 6.0, 73.2% of phages survived after 60 min incubation at 50°C. When phage AB1 was used to infect four additional clinical isolates of A. baumannii, one clinical isolate of Stenotrophomonas maltophilia, and Pseudomonas aeruginosa lab strains PAK and PAO1, none of the tested strains was found susceptible, indicating a relatively narrow host range for phage AB1. Conclusion Phage AB1 was capable of eliciting efficient lysis

  16. Mechanisms for the initiation of bacteriophage T7 DNA replication

    International Nuclear Information System (INIS)

    Fuller, C.W.; Beauchamp, B.B.; Engler, M.J.; Lechner, R.L.; Matson, S.W.; Tabor, S.; White, J.H.; Richardson, C.C.

    1983-01-01

    Genetic analysis of bacteriophage T7 has shown that the products of phage genes 1, 2, 3, 4, 5, and 6 are required for phage DNA synthesis in vivo. T7 RNA polymerase is the translation product of gene 1. This RNA polymerase is required for transcription of most of the phage genome, including genes 2 through 6. T7 RNA polymerase promoters consist of a highly conserved 23-bp DNA sequence. There are 17 such promoters in the T7 DNA molecule, all of which direct transcription from the same strand of the DNA. 70 references, 11 figures

  17. Direct detection of methicillin resistance in Staphylococcus aureus in blood culture broth by use of a penicillin binding protein 2a latex agglutination test.

    Science.gov (United States)

    Qian, Qinfang; Venkataraman, Lata; Kirby, James E; Gold, Howard S; Yamazumi, Toshiaki

    2010-04-01

    We studied the utility of performing a penicillin binding protein 2a latex agglutination (PBP-LA) assay directly on Bactec blood culture broth samples containing Staphylococcus aureus to rapidly detect methicillin resistance. The sensitivity, specificity, positive predictive value, and negative predictive value of this method were 94.1%, 97.5%, 98%, and 92.9%, respectively.

  18. Direct Detection of Methicillin Resistance in Staphylococcus aureus in Blood Culture Broth by Use of a Penicillin Binding Protein 2a Latex Agglutination Test▿

    OpenAIRE

    Qian, Qinfang; Venkataraman, Lata; Kirby, James E.; Gold, Howard S.; Yamazumi, Toshiaki

    2010-01-01

    We studied the utility of performing a penicillin binding protein 2a latex agglutination (PBP-LA) assay directly on Bactec blood culture broth samples containing Staphylococcus aureus to rapidly detect methicillin resistance. The sensitivity, specificity, positive predictive value, and negative predictive value of this method were 94.1%, 97.5%, 98%, and 92.9%, respectively.

  19. Branched Lateral Tail Fiber Organization in T5-Like Bacteriophages DT57C and DT571/2 is Revealed by Genetic and Functional Analysis

    Directory of Open Access Journals (Sweden)

    Alla K. Golomidova

    2016-01-01

    Full Text Available The T5-like siphoviruses DT57C and DT571/2, isolated from horse feces, are very closely related to each other, and most of their structural proteins are also nearly identical to T5 phage. Their LTFs (L-shaped tail fibers, however, are composed of two proteins, LtfA and LtfB, instead of the single Ltf of bacteriophage T5. In silico and mutant analysis suggests a possible branched structure of DT57C and DT571/2 LTFs, where the LtfB protein is connected to the phage tail via the LtfA protein and with both proteins carrying receptor recognition domains. Such adhesin arrangement has not been previously recognized in siphoviruses. The LtfA proteins of our phages are found to recognize different host O-antigen types: E. coli O22-like for DT57C phage and E. coli O87 for DT571/2. LtfB proteins are identical in both phages and recognize another host receptor, most probably lipopolysaccharide (LPS of E. coli O81 type. In these two bacteriophages, LTF function is essential to penetrate the shield of the host’s O-antigens. We also demonstrate that LTF-mediated adsorption becomes superfluous when the non-specific cell protection by O-antigen is missing, allowing the phages to bind directly to their common secondary receptor, the outer membrane protein BtuB. The LTF independent adsorption was also demonstrated on an O22-like host mutant missing O-antigen O-acetylation, thus showing the biological value of this O-antigen modification for cell protection against phages.

  20. Design of a CRISPR-Cas system to increase resistance of Bacillus subtilis to bacteriophage SPP1.

    Science.gov (United States)

    Jakutyte-Giraitiene, Lina; Gasiunas, Giedrius

    2016-08-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) together with CRISPR-associated (cas) genes form an adaptive prokaryotic immune system which provides acquired resistance against viruses and plasmids. Bacillus subtilis presently is the best-characterized laboratory model for Gram-positive bacteria and also widely used for industrial production of enzymes, vitamins and antibiotics. In this study, we show that type II-A CRISPR-Cas system from Streptococcus thermophilus can be transferred into B. subtilis and provides heterologous protection against phage infection. We engineered a heterologous host by cloning S. thermophilus Cas9 and a spacer targeting bacteriophage SPP1 into the chromosome of B. subtilis, which does not harbor its own CRISPR-Cas systems. We found that the heterologous CRISPR-Cas system is functionally active in B. subtilis and provides resistance against bacteriophage SPP1 infection. The high efficiency of the acquired immunity against phage could be useful in generation of biotechnologically important B. subtilis strains with engineered chromosomes.

  1. Selective Inhibition of Histone Deacetylation in Melanoma Increases Targeted Gene Delivery by a Bacteriophage Viral Vector

    Directory of Open Access Journals (Sweden)

    Samuel Campbell

    2018-04-01

    Full Text Available The previously developed adeno-associated virus/phage (AAVP vector, a hybrid between M13 bacteriophage (phage viruses that infect bacteria only and human Adeno-Associated Virus (AAV, is a promising tool in targeted gene therapy against cancer. AAVP can be administered systemically and made tissue specific through the use of ligand-directed targeting. Cancer cells and tumor-associated blood vessels overexpress the αν integrin receptors, which are involved in tumor angiogenesis and tumor invasion. AAVP is targeted to these integrins via a double cyclic RGD4C ligand displayed on the phage capsid. Nevertheless, there remain significant host-defense hurdles to the use of AAVP in targeted gene delivery and subsequently in gene therapy. We previously reported that histone deacetylation in cancer constitutes a barrier to AAVP. Herein, to improve AAVP-mediated gene delivery to cancer cells, we combined the vector with selective adjuvant chemicals that inhibit specific histone deacetylases (HDAC. We examined the effects of the HDAC inhibitor C1A that mainly targets HDAC6 and compared this to sodium butyrate, a pan-HDAC inhibitor with broad spectrum HDAC inhibition. We tested the effects on melanoma, known for HDAC6 up-regulation, and compared this side by side with a normal human kidney HEK293 cell line. Varying concentrations were tested to determine cytotoxic levels as well as effects on AAVP gene delivery. We report that the HDAC inhibitor C1A increased AAVP-mediated transgene expression by up to ~9-fold. These findings indicate that selective HDAC inhibition is a promising adjuvant treatment for increasing the therapeutic value of AAVP.

  2. Review: elimination of bacteriophages in whey and whey products

    Directory of Open Access Journals (Sweden)

    Zeynep eAtamer

    2013-07-01

    Full Text Available As the cheese market faces strong international competition, the optimization of production processes becomes more important for the economic success of dairy companies. In dairy productions, whey from former cheese batches is frequently re-used to increase the yield, to improve the texture and to increase the nutrient value of the final product. Recycling of whey cream and particulated whey proteins is also routinely performed. Most bacteriophages, however, survive pasteurization and may re-enter the cheese manufacturing process. There is a risk that phages multiply to high numbers during the production. Contamination of whey samples with bacteriophages may cause problems in cheese factories because whey separation often leads to aerosol-borne phages and thus contamination of the factory environment. Furthermore, whey cream or whey proteins used for recycling into cheese matrices may contain thermo-resistant phages. Drained cheese whey can be contaminated with phages as high as 109 phages per mL. When whey batches are concentrated, phage titers can increase significantly by a factor of 10 hindering a complete elimination of phages. To eliminate the risk of fermentation failure during recycling of whey, whey treatments assuring an efficient reduction of phages are indispensable. This review focuses on inactivation of phages in whey by thermal treatment, ultraviolet (UV light irradiation and membrane filtration. Inactivation by heat is the most common procedure. However, application of heat for inactivation of thermo-resistant phages in whey is restricted due to negative effects on the functional properties of native whey proteins. Therefore an alternative strategy applying combined treatments should be favoured - rather than heating the dairy product at extreme temperature/time combinations. By using membrane filtration or UV treatment in combination with thermal treatment, phage numbers in whey can be reduced sufficiently to prevent subsequent

  3. Review: elimination of bacteriophages in whey and whey products

    Science.gov (United States)

    Atamer, Zeynep; Samtlebe, Meike; Neve, Horst; J. Heller, Knut; Hinrichs, Joerg

    2013-01-01

    As the cheese market faces strong international competition, the optimization of production processes becomes more important for the economic success of dairy companies. In dairy productions, whey from former cheese batches is frequently re-used to increase the yield, to improve the texture and to increase the nutrient value of the final product. Recycling of whey cream and particulated whey proteins is also routinely performed. Most bacteriophages, however, survive pasteurization and may re-enter the cheese manufacturing process. There is a risk that phages multiply to high numbers during the production. Contamination of whey samples with bacteriophages may cause problems in cheese factories because whey separation often leads to aerosol-borne phages and thus contamination of the factory environment. Furthermore, whey cream or whey proteins used for recycling into cheese matrices may contain thermo-resistant phages. Drained cheese whey can be contaminated with phages as high as 109 phages mL-1. When whey batches are concentrated, phage titers can increase significantly by a factor of 10 hindering a complete elimination of phages. To eliminate the risk of fermentation failure during recycling of whey, whey treatments assuring an efficient reduction of phages are indispensable. This review focuses on inactivation of phages in whey by thermal treatment, ultraviolet (UV) light irradiation, and membrane filtration. Inactivation by heat is the most common procedure. However, application of heat for inactivation of thermo-resistant phages in whey is restricted due to negative effects on the functional properties of native whey proteins. Therefore an alternative strategy applying combined treatments should be favored – rather than heating the dairy product at extreme temperature/time combinations. By using membrane filtration or UV treatment in combination with thermal treatment, phage numbers in whey can be reduced sufficiently to prevent subsequent phage accumulations

  4. Bacteriophage-encoded shiga toxin gene in atypical bacterial host

    Directory of Open Access Journals (Sweden)

    Casas Veronica

    2011-07-01

    Full Text Available Abstract Background Contamination from fecal bacteria in recreational waters is a major health concern since bacteria capable of causing human disease can be found in animal feces. The Dog Beach area of Ocean Beach in San Diego, California is a beach prone to closures due to high levels of fecal indicator bacteria (FIB. A potential source of these FIB could be the canine feces left behind by owners who do not clean up after their pets. We tested this hypothesis by screening the DNA isolated from canine feces for the bacteriophage-encoded stx gene normally found in the virulent strains of the fecal bacterium Escherichia coli. Results Twenty canine fecal samples were collected, processed for total and bacterial fraction DNA, and screened by PCR for the stx gene. The stx gene was detected in the total and bacterial fraction DNA of one fecal sample. Bacterial isolates were then cultivated from the stx-positive fecal sample. Eighty nine of these canine fecal bacterial isolates were screened by PCR for the stx gene. The stx gene was detected in five of these isolates. Sequencing and phylogenetic analyses of 16S rRNA gene PCR products from the canine fecal bacterial isolates indicated that they were Enterococcus and not E. coli. Conclusions The bacteriophage-encoded stx gene was found in multiple species of bacteria cultivated from canine fecal samples gathered at the shoreline of the Dog Beach area of Ocean Beach in San Diego, California. The canine fecal bacteria carrying the stx gene were not the typical E. coli host and were instead identified through phylogenetic analyses as Enterococcus. This suggests a large degree of horizontal gene transfer of exotoxin genes in recreational waters.

  5. Prevalence of agglutinating antibodies to Toxoplasma gondii in striped skunks (Mephitis mephitis), opossums (Didelphis virginiana), and raccoons (Procyon lotor) from Connecticut

    OpenAIRE

    Mitchell, S. M.; Richardson, D. J.; Lindsay, D. S.

    2006-01-01

    The prevalence of agglutinating antibodies to Toxoplasma gondii was examined in striped skunks (Mephitis mephitis), opossums (Didelphis virginiana), and raccoons (Procyon lotor) from 8 cities in Connecticut. Ten (42%) of the 24 striped skunks, 2 of 7 (29%) opossums, and 12 of 12 (100%) raccoons were positive at dilutions of 1:50 or greater. These results suggest that T. gondii is prevalent in the environment, or prey items, or both, of these omnivores in Connecticut.

  6. Prevalence of agglutinating antibodies to Toxoplasma gondii in striped skunks (Mephitis mephitis), opossums (Didelphis virginiana), and raccoons (Procyon lotor) from Connecticut.

    Science.gov (United States)

    Mitchell, Sheila M; Richardson, Dennis J; Lindsay, David S

    2006-06-01

    The prevalence of agglutinating antibodies to Toxoplasma gondii was examined in striped skunks (Mephitis mephitis), opossums (Didelphis virginiana), and raccoons (Procyon lotor) from 8 cities in Connecticut. Ten (42%) of the 24 striped skunks, 2 of 7 (29%) opossums, and 12 of 12 (100%) raccoons were positive at dilutions of 1:50 or greater. These results suggest that T. gondii is prevalent in the environment, or prey items, or both, of these omnivores in Connecticut.

  7. Bacteriophage therapy to combat bacterial infections in poultry.

    Science.gov (United States)

    Wernicki, Andrzej; Nowaczek, Anna; Urban-Chmiel, Renata

    2017-09-16

    Infections in poultry are an economic and health problem in Europe and worldwide. The most common infections are associated with salmonellosis, colibacillosis, campylobacteriosis, and others. The prevalence of Campylobacter-positive poultry flocks in European countries varies from 18% to 90%. In the United States, the prevalence of infected flocks is nearly 90%. A similar percentage of infection has been noted for salmonellosis (about 75-90%) and E. coli (90-95%). The occurence of Clostridium perfringens is a major problem for the poultry industry, with some estimates suggesting colonization of as many as 95% of chickens, resulting in clinical or subclinical infections. In the US, annual economic losses due to Salmonella infections run from $1.188 billion to over $11.588 billion, based on an estimated 1.92 million cases. Similar costs are observed in the case of other types of infections. In 2005 economic losses in the the poultry industry due to mortalities reached 1,000,000 USD.Infections caused by these pathogens, often through poultry products, are also a serious public health issue.The progressive increase in the number of multi-drug resistant bacteria and the complete ban on the use of antibiotics in livestock feed in the EU, as well as the partial ban in the US, have led to the growth of research on the use of bacteriophages to combat bacterial infections in humans and animals.The high success rate and safety of phage therapy in comparison with antibiotics are partly due to their specificity for selected bacteria and the ability to infect only one species, serotype or strain. This mechanism does not cause the destruction of commensal bacterial flora. Phages are currently being used with success in humans and animals in targeted therapies for slow-healing infections. They have also found application in the US in eliminating pathogens from the surface of foods of animal and plant origin. At a time of growing antibiotic resistance in bacteria and the resulting

  8. The efficacy of sewage influent-isolated bacteriophages on Pseudomonas aeruginosa in a mixed-species biofilm

    KAUST Repository

    Yap, Scott

    2016-12-01

    The growth of environmentally persistent biofilms in cooling towers causes several associated problems, including microbiologically-induced corrosion (MIC) and biofouling. Current chemical control methods are not only ineffective against biofilms and costly to procure, they also have downstream environmental impacts when released untreated, or incur additional treatment costs. Bacteriophages are alternative biofilm control agents that have the potential to be more effective, cheaper to produce and yet have a more benign effect on the environment. In this study, biofilms grown under conditions simulating seawater fed cooling towers were characterized and the differences in growth and community make-up across time and different substrates were assessed. An MIC associated bacterium common in cooling tower water, P. aeruginosa, was chosen. Seven bacteriophage strains found to be effective against the chosen bacterium were isolated from wastewater influent. The relative effectiveness of these strains was measured against P. aeruginosa across different salinities. Separate biofilms fed with P. aeruginosa enriched seawater were characterized and the effectiveness of the isolated strains, singly and in cocktails, against the enriched biofilms was measured.

  9. Selection of Functional Quorum Sensing Systems by Lysogenic Bacteriophages in Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Miguel A. Saucedo-Mora

    2017-08-01

    Full Text Available Quorum sensing (QS in Pseudomonas aeruginosa coordinates the expression of virulence factors, some of which are used as public goods. Since their production is a cooperative behavior, it is susceptible to social cheating in which non-cooperative QS deficient mutants use the resources without investing in their production. Nevertheless, functional QS systems are abundant; hence, mechanisms regulating the amount of cheating should exist. Evidence that demonstrates a tight relationship between QS and the susceptibility of bacteria against the attack of lytic phages is increasing; nevertheless, the relationship between temperate phages and QS has been much less explored. Therefore, in this work, we studied the effects of having a functional QS system on the susceptibility to temperate bacteriophages and how this affects the bacterial and phage dynamics. We find that both experimentally and using mathematical models, that the lysogenic bacteriophages D3112 and JBD30 select QS-proficient P. aeruginosa phenotypes as compared to the QS-deficient mutants during competition experiments with mixed strain populations in vitro and in vivo in Galleria mellonella, in spite of the fact that both phages replicate better in the wild-type background. We show that this phenomenon restricts social cheating, and we propose that temperate phages may constitute an important selective pressure toward the conservation of bacterial QS.

  10. Widal agglutination titre: a rapid serological diagnosis of typhoid fever in developing countries

    International Nuclear Information System (INIS)

    Aftab, R.; Khurshid, R.

    2009-01-01

    To study the reliability of a single Widal test and to find out the diagnostic significance of 'O' and 'H' agglutinin titre in the diagnosis of typhoid fever. Community-based case-control study conducted from Jan 2001 to June 2007. The blood samples were collected from the medical and out door department of Sir Ganga Ram Hospitals, Lahore. The diagnostic value of an acute phase single Widal agglutination test for suspected typhoid fever was evaluated in 733 consecutive patients with fever lasting 6 or more days. In 733 patients with fever 84 (11.45%) were positive for Widal test. A noteworthy rise 1/320 of H and/or O agglutinin titre was observed in 86 (11.3%) of patients with typhoid fever. In the absence of vaccination an elevated level of H and/or O agglutinin titre of 1: 320 is of diagnostic value for typhoid fever especially in our setting where a single sample of serum is relied on for the diagnosis of typhoid fever. (author)

  11. Widal agglutination titre: a rapid serological diagnosis of typhoid fever in developing countries

    Energy Technology Data Exchange (ETDEWEB)

    Aftab, R [Fatima Jinnah Medical College Lahore, Lahore (Pakistan). Dept. of Pathology; Khurshid, R [Fatima Jinnah Medical College Lahore, Lahore (Pakistan). Dept. of Biochemistry

    2009-01-15

    To study the reliability of a single Widal test and to find out the diagnostic significance of 'O' and 'H' agglutinin titre in the diagnosis of typhoid fever. Community-based case-control study conducted from Jan 2001 to June 2007. The blood samples were collected from the medical and out door department of Sir Ganga Ram Hospitals, Lahore. The diagnostic value of an acute phase single Widal agglutination test for suspected typhoid fever was evaluated in 733 consecutive patients with fever lasting 6 or more days. In 733 patients with fever 84 (11.45%) were positive for Widal test. A noteworthy rise 1/320 of H and/or O agglutinin titre was observed in 86 (11.3%) of patients with typhoid fever. In the absence of vaccination an elevated level of H and/or O agglutinin titre of 1: 320 is of diagnostic value for typhoid fever especially in our setting where a single sample of serum is relied on for the diagnosis of typhoid fever. (author)

  12. Multiple factors and processes involved in host cell killing by bacteriophage Mu: characterization and mapping.

    Science.gov (United States)

    Waggoner, B T; Marrs, C F; Howe, M M; Pato, M L

    1984-07-15

    The regions of bacteriophage Mu involved in host cell killing were determined by infection of a lambda-immune host with 12 lambda pMu-transducing phages carrying different amounts of Mu DNA beginning at the left end. Infecting lambda pMu phages containing 5.0 (+/- 0.2) kb or less of the left end of Mu DNA did not kill the lambda-immune host, whereas lambda pMu containing 5.1 kb did kill, thus locating the right end of the kil gene between approximately 5.0 and 5.1 kb. For the Kil+ phages the extent of killing increased as the multiplicity of infection (m.o.i.) increased. In addition, killing was also affected by the presence of at least two other regions of Mu DNA: one, located between 5.1 and 5.8 kb, decreased the extent of killing; the other, located between 6.3 and 7.9 kb, greatly increased host cell killing. Killing was also assayed after lambda pMu infection of a lambda-immune host carrying a mini-Mu deleted for most of the B gene and the middle region of Mu DNA. Complementation of mini-Mu replication by infecting B+ lambda pMu phages resulted in killing of the lambda-immune, mini-Mu-containing host, regardless of the presence or absence of the Mu kil gene. The extent of host cell killing increased as the m.o.i. of the infecting lambda pMu increased, and was further enhanced by both the presence of the kil gene and the region located between 6.3 and 7.9 kb. These distinct processes of kil-mediated killing in the absence of replication and non-kil-mediated killing in the presence of replication were also observed after induction of replication-deficient and kil mutant prophages, respectively.

  13. Antibiotic Resistant Bacterial Isolates from Captive Green Turtles and In Vitro Sensitivity to Bacteriophages

    Directory of Open Access Journals (Sweden)

    Alessandro Delli Paoli Carini

    2017-01-01

    Full Text Available This study aimed to test multidrug resistant isolates from hospitalised green turtles (Chelonia mydas and their environment in North Queensland, Australia, for in vitro susceptibility to bacteriophages. Seventy-one Gram-negative bacteria were isolated from green turtle eye swabs and water samples. Broth microdilution tests were used to determine antibiotic susceptibility. All isolates were resistant to at least two antibiotics, with 24% being resistant to seven of the eight antibiotics. Highest resistance rates were detected to enrofloxacin (77% and ampicillin (69.2%. More than 50% resistance was also found to amoxicillin/clavulanic acid (62.5%, ceftiofur (53.8%, and erythromycin (53.3%. All the enriched phage filtrate mixtures resulted in the lysis of one or more of the multidrug resistant bacteria, including Vibrio harveyi and V. parahaemolyticus. These results indicate that antibiotic resistance is common in Gram-negative bacteria isolated from hospitalised sea turtles and their marine environment in North Queensland, supporting global concern over the rapid evolution of multidrug resistant genes in the environment. Using virulent bacteriophages as antibiotic alternatives would not only be beneficial to turtle health but also prevent further addition of multidrug resistant genes to coastal waters.

  14. Characterization of bacteriophages infecting clinical isolates of Pseudomonas aeruginosa stored in a culture collection

    Directory of Open Access Journals (Sweden)

    C.C.S. Zanetti

    2013-08-01

    Full Text Available Some clinical isolates of Pseudomonas aeruginosa stored in our culture collection did not grow or grew poorly and showed lysis on the culture plates when removed from the collection and inoculated on MacConkey agar. One hypothesis was that bacteriophages had infected and killed those clinical isolates. To check the best storage conditions to maintain viable P. aeruginosa for a longer time, clinical isolates were stored at various temperatures and were grown monthly. We investigated the presence of phage in 10 clinical isolates of P. aeruginosa stored in our culture collection. Four strains of P. aeruginosa were infected by phages that were characterized by electron microscopy and isolated to assess their ability to infect. The best condition to maintain the viability of the strains during storage was in water at room temperature. Three Siphoviridae and two Myoviridae phages were visualized and characterized by morphology. We confirmed the presence of bacteriophages infecting clinical isolates, and their ability to infect and lyse alternative hosts. Strain PAO1, however, did not show lysis to any phage. Mucoid and multidrug resistant strains of P. aeruginosa showed lysis to 50% of the phages tested.

  15. Experimental Examination of Bacteriophage Latent-Period Evolution as a Response to Bacterial Availability

    OpenAIRE

    Abedon, Stephen T.; Hyman, Paul; Thomas, Cameron

    2003-01-01

    For obligately lytic bacteriophage (phage) a trade-off exists between fecundity (burst size) and latent period (a component of generation time). This trade-off occurs because release of phage progeny from infected bacteria coincides with destruction of the machinery necessary to produce more phage progeny. Here we employ phage mutants to explore issues of phage latent-period evolution as a function of the density of phage-susceptible bacteria. Theory suggests that higher bacterial densities s...

  16. Bacteriophage-based tools: recent advances and novel applications [version 1; referees: 3 approved

    Directory of Open Access Journals (Sweden)

    Lisa O'Sullivan

    2016-11-01

    Full Text Available Bacteriophages (phages are viruses that infect bacterial hosts, and since their discovery over a century ago they have been primarily exploited to control bacterial populations and to serve as tools in molecular biology. In this commentary, we highlight recent diverse advances in the field of phage research, going beyond bacterial control using whole phage, to areas including biocontrol using phage-derived enzybiotics, diagnostics, drug discovery, novel drug delivery systems and bionanotechnology.

  17. Characterization of the integrated filamentous phage Pf5 and its involvement in small-colony formation

    NARCIS (Netherlands)

    Mooij, Marlies J.; Drenkard, Eliana; Llamas, María A.; Vandenbroucke-Grauls, Christina M. J. E.; Savelkoul, Paul H. M.; Ausubel, Frederick M.; Bitter, Wilbert

    2007-01-01

    Bacteriophages play an important role in bacterial virulence and phenotypic variation. It has been shown that filamentous bacteriophage Pf4 of Pseudomonas aeruginosa strain PAO1 mediates the formation of small-colony variants (SCVs) in biofilms. This morphology type is associated with parameters of

  18. Momordica charantia seed lectin: toxicity, bacterial agglutination and antitumor properties.

    Science.gov (United States)

    Kabir, Syed Rashel; Nabi, Md Mahamodun; Nurujjaman, Md; Abu Reza, Md; Alam, A H M Khurshid; Uz Zaman, Rokon; Khalid-Bin-Ferdaus, Khandaker Md; Amin, Ruhul; Khan, Md Masudul Hasan; Hossain, Md Anowar; Uddin, Md Salim; Mahmud, Zahid Hayat

    2015-03-01

    In last three decades, several studies were carried out on the D-galactose-specific lectin of Momordica charantia seeds (MCL). In the present study, in vitro growth inhibition (8-23 %) at different concentrations (6-24 μg/ml) of MCL was observed against Ehrlich ascites carcinoma (EAC) cells by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. MCL also showed 28, 45, and 75 % growth inhibitions against EAC cells when administered 1.2, 2.0, and 2.8 mg/kg/day (i.p.), respectively for five consequent days in vivo in mice. After lectin treatment, the level of red blood cell and hemoglobin was increased significantly with the decrease of white blood cell and maintained the normal level when compared with EAC-bearing control and normal mice without EAC cells. Although MCL caused cell cycle arrest at G0/G1 phase of EAC cells, any irregular shape or apoptotic morphological alterations in the lectin-treated EAC cells was not observed by an optical and fluorescence microscope. Lectin showed toxicity against brine shrimp nauplii with an LC50 value of 49.7 μg/ml. Four out of seven pathogenic bacteria were agglutinated by MCL in the absence of inhibitory sugar D-lactose/D-galactose. In conclusion, MCL showed strong cytotoxic effect and therefore can be used as a potent anticancer chemotherapeutic agent.

  19. Identification of Quaternary Structure and Functional Domains of the CI Repressor from Bacteriophage TP901-1

    DEFF Research Database (Denmark)

    Pedersen, Margit; Lo Leggio, Leila; Grossmann, J. Günter

    2008-01-01

    is involved in the interaction with host proteins. By using small-angle X-ray scattering, we show for the first time the overall solution structure of a full-length wild-type bacteriophage repressor at low resolution revealing that the TP901-1 repressor forms a flat oligomer, most probably a trimer of dimers....

  20. TOTAL HEMOCYTE COUNT AND HEMOLYMPH GLUCOSE CONCENRATION RESPONSE OF SPINY LOBSTER Panulirus homarus ON RATIO OF SHELTER

    Directory of Open Access Journals (Sweden)

    Suhaiba Djai

    2017-11-01

    Full Text Available This research was conducted to assess the physiological response of the lobster Panulirus homarus for the ratio of the shelters. The method used completely randomized design with two replicates of each treatments with shelter ratio (A 1 : 5, (B 3 : 5, (C 4 : 5, (D 5 : 5. Weight average for 184 lobsters with the stocking density of 23 lobsters for each treatment was 32.64 ± 0.58 g. The experiment was conducted for 60 days. The lobster was fed with trash fish and acclimatized for 7 days before the experiment. Observations on the physiologycal of every 10 days. The physiological responses that observed were total hemocyte count (THC and hemolymph glucose concentration. The results showed that 4:5 was the best lobster shelter ratio because it could reduce stress levels. This is indicated by the stable values of THC and hemolymph glucose level during the experiment and supported by the growth of 57.28 ± 0.15 g and survival rate of 91.31 ± 2.60%. Keywords: lobster, Panulirus homarus, ratio, shelter, THC, glucose

  1. Fluorescent nanodiamond-bacteriophage conjugates maintain host specificity.

    Science.gov (United States)

    Trinh, Jimmy T; Alkahtani, Masfer H; Rampersaud, Isaac; Rampersaud, Arfaan; Scully, Marlan; Young, Ryland F; Hemmer, Philip; Zeng, Lanying

    2018-06-01

    Rapid identification of specific bacterial strains within clinical, environmental, and food samples can facilitate the prevention and treatment of disease. Fluorescent nanodiamonds (FNDs) are being developed as biomarkers in biology and medicine, due to their excellent imaging properties, ability to accept surface modifications, and lack of toxicity. Bacteriophages, the viruses of bacteria, can have exquisite specificity for certain hosts. We propose to exploit the properties of FNDs and phages to develop phages conjugated with FNDs as long-lived fluorescent diagnostic reagents. In this study, we develop a simple procedure to create such fluorescent probes by functionalizing the FNDs and phages with streptavidin and biotin, respectively. We find that the FND-phage conjugates retain the favorable characteristics of the individual components and can discern their proper host within a mixture. This technology may be further explored using different phage/bacteria systems, different FND color centers and alternate chemical labeling schemes for additional means of bacterial identification and new single-cell/virus studies. © 2018 Wiley Periodicals, Inc.

  2. denV gene of bacteriophage T4 restores DNA excision repair to mei-9 and mus201 mutants of Drosophila melanogaster

    International Nuclear Information System (INIS)

    Banga, S.S.; Boyd, J.B.; Valerie, K.; Harris, P.V.; Kurz, E.M.; de Riel, J.K.

    1989-01-01

    The denV gene of bacteriophage T4 was fused to a Drosophila hsp70 (70-kDa heat shock protein) promoter and introduced into the germ line of Drosophila by P-element-mediated transformation. The protein product of that gene (endonuclease V) was detected in extracts of heat-shocked transformants with both enzymological and immunoblotting procedures. That protein restores both excision repair and UV resistance to mei-9 and mus201 mutants of this organism. These results reveal that the denV gene can compensate for excision-repair defects in two very different eukayotic mutants, in that the mus201 mutants are typical of excision-deficient mutants in other organisms, whereas the mei-9 mutants exhibit a broad pleiotropism that includes a strong meiotic deficiency. This study permits an extension of the molecular analysis of DNA repair to the germ line of higher eukaryotes. It also provides a model system for future investigations of other well-characterized microbial repair genes on DNA damage in the germ line of this metazoan organism

  3. Prostaglandin-mediated recovery from bacteriosemia delays larval development in fall armyworms, Spodoptera frugiperda

    Science.gov (United States)

    Insect immunity includes a surveillance system that detects and signals infections, coupled with hemocytic and humoral immune functions. These functions are signaled and coordinated by several biochemicals, including biogenic amines, insect cytokines, peptides and prostaglandins (PGs). The actions o...

  4. Some aspects of the mechanism of bacteriophage function. Final progress report

    International Nuclear Information System (INIS)

    Freifelder, D.

    1977-01-01

    Data are summarized from a ten-year study on the radiobiology of phages. The results showed that: phages are inactivated principally by damage to DNA; DNA damage is of two types, base damage and double-strand breakage; double-strand breakage may be lethal because of interruption within a gene, however in phage systems the damage is more fundamental in that only a single DNA fragment is injected into the host; E. coli phage T4 is relatively resistant to inactivation by x-rays; and the rate of production of strand breaks and base damage is nearly the same in bacteriophage and bacteria

  5. Biology and genomics of an historic therapeutic Escherichia coli bacteriophage collection

    DEFF Research Database (Denmark)

    Baig, Abiyad; Colom, Joan; Barrow, Paul

    2017-01-01

    We have performed microbiological and genomic characterization of an historic collection of nine bacteriophages, specifically infecting a K1 E. coli O18:K1:H7 ColV+ strain. These phages were isolated from sewage and tested for their efficacy in vivo for the treatment of systemic E. coli infection...... in a mouse infection model by Smith and Huggins (1982). The aim of the study was to identify common microbiological and genomic characteristics, which co-relate to the performance of these phages in in vivo study. These features will allow an informed selection of phages for use as therapeutic agents...

  6. Feasibility of using a bacteriophage-based structural color sensor for screening the geographical origins of agricultural products

    Science.gov (United States)

    Seol, Daun; Moon, Jong-Sik; Lee, Yujin; Han, Jiye; Jang, Daeil; Kang, Dong-Jin; Moon, Jiyoung; Jang, Eunjin; Oh, Jin-Woo; Chung, Hoeil

    2018-05-01

    An M13 bacteriophage-based color sensor, which can change its structural color upon interaction with a gaseous molecule, was evaluated as a screening tool for the discrimination of the geographical origins of three different agricultural products (garlic, onion, and perilla). Exposure of the color sensor to sample odors induced the self-assembled M13 bacteriophage bundles to swell by the interaction of amino acid residues (repeating units of four glutamates) on the bacteriophage with the odor components, resulting in a change in the structural color of the sensor. When the sensor was exposed to the odors of garlic and onion samples, the RGB color changes were considerable because of the strong interactions of the odor components such as disulfides with the glutamate residues on the sensor. Although the patterns of the color variations were generally similar between the domestic and imported samples, some degrees of dissimilarities in their intensities were also observed. Although the magnitude of color change decreased for perilla, the color change patterns between the two groups were somewhat different. With the acquired RGB data, a support vector machine was employed to distinguish the domestic and imported samples, and the resulting accuracies in the measurements of garlic, onion, and perilla samples were 94.1, 88.7, and 91.6%, respectively. The differences in the concentrations of the odor components between both groups and/or the presence of specific components exclusively in the odor of one group allowed the color sensor-based discrimination. The demonstrated color sensor was thus shown to be a potentially versatile and simple as an on-site screening tool. Strategies able to further improve the sensor performance were also discussed.

  7. Versatile de novo enzyme activity in capsid proteins from an engineered M13 bacteriophage library.

    Science.gov (United States)

    Casey, John P; Barbero, Roberto J; Heldman, Nimrod; Belcher, Angela M

    2014-11-26

    Biocatalysis has grown rapidly in recent decades as a solution to the evolving demands of industrial chemical processes. Mounting environmental pressures and shifting supply chains underscore the need for novel chemical activities, while rapid biotechnological progress has greatly increased the utility of enzymatic methods. Enzymes, though capable of high catalytic efficiency and remarkable reaction selectivity, still suffer from relative instability, high costs of scaling, and functional inflexibility. Herein, we developed a biochemical platform for engineering de novo semisynthetic enzymes, functionally modular and widely stable, based on the M13 bacteriophage. The hydrolytic bacteriophage described in this paper catalyzes a range of carboxylic esters, is active from 25 to 80 °C, and demonstrates greater efficiency in DMSO than in water. The platform complements biocatalysts with characteristics of heterogeneous catalysis, yielding high-surface area, thermostable biochemical structures readily adaptable to reactions in myriad solvents. As the viral structure ensures semisynthetic enzymes remain linked to the genetic sequences responsible for catalysis, future work will tailor the biocatalysts to high-demand synthetic processes by evolving new activities, utilizing high-throughput screening technology and harnessing M13's multifunctionality.

  8. Characterization of the genome of the dairy Lactobacillus helveticus bacteriophage {Phi}AQ113.

    Science.gov (United States)

    Zago, Miriam; Scaltriti, Erika; Rossetti, Lia; Guffanti, Alessandro; Armiento, Angelarita; Fornasari, Maria Emanuela; Grolli, Stefano; Carminati, Domenico; Brini, Elena; Pavan, Paolo; Felsani, Armando; D'Urzo, Annalisa; Moles, Anna; Claude, Jean-Baptiste; Grandori, Rita; Ramoni, Roberto; Giraffa, Giorgio

    2013-08-01

    The complete genomic sequence of the dairy Lactobacillus helveticus bacteriophage ΦAQ113 was determined. Phage ΦAQ113 is a Myoviridae bacteriophage with an isometric capsid and a contractile tail. The final assembled consensus sequence revealed a linear, circularly permuted, double-stranded DNA genome with a size of 36,566 bp and a G+C content of 37%. Fifty-six open reading frames (ORFs) were predicted, and a putative function was assigned to approximately 90% of them. The ΦAQ113 genome shows functionally related genes clustered together in a genome structure composed of modules for DNA replication/regulation, DNA packaging, head and tail morphogenesis, cell lysis, and lysogeny. The identification of genes involved in the establishment of lysogeny indicates that it may have originated as a temperate phage, even if it was isolated from natural cheese whey starters as a virulent phage, because it is able to propagate in a sensitive host strain. Additionally, we discovered that the ΦAQ113 phage genome is closely related to Lactobacillus gasseri phage KC5a and Lactobacillus johnsonii phage Lj771 genomes. The phylogenetic similarities between L. helveticus phage ΦAQ113 and two phages that belong to gut species confirm a possible common ancestral origin and support the increasing consideration of L. helveticus as a health-promoting organism.

  9. Phenotypic heterogeneity in a bacteriophage population only appears as stress-induced mutagenesis.

    Science.gov (United States)

    Yosef, Ido; Edgar, Rotem; Qimron, Udi

    2016-11-01

    Stress-induced mutagenesis has been studied in cancer cells, yeast, bacteria, and archaea, but not in viruses. In a recent publication, we present a bacteriophage model showing an apparent stress-induced mutagenesis. We show that the stress does not drive the mutagenesis, but only selects the fittest mutants. The mechanism underlying the observed phenomenon is a phenotypic heterogeneity that resembles persistence of the viral population. The new findings, the background for the ongoing debate on stress-induced mutagenesis, and the phenotypic heterogeneity underlying a novel phage infection strategy are discussed in this short manuscript.

  10. Extraction and Study of Bacteriophages, Used against Agents of Potato Soft Rot

    Directory of Open Access Journals (Sweden)

    Magda D. Davitashvili

    2012-12-01

    Full Text Available The use of specific bacteriophages and their complex mixtures against bacterial diseases is very effective. As for causative agent of potato soft rot Erwinia carotovora, specific phages (25 phages in total were extracted from diseased potato, soil and sewage. The study of their biological properties showed the diversity of phages in terms of lytic action, virion plaque and morphology, as well as in relation to different environmental factors. Phages showed explicit antibacterial activity in vitro in liquid and solid media, as well as during model tests of potato tubers artificial inoculation.

  11. Bacteriophage and their potential roles in the human oral cavity

    Directory of Open Access Journals (Sweden)

    Anna Edlund

    2015-04-01

    Full Text Available The human oral cavity provides the perfect portal of entry for viruses and bacteria in the environment to access new hosts. Hence, the oral cavity is one of the most densely populated habitats of the human body containing some 6 billion bacteria and potentially 35 times that many viruses. The role of these viral communities remains unclear; however, many are bacteriophage that may have active roles in shaping the ecology of oral bacterial communities. Other implications for the presence of such vast oral phage communities include accelerating the molecular diversity of their bacterial hosts as both host and phage mutate to gain evolutionary advantages. Additional roles include the acquisitions of new gene functions through lysogenic conversions that may provide selective advantages to host bacteria in response to antibiotics or other types of disturbances, and protection of the human host from invading pathogens by binding to and preventing pathogens from crossing oral mucosal barriers. Recent evidence suggests that phage may be more involved in periodontal diseases than were previously thought, as their compositions in the subgingival crevice in moderate to severe periodontitis are known to be significantly altered. However, it is unclear to what extent they contribute to dysbiosis or the transition of the microbial community into a state promoting oral disease. Bacteriophage communities are distinct in saliva compared to sub- and supragingival areas, suggesting that different oral biogeographic niches have unique phage ecology shaping their bacterial biota. In this review, we summarize what is known about phage communities in the oral cavity, the possible contributions of phage in shaping oral bacterial ecology, and the risks to public health oral phage may pose through their potential to spread antibiotic resistance gene functions to close contacts.

  12. Repair of ultraviolet light-induced DNA damage in cholera bacteriophages

    International Nuclear Information System (INIS)

    Palit, B.N.; Das, G.; Das, J.

    1983-01-01

    DNA repair-proficient and -deficient strains of Vibrio cholerae were used to examine host cell reactivation, Weigle reactivation and photoreactivation of u.v.-irradiated cholera bacteriophages. U.v. light-induced DNA damage in phages of different morphological and serological groups could be efficiently photoreactivated. Host cell reactivation of irradiated phages of different groups was different on the same indicator host. Phage phi149 was the most sensitive, and phi138 the most resistant to u.v. irradiation. While phi138 showed appreciable host cell reactivation, this was minimal for phi149. Attempts to demonstrate Weigle reactivation of u.v.-irradiated cholera phages were not successful, although u.v.-induced filamentation of host cells was observed. (author)

  13. Recovery status of bacteriophages of different livestock farms of Veterinary College, Adhartal, Jabalpur, India

    OpenAIRE

    Sanjay Shukla and S. D. Hirpurkar

    2011-01-01

    Study was conducted to know the presence of bacteriophage in sewage material which can play a very important role during therapy against the some antibiotic resistance organisms. During study waste water samples were collected from different depths of the wastewater collection tanks located in livestock farms of different species (Cattle, pig, goat and poultry). These samples were subjected primarily to rapid detection by streak plate method for the detection of lytic activity followed by pri...

  14. Determination of fecal bacteriophages fecales from pre-potable waters using a simple membrane filtration technique; Determinacion de bacteriofagos fecales en aguas pre-potables por la tecnica de filtracion por membrana

    Energy Technology Data Exchange (ETDEWEB)

    Luque, A.; Basanta, A. [EMASESA. Sevilla (Spain); Fuentes, M. V.; Alonso, M. C.; Borrego, J. J. [Universidad de Malaga (Spain)

    2000-07-01

    A modification of the simple filtration technique used for the detection and enumeration of fecal bacteriophages from pre-treated and treated drinking water is described. The filters and equipments used are similar to those employed for the classical bacterial indicator analysis, and the titer of bacteriophages may be recorded in only 6 hours. The proposed modification is based on the elimination of the bacterial decontamination of the water sample by use the selective medium mFC agar; and, in addition, an optimal visualization of the lysis plaques is obtained in this medium, which avoid the use of specific stains like triphenyl tetrazolium chloride (TTC). On the other hand, the proposed modification of the technique presents a high accuracy, since bacteriophage titers of<10 pfu/100 ml of water are detected. (Author) 46 refs.

  15. Lysogeny with Shiga Toxin 2-Encoding Bacteriophages Represses Type III Secretion in Enterohemorrhagic Escherichia coli

    Science.gov (United States)

    Xu, Xuefang; McAteer, Sean P.; Tree, Jai J.; Shaw, Darren J.; Wolfson, Eliza B. K.; Beatson, Scott A.; Roe, Andrew J.; Allison, Lesley J.; Chase-Topping, Margo E.; Mahajan, Arvind; Tozzoli, Rosangela; Woolhouse, Mark E. J.; Morabito, Stefano; Gally, David L.

    2012-01-01

    Lytic or lysogenic infections by bacteriophages drive the evolution of enteric bacteria. Enterohemorrhagic Escherichia coli (EHEC) have recently emerged as a significant zoonotic infection of humans with the main serotypes carried by ruminants. Typical EHEC strains are defined by the expression of a type III secretion (T3S) system, the production of Shiga toxins (Stx) and association with specific clinical symptoms. The genes for Stx are present on lambdoid bacteriophages integrated into the E. coli genome. Phage type (PT) 21/28 is the most prevalent strain type linked with human EHEC infections in the United Kingdom and is more likely to be associated with cattle shedding high levels of the organism than PT32 strains. In this study we have demonstrated that the majority (90%) of PT 21/28 strains contain both Stx2 and Stx2c phages, irrespective of source. This is in contrast to PT 32 strains for which only a minority of strains contain both Stx2 and 2c phages (28%). PT21/28 strains had a lower median level of T3S compared to PT32 strains and so the relationship between Stx phage lysogeny and T3S was investigated. Deletion of Stx2 phages from EHEC strains increased the level of T3S whereas lysogeny decreased T3S. This regulation was confirmed in an E. coli K12 background transduced with a marked Stx2 phage followed by measurement of a T3S reporter controlled by induced levels of the LEE-encoded regulator (Ler). The presence of an integrated Stx2 phage was shown to repress Ler induction of LEE1 and this regulation involved the CII phage regulator. This repression could be relieved by ectopic expression of a cognate CI regulator. A model is proposed in which Stx2-encoding bacteriophages regulate T3S to co-ordinate epithelial cell colonisation that is promoted by Stx and secreted effector proteins. PMID:22615557

  16. Induction of genetic recombination in the lambda bacteriophage by ultraviolet radiation of the Escherichia Coli cells

    International Nuclear Information System (INIS)

    Alcantara D, D.

    1986-12-01

    In this work there are reported the results that show that although the stimulation of the recombination of the Lambda bacteriophage, by UV irradiation of the cells of Escherichia Coli, it looks to be the result of the high expression of the functions of the SOS system, doesn't keep some relationship with the high concentration of protein reached RecA. (Author)

  17. Lytic bacteriophages reduce Escherichia coli O157: H7 on fresh cut lettuce introduced through cross-contamination.

    Science.gov (United States)

    Ferguson, Sean; Roberts, Cheryl; Handy, Eric; Sharma, Manan

    2013-01-01

    The role of lytic bacteriophages in preventing cross contamination of produce has not been evaluated. A cocktail of three lytic phages specific for E. coli O157:H7 (EcoShield™) or a control (phosphate buffered saline, PBS) was applied to lettuce by either; (1) immersion of lettuce in 500 ml of EcoShield™ 8.3 log PFU/ml or 9.8 log PFU/ml for up to 2 min before inoculation with E. coli O157:H7; (2) spray-application of EcoShield™ (9.3 log PFU/ml) to lettuce after inoculation with E. coli O157:H7 (4.10 CFU/cm 2 ) following exposure to 50 μg/ml chlorine for 30 sec. After immersion studies, lettuce was spot-inoculated with E. coli O157:H7 (2.38 CFU/cm 2 ). Phage-treated, inoculated lettuce pieces were stored at 4°C for and analyzed for E. coli O157:H7 populations for up to 7 d. Immersion of lettuce in 9.8 log PFU/ml EcoShield™ for 2 min significantly (p PFU/ml) resulted in the deposition of high concentrations (7.8 log log PFU/cm 2 ) of bacteriophages on the surface of fresh cut lettuce, potentially contributing to the efficacy of the lytic phages on lettuce. Spraying phages on to inoculated fresh cut lettuce after being washed in hypochlorite solution was significantly more effective in reducing E. coli O157:H7 populations (2.22 log CFU/cm 2 ) on day 0 compared with control treatments (4.10 log CFU/cm 2 ). Both immersion and spray treatments provided protection from E. coli O157:H7 contamination on lettuce, but spray application of lytic bacteriophages to lettuce was more effective in immediately reducing E. coli O157:H7 populations fresh cut lettuce.

  18. Isolation and characterization of polyvalent bacteriophages infecting multi drug resistant Salmonella serovars isolated from broilers in Egypt.

    Science.gov (United States)

    Mahmoud, Mayada; Askora, Ahmed; Barakat, Ahmed Barakat; Rabie, Omar El-Farouk; Hassan, Sayed Emam

    2018-02-02

    In this study, we isolated and characterized three phages named as Salmacey1, Salmacey2 and Salmacey3, infecting multi drug resistant Salmonella serovars isolated from broilers in Egypt. The most prevalent Salmonella serovars were S. typhimurium, S. enteritidis, and S. kentucky. All these Salmonella serovars were found to be resistant to more than two of the ten antimicrobial agents tested. Only S. kentucky was found to be resistant to seven antimicrobial agents. Examination of these phage particles by transmission electron microscopy (TEM), demonstrated that two phages (Salmacey1, Salmacey2) were found to belong to family Siphoviridae, and Salmacey3 was assigned to the family Myoviridae. The results of host range assay revealed that these bacteriophages were polyvalent and thus capable of infecting four strains of Salmonella serovars and Citrobacter freundii. Moreover, the two phages (Salmacey1, Salmacey2) had a lytic effect on Enterobacter cloacae and Salmacey3 was able to infect E. coli. All phages could not infect S. para Typhi, Staphylococus aureus and Bacillus cereus. One-step growth curves of bacteriophages revealed that siphovirus phages (Salmacey1, Salmacey2) have burst size (80 and 90pfu per infected cell with latent period 35min and 40min respectively), and for the myovirus Salmacey3 had a burst size 110pfu per infected cell with latent period 60min. Molecular analyses indicated that these phages contained double-stranded DNA genomes. The lytic activity of the phages against the most multidrug resistant serovars S. kentucky as host strain was evaluated. The result showed that these bacteriophages were able to completely stop the growth of S. kentucky in vitro. These results suggest that phages have a high potential for phage application to control Salmonella serovars isolated from broilers in Egypt. Copyright © 2017. Published by Elsevier B.V.

  19. Feasibility of using a bacteriophage-based structural color sensor for screening the geographical origins of agricultural products.

    Science.gov (United States)

    Seol, Daun; Moon, Jong-Sik; Lee, Yujin; Han, Jiye; Jang, Daeil; Kang, Dong-Jin; Moon, Jiyoung; Jang, Eunjin; Oh, Jin-Woo; Chung, Hoeil

    2018-05-15

    An M13 bacteriophage-based color sensor, which can change its structural color upon interaction with a gaseous molecule, was evaluated as a screening tool for the discrimination of the geographical origins of three different agricultural products (garlic, onion, and perilla). Exposure of the color sensor to sample odors induced the self-assembled M13 bacteriophage bundles to swell by the interaction of amino acid residues (repeating units of four glutamates) on the bacteriophage with the odor components, resulting in a change in the structural color of the sensor. When the sensor was exposed to the odors of garlic and onion samples, the RGB color changes were considerable because of the strong interactions of the odor components such as disulfides with the glutamate residues on the sensor. Although the patterns of the color variations were generally similar between the domestic and imported samples, some degrees of dissimilarities in their intensities were also observed. Although the magnitude of color change decreased for perilla, the color change patterns between the two groups were somewhat different. With the acquired RGB data, a support vector machine was employed to distinguish the domestic and imported samples, and the resulting accuracies in the measurements of garlic, onion, and perilla samples were 94.1, 88.7, and 91.6%, respectively. The differences in the concentrations of the odor components between both groups and/or the presence of specific components exclusively in the odor of one group allowed the color sensor-based discrimination. The demonstrated color sensor was thus shown to be a potentially versatile and simple as an on-site screening tool. Strategies able to further improve the sensor performance were also discussed. Copyright © 2018. Published by Elsevier B.V.

  20. Characterization of a bacteriophage T4 mutant lacking DNA-dependent ATPase

    International Nuclear Information System (INIS)

    Behme, M.T.; Ebisuzaki, K.

    1975-01-01

    A DNA-dependent ATPase has previously been purified from bacteriophage T4-infected Escherichia coli. A mutant phage strain lacking this enzyme has been isolated and characterized. Although the mutant strain produced no detectable DNA-dependent ATPase, growth properties were not affected. Burst sizes were similar for the mutant phage and T4D in polAl, recB, recC, uvrA, uvrB, uvrC, and various DNA-negative E. coli. UV sensitivity and genetic recombination were normal in a variety of E. coli hosts. Mapping data indicate that the genetic locus controlling the mutant occurs near gene 56. The nonessential nature of this gene is discussed

  1. Application of bacteriophages to reduce biofilms formed by hydrogen sulfide producing bacteria on surfaces in a rendering plant.

    Science.gov (United States)

    Gong, Chao; Jiang, Xiuping

    2015-08-01

    Hydrogen sulfide producing bacteria (SPB) in raw animal by-products are likely to grow and form biofilms in the rendering processing environments, resulting in the release of harmful hydrogen sulfide (H2S) gas. The objective of this study was to reduce SPB biofilms formed on different surfaces typically found in rendering plants by applying a bacteriophage cocktail. Using a 96-well microplate method, we determined that 3 SPB strains of Citrobacter freundii and Hafnia alvei are strong biofilm formers. Application of 9 bacteriophages (10(7) PFU/mL) from families of Siphoviridae and Myoviridae resulted in a 33%-70% reduction of biofilm formation by each SPB strain. On stainless steel and plastic templates, phage treatment (10(8) PFU/mL) reduced the attached cells of a mixed SPB culture (no biofilm) by 2.3 and 2.7 log CFU/cm(2) within 6 h at 30 °C, respectively, as compared with 2 and 1.5 log CFU/cm(2) reductions of SPB biofilms within 6 h at 30 °C. Phage treatment was also applied to indigenous SPB biofilms formed on the environmental surface, stainless steel, high-density polyethylene plastic, and rubber templates in a rendering plant. With phage treatment (10(9) PFU/mL), SPB biofilms were reduced by 0.7-1.4, 0.3-0.6, and 0.2-0.6 log CFU/cm(2) in spring, summer, and fall trials, respectively. Our study demonstrated that bacteriophages could effectively reduce the selected SPB strains either attached to or in formed biofilms on various surfaces and could to some extent reduce the indigenous SPB biofilms on the surfaces in the rendering environment.

  2. Examining a DNA Replication Requirement for Bacteriophage λ Red- and Rac Prophage RecET-Promoted Recombination in Escherichia coli.

    Science.gov (United States)

    Thomason, Lynn C; Costantino, Nina; Court, Donald L

    2016-09-13

    Recombineering, in vivo genetic engineering with bacteriophage homologous recombination systems, is a powerful technique for making genetic modifications in bacteria. Two systems widely used in Escherichia coli are the Red system from phage λ and RecET from the defective Rac prophage. We investigated the in vivo dependence of recombineering on DNA replication of the recombining substrate using plasmid targets. For λ Red recombination, when DNA replication of a circular target plasmid is prevented, recombination with single-stranded DNA oligonucleotides is greatly reduced compared to that under replicating conditions. For RecET recombination, when DNA replication of the targeted plasmid is prevented, the recombination frequency is also reduced, to a level identical to that seen for the Red system in the absence of replication. The very low level of oligonucleotide recombination observed in the absence of any phage recombination functions is the same in the presence or absence of DNA replication. In contrast, both the Red and RecET systems recombine a nonreplicating linear dimer plasmid with high efficiency to yield a circular monomer. Therefore, the DNA replication requirement is substrate dependent. Our data are consistent with recombination by both the Red and RecET systems occurring predominately by single-strand annealing rather than by strand invasion. Bacteriophage homologous recombination systems are widely used for in vivo genetic engineering in bacteria. Single- or double-stranded linear DNA substrates containing short flanking homologies to chromosome targets are used to generate precise and accurate genetic modifications when introduced into bacteria expressing phage recombinases. Understanding the molecular mechanism of these recombination systems will facilitate improvements in the technology. Here, two phage-specific systems are shown to require exposure of complementary single-strand homologous targets for efficient recombination; these single

  3. Investigation of the bacteriophage community in induced lysates of undefined mesophilic mixed-strain DL-cultures using classical and metagenomic approaches

    DEFF Research Database (Denmark)

    Muhammed, Musemma K.; Olsen, Mette L.; Kot, Witold

    2018-01-01

    To investigate the notion that starter cultures can be a reservoir of bacteriophages (phages) in the dairy environment, strains of three DL-starters (undefined mesophilic mixed-strain starters containing Lactococcus lactis subsp. lactis biovar. diacetylactis and Leuconostoc species) were selected...

  4. Repair capability of mammalian cell fractions demonstrated using infectivity of bacteriophage DNA

    International Nuclear Information System (INIS)

    Lai, S.P.; Lytle, C.D.; Benane, S.G.

    1976-01-01

    Extracts of Potoroo kidney cells (PtK2) were examined for ability to provide a repair function in vitro. The biological activity (infectivity) of uv-irradiated replicative form (RF) DNA of bacteriophage phiX174 was restored during incubation of the DNA with a nuclear extract but not with a cytoplasmic extract. The infectivity of the RF-DNA was determined in spheroplasts of E. coli C/sub s/, which is HCR - . This system for biological assay of uv-irradiated DNA repaired in vitro may be used to complement biochemical and biophysical investigations of molecular repair mechanisms in mammalian cells

  5. Direct interaction of the bacteriophage SPP1 packaging ATPase with the portal protein.

    Science.gov (United States)

    Oliveira, Leonor; Cuervo, Ana; Tavares, Paulo

    2010-03-05

    DNA packaging in tailed bacteriophages and other viruses requires assembly of a complex molecular machine at a specific vertex of the procapsid. This machine is composed of the portal protein that provides a tunnel for DNA entry, an ATPase that fuels DNA translocation (large terminase subunit), and most frequently, a small terminase subunit. Here we characterized the interaction between the terminase ATPase subunit of bacteriophage SPP1 (gp2) and the procapsid portal vertex. We found, by affinity pulldown assays with purified proteins, that gp2 interacts with the portal protein, gp6, independently of the terminase small subunit gp1, DNA, or ATP. The gp2-procapsid interaction via the portal protein depends on gp2 concentration and requires the presence of divalent cations. Competition experiments showed that isolated gp6 can only inhibit gp2-procapsid interactions and DNA packaging at gp6:procapsid molar ratios above 10-fold. Assays with gp6 carrying mutations in distinct regions of its structure that affect the portal-induced stimulation of ATPase and DNA packaging revealed that none of these mutations impedes gp2-gp6 binding. Our results demonstrate that the SPP1 packaging ATPase binds directly to the portal and that the interaction is stronger with the portal embedded in procapsids. Identification of mutations in gp6 that allow for assembly of the ATPase-portal complex but impair DNA packaging support an intricate cross-talk between the two proteins for activity of the DNA translocation motor.

  6. In vivo studies of genomic packaging in the dsRNA bacteriophage Φ8

    Directory of Open Access Journals (Sweden)

    Mindich Leonard

    2005-03-01

    Full Text Available Abstract Background Φ8 is a bacteriophage containing a genome of three segments of double-stranded RNA inside a polyhedral capsid enveloped in a lipid-containing membrane. Plus strand RNA binds and is packaged by empty procapsids. Whereas Φ6, another member of the Cystoviridae, shows high stringency, serial dependence and precision in its genomic packaging in vitro and in vivo, Φ8 packaging is more flexible. Unique sequences (pac near the 5' ends of plus strands are necessary and sufficient for Φ6 genomic packaging and the RNA binding sites are located on P1, the major structural protein of the procapsid. Results In this paper the boundaries of the Φ8 pac sequences have been explored by testing the in vivo packaging efficacy of transcripts containing deletions or changes in the RNA sequences. The pac sequences have been localized to the 5' untranslated regions of the viral transcripts. Major changes in the pac sequences are either tolerated or ameliorated by suppressor mutations in the RNA sequence. Changes in the genomic packaging program can be established as a result of mutations in P1, the major structural protein of the procapsid and the determinant of RNA binding specificity. Conclusion Although Φ8 is distantly related to bacteriophage Φ6, and does not show sequence similarity, it has a similar genomic packaging program. This program, however, is less stringent than that of Φ6.

  7. Regulation of gene expression in Escherichia coli and its bacteriophage

    International Nuclear Information System (INIS)

    Higgins, C.F.

    1986-01-01

    This chapter reviews the study of prokaryotic gene expression beginning with a look at the regulation of the lactose operon and the mechanism of attenuation in the tryptophan operon to the more recent development of recombinant DNA technology. The chapter deals almost entirely with escherichia coli and its bacteriophage. The only experimental technique which the authors explore in some detail is the construction and use of gene and operon fusions which have revolutionized the study of gene expression. Various mechanisms by which E. Coli regulate the cellular levels of individual messenger-RNA species are described. Translational regulation of the cellular levels of messenger-RNA include signals encoded within the messenger-RNA molecule itself and regulatory molecules which interact with the messenger-RNA and alter it translational efficiency

  8. Comparative Prevalence of Immune Evasion Complex Genes Associated with β-Hemolysin Converting Bacteriophages in MRSA ST5 Isolates from Swine, Swine Facilities, Humans with Swine Contact, and Humans with No Swine Contact.

    Directory of Open Access Journals (Sweden)

    Samantha J Hau

    Full Text Available Livestock associated methicillin-resistant Staphylococcus aureus (LA-MRSA draws concern from the public health community because in some countries these organisms may represent the largest reservoir of MRSA outside hospital settings. Recent studies indicate LA-MRSA strains from swine are more genetically diverse than the first reported sequence type ST398. In the US, a diverse population of LA-MRSA is found including organisms of the ST398, ST9, and ST5 lineages. Occurrence of ST5 MRSA in swine is of particular concern since ST5 is among the most prevalent lineages causing clinical infections in humans. The prominence of ST5 in clinical disease is believed to result from acquisition of bacteriophages containing virulence or host-adapted genes including the immune-evasion cluster (IEC genes carried by β-hemolysin converting bacteriophages, whose absence in LA-MRSA ST398 is thought to contribute to reduced rates of human infection and transmission associated with this lineage. The goal of this study was to investigate the prevalence of IEC genes associated with β-hemolysin converting bacteriophages in MRSA ST5 isolates obtained from agricultural sources, including swine, swine facilities, and humans with short- or long-term swine exposure. To gain a broader perspective, the prevalence of these genes in LA-MRSA ST5 strains was compared to the prevalence in clinical MRSA ST5 strains from humans with no known exposure to swine. IEC genes were not present in any of the tested MRSA ST5 strains from agricultural sources and the β-hemolysin gene was intact in these strains, indicating the bacteriophage's absence. In contrast, the prevalence of the β-hemolysin converting bacteriophage in MRSA ST5 strains from humans with no exposure to swine was 90.4%. The absence of β-hemolysin converting bacteriophage in LA-MRSA ST5 isolates is consistent with previous reports evaluating ST398 strains and provides genetic evidence indicating LA-MRSA ST5 isolates

  9. phiGENOME: an integrative navigation throughout bacteriophage genomes.

    Science.gov (United States)

    Stano, Matej; Klucar, Lubos

    2011-11-01

    phiGENOME is a web-based genome browser generating dynamic and interactive graphical representation of phage genomes stored in the phiSITE, database of gene regulation in bacteriophages. phiGENOME is an integral part of the phiSITE web portal (http://www.phisite.org/phigenome) and it was optimised for visualisation of phage genomes with the emphasis on the gene regulatory elements. phiGENOME consists of three components: (i) genome map viewer built using Adobe Flash technology, providing dynamic and interactive graphical display of phage genomes; (ii) sequence browser based on precisely formatted HTML tags, providing detailed exploration of genome features on the sequence level and (iii) regulation illustrator, based on Scalable Vector Graphics (SVG) and designed for graphical representation of gene regulations. Bringing 542 complete genome sequences accompanied with their rich annotations and references, makes phiGENOME a unique information resource in the field of phage genomics. Copyright © 2011 Elsevier Inc. All rights reserved.

  10. Downstream processing and chromatography based analytical methods for production of vaccines, gene therapy vectors, and bacteriophages

    Science.gov (United States)

    Kramberger, Petra; Urbas, Lidija; Štrancar, Aleš

    2015-01-01

    Downstream processing of nanoplexes (viruses, virus-like particles, bacteriophages) is characterized by complexity of the starting material, number of purification methods to choose from, regulations that are setting the frame for the final product and analytical methods for upstream and downstream monitoring. This review gives an overview on the nanoplex downstream challenges and chromatography based analytical methods for efficient monitoring of the nanoplex production. PMID:25751122

  11. Bacteriophage-Derived Peptidase CHAPK Eliminates and Prevents Staphylococcal Biofilms

    Directory of Open Access Journals (Sweden)

    Mark Fenton

    2013-01-01

    Full Text Available New antibacterial agents are urgently needed for the elimination of biofilm-forming bacteria that are highly resistant to traditional antimicrobial agents. Proliferation of such bacteria can lead to significant economic losses in the agri-food sector. This study demonstrates the potential of the bacteriophage-derived peptidase, CHAPK, as a biocidal agent for the rapid disruption of biofilm-forming staphylococci, commonly associated with bovine mastitis. Purified CHAPK applied to biofilms of Staphylococcus aureus DPC5246 completely eliminated the staphylococcal biofilms within 4 h. In addition, CHAPK was able to prevent biofilm formation by this strain. The CHAPK lysin also reduced S. aureus in a skin decolonization model. Our data demonstrates the potential of CHAPK as a biocidal agent for prevention and treatment of biofilm-associated staphylococcal infections or as a decontaminating agent in the food and healthcare sectors.

  12. Genetic and biochemical studies of the lipid-containing bacteriophage PR4

    International Nuclear Information System (INIS)

    Vanden Boom, T.J.

    1989-01-01

    Bacteriophage PR4 is a lipid-containing bacterial virus able to infect Escherichia coli and Salmonella typhimurium. The icosahedral virion consists of an external protein capsid layer which surrounds a membrane vesicle enclosed ds DNA genome. The author has analyzed the time course of phage PR4 protein synthesis and have identified at least 34 proteins present in phage infected cells not detected in uninfected control cultures. In addition, he has isolated a more extensive set of conditional-lethal nonsense mutants of this virus. This collection of mutants permitted the identification of seven additional phage PR4 gene products, including the terminal genome protein and an accessory lytic factor. The present collection of phage PR4 mutants has been assigned to 19 distinct genetic groups on the basis of genetic complementation tests and sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis of the proteins produced in mutant-infected UV-irradiated cells. A restriction endonuclease map of the phage PR4 genome was constructed which includes 59 sites for ten restriction endonucleases. In addition, he has constructed a collection of recombinant plasmids containing subgenomic DNA fragments of bacteriophage PR4. He has used this collection of plasmids to generate a physical-genetic map of the PR4 genome. The physical-genetic map localizes mutations in 13 phage PR4 genetic groups on the viral DNA molecule. To investigate the role of phosphatidylglycerol (PG) in phage assembly and infectivity, he propagated PR4 on an E. coli mutant defective in PG synthesis. The PG content of phage PR4 grown on the mutant host accounted for 0.4% of the total viral phospholipids, representing a 90-fold decrease in PG relative to the PG content of phage grown on a wild type host

  13. Evolution of the Quorum network and the mobilome (plasmids and bacteriophages) in clinical strains of Acinetobacter baumannii during a decade.

    Science.gov (United States)

    López, M; Rueda, A; Florido, J P; Blasco, L; Fernández-García, L; Trastoy, R; Fernández-Cuenca, F; Martínez-Martínez, L; Vila, J; Pascual, A; Bou, G; Tomas, M

    2018-02-06

    In this study, we compared eighteen clinical strains of A. baumannii belonging to the ST-2 clone and isolated from patients in the same intensive care unit (ICU) in 2000 (9 strains referred to collectively as Ab_GEIH-2000) and 2010 (9 strains referred to collectively as Ab_GEIH-2010), during the GEIH-REIPI project (Umbrella BioProject PRJNA422585). We observed two main molecular differences between the Ab_GEIH-2010 and the Ab_GEIH-2000 collections, acquired over the course of the decade long sampling interval and involving the mobilome: i) a plasmid harbouring genes for bla OXA 24/40 ß-lactamase and abKA/abkB proteins of a toxin-antitoxin system; and ii) two temperate bacteriophages, Ab105-1ϕ (63 proteins) and Ab105-2ϕ (93 proteins), containing important viral defence proteins. Moreover, all Ab_GEIH-2010 strains contained a Quorum functional network of Quorum Sensing (QS) and Quorum Quenching (QQ) mechanisms, including a new QQ enzyme, AidA, which acts as a bacterial defence mechanism against the exogenous 3-oxo-C12-HSL. Interestingly, the infective capacity of the bacteriophages isolated in this study (Ab105-1ϕ and Ab105-2ϕ) was higher in the Ab_GEIH-2010 strains (carrying a functional Quorum network) than in the Ab_GEIH-2000 strains (carrying a deficient Quorum network), in which the bacteriophages showed little or no infectivity. This is the first study about the evolution of the Quorum network and the mobilome in clinical strains of Acinetobacter baumannii during a decade.

  14. Effects of Lactobacillus Probiotic, P22 Bacteriophage and Salmonella Typhimurium on the Heterophilic Burst Activity of Broiler Chickens

    Directory of Open Access Journals (Sweden)

    GA Marietto-Gonçalves

    2014-09-01

    Full Text Available Due to the constant evolution of industrial poultry production and the global emergence of bacterial resistance to antibiotics there has been an increasing interest in alternatives for the treatment of poultry salmonellosis, such as phage therapy and probiotics. The present study evaluated the effects of the oral administration of the bacteriophage P22 and of a probiotic, consisting of four Lactobacillus species, on the level of circulating heterophils containing a superoxide anion of one-day-old broilers challenged with Salmonella Typhimurium for seven days. It was concluded that the treatment with a probiotic with lactobacilli of broilers experimentally infected with Salmonella spp eliminates this pathogen by increasing the circulating levels of reactive heterophils. When chicks are treated with a probiotic and a bacteriophage, the agent is eliminated with no changes in circulating reactive heterophil counts. It is also concluded that the heterophils of day-old chicks are not capable of producing superoxide anion. However, this capacity is detected after 48 h of life, indicating that heterophils mature as birds age.

  15. Draft genome sequences of three virulent Streptococcus thermophilus bacteriophages isolated from the dairy environment in the Veneto region of Italy

    DEFF Research Database (Denmark)

    Duarte, Viní­cius da Silva; Giaretta, Sabrina; Treu, Laura

    2018-01-01

    Streptococcus thermophilus, a very important dairy species, is constantly threatened by phage infection. We report the genome sequences of three S. thermophilus bacteriophages isolated from a dairy environment in the Veneto region of Italy. These sequences will be used for the development of new ...

  16. E-Hitz: a word frequency list and a program for deriving psycholinguistic statistics in an agglutinative language (Basque).

    Science.gov (United States)

    Perea, Manuel; Urkia, Miriam; Davis, Colin J; Agirre, Ainhoa; Laseka, Edurne; Carreiras, Manuel

    2006-11-01

    We describe a Windows program that enables users to obtain a broad range of statistics concerning the properties of word and nonword stimuli in an agglutinative language (Basque), including measures of word frequency (at the whole-word and lemma levels), bigram and biphone frequency, orthographic similarity, orthographic and phonological structure, and syllable-based measures. It is designed for use by researchers in psycholinguistics, particularly those concerned with recognition of isolated words and morphology. In addition to providing standard orthographic and phonological neighborhood measures, the program can be used to obtain information about other forms of orthographic similarity, such as transposed-letter similarity and embedded-word similarity. It is available free of charge from www .uv.es/mperea/E-Hitz.zip.

  17. Structure and assembly of bacteriophage T4 head

    Directory of Open Access Journals (Sweden)

    Black Lindsay W

    2010-12-01

    Full Text Available Abstract The bacteriophage T4 capsid is an elongated icosahedron, 120 nm long and 86 nm wide, and is built with three essential proteins; gp23*, which forms the hexagonal capsid lattice, gp24*, which forms pentamers at eleven of the twelve vertices, and gp20, which forms the unique dodecameric portal vertex through which DNA enters during packaging and exits during infection. The past twenty years of research has greatly elevated the understanding of phage T4 head assembly and DNA packaging. The atomic structure of gp24 has been determined. A structural model built for gp23 using its similarity to gp24 showed that the phage T4 major capsid protein has the same fold as that found in phage HK97 and several other icosahedral bacteriophages. Folding of gp23 requires the assistance of two chaperones, the E. coli chaperone GroEL and the phage coded gp23-specific chaperone, gp31. The capsid also contains two non-essential outer capsid proteins, Hoc and Soc, which decorate the capsid surface. The structure of Soc shows two capsid binding sites which, through binding to adjacent gp23 subunits, reinforce the capsid structure. Hoc and Soc have been extensively used in bipartite peptide display libraries and to display pathogen antigens including those from HIV, Neisseria meningitides, Bacillus anthracis, and FMDV. The structure of Ip1*, one of the components of the core, has been determined, which provided insights on how IPs protect T4 genome against the E. coli nucleases that degrade hydroxymethylated and glycosylated T4 DNA. Extensive mutagenesis combined with the atomic structures of the DNA packaging/terminase proteins gp16 and gp17 elucidated the ATPase and nuclease functional motifs involved in DNA translocation and headful DNA cutting. Cryo-EM structure of the T4 packaging machine showed a pentameric motor assembled with gp17 subunits on the portal vertex. Single molecule optical tweezers and fluorescence studies showed that the T4 motor packages

  18. Prostaglandin-mediated recovery from bacteremia delays larval development in fall armyworm, Spodoptera frugiperda.

    Science.gov (United States)

    Zhang, Lei; Ringbauer, Joseph A; Goodman, Cynthia L; Reall, Tamra; Jiang, Xing-Fu; Stanley, David

    2018-04-01

    Insect immunity includes a surveillance system that detects and signals infections, coupled with hemocytic and humoral immune functions. These functions are signaled and coordinated by several biochemicals, including biogenic amines, insect cytokines, peptides, and prostaglandins (PGs). The actions of these mediators are coordinated within cells by various forms of cross-talk among the signaling systems and they result in effective reactions to infection. While this is well understood, we lack information on how immune-mediated recovery influences subsequent juvenile development in surviving insects. We investigated this point by posing the hypothesis that PG signaling is necessary for larval recovery, although the recovery imposes biological costs, registered in developmental delays and failures in surviving individuals. Here, we report that nodulation responses to infections by the bacterium, Serratia marcescens, increased over time up to 5 h postinfection, with no further nodulation; it increased in a linear manner with increasing bacterial dosages. Larval survivorship decreased with increasing bacterial doses. Treating larvae with the PG-biosynthesis inhibitor, indomethacin, led to sharply decreased nodulation reactions to infection, which were rescued in larvae cotreated with indomethacin and the PG-precursor, arachidonic acid. Although nodulation was fully rescued, all bacterial challenged larvae suffered reduced survivorship compared to controls. Bacterial infection led to reduced developmental rates in larvae, but not pupae. Adult emergence from pupae that developed from experimental larvae was also decreased. Taken together, our data potently bolster our hypothesis. © 2018 Wiley Periodicals, Inc.

  19. Yersinia enterocolitica-Specific Infection by Bacteriophages TG1 and ϕR1-RT Is Dependent on Temperature-Regulated Expression of the Phage Host Receptor OmpF.

    Science.gov (United States)

    Leon-Velarde, Carlos G; Happonen, Lotta; Pajunen, Maria; Leskinen, Katarzyna; Kropinski, Andrew M; Mattinen, Laura; Rajtor, Monika; Zur, Joanna; Smith, Darren; Chen, Shu; Nawaz, Ayesha; Johnson, Roger P; Odumeru, Joseph A; Griffiths, Mansel W; Skurnik, Mikael

    2016-09-01

    Bacteriophages present huge potential both as a resource for developing novel tools for bacterial diagnostics and for use in phage therapy. This potential is also valid for bacteriophages specific for Yersinia enterocolitica To increase our knowledge of Y. enterocolitica-specific phages, we characterized two novel yersiniophages. The genomes of the bacteriophages vB_YenM_TG1 (TG1) and vB_YenM_ϕR1-RT (ϕR1-RT), isolated from pig manure in Canada and from sewage in Finland, consist of linear double-stranded DNA of 162,101 and 168,809 bp, respectively. Their genomes comprise 262 putative coding sequences and 4 tRNA genes and share 91% overall nucleotide identity. Based on phylogenetic analyses of their whole-genome sequences and large terminase subunit protein sequences, a genus named Tg1virus within the family Myoviridae is proposed, with TG1 and ϕR1-RT (R1RT in the ICTV database) as member species. These bacteriophages exhibit a host range restricted to Y. enterocolitica and display lytic activity against the epidemiologically significant serotypes O:3, O:5,27, and O:9 at and below 25°C. Adsorption analyses of lipopolysaccharide (LPS) and OmpF mutants demonstrate that these phages use both the LPS inner core heptosyl residues and the outer membrane protein OmpF as phage receptors. Based on RNA sequencing and quantitative proteomics, we also demonstrate that temperature-dependent infection is due to strong repression of OmpF at 37°C. In addition, ϕR1-RT was shown to be able to enter into a pseudolysogenic state. Together, this work provides further insight into phage-host cell interactions by highlighting the importance of understanding underlying factors which may affect the abundance of phage host receptors on the cell surface. Only a small number of bacteriophages infecting Y. enterocolitica, the predominant causative agent of yersiniosis, have been previously described. Here, two newly isolated Y. enterocolitica phages were studied in detail, with the aim of

  20. Biocontrol of Escherichia coli O157:H7 on fresh-cut spinach and lettuce using a bacteriophage cocktail

    Science.gov (United States)

    The effect of an E. coli O157:H7-specific bacteriophage cocktail (EcoShield™) was evaluated against nalidixic acid resistant (NalR) E. coli O157:H7 strains in either a) laboratory medium or b) on leafy greens. Laboratory medium cultures were inoculated with 5 log CFU/ml and treated with 7 log PFU/ml...

  1. Prevalence of agglutinating antibodies to Toxoplasma gondii and Sarcocystis neurona in beavers (Castor canadensis) from Massachusetts

    Science.gov (United States)

    Jordan, C.N.; Kaur, T.; Koenen, K.; DeStefano, S.; Zajac, A.M.; Lindsay, D.S.

    2005-01-01

    The present study examined the seroprevalence of Toxoplasma gondii and Sarcocystls neurona in a population of beavers (Castor canadensis) from Massachusetts. Sixty-two blood samples were collected during the field seasons over 3 consecutive years from different animals. Blood was collected onto filter paper and shipped to the Department of Biomedical Sciences, Virginia Tech, Blacksburg, Virginia, for parasite testing. The samples were tested at dilutions of 1:25, 1:50, and 1:100 against each parasite antigen by modified agglutination tests to determine whether antibodies to either parasite were present in the blood. Six of 62 samples (10%) were positive for T. gondii, with 2 samples having titers of 1:25 and 4 having titers of 1:50. Four of 62 samples (6%) were positive for S. neurona, with 2 samples having titers of 1:25 and 2 having titers of 1:50. ?? American Society of Pathologists 2005.

  2. RNA secondary structures of the bacteriophage phi6 packaging regions.

    Science.gov (United States)

    Pirttimaa, M J; Bamford, D H

    2000-06-01

    Bacteriophage phi6 genome consists of three segments of double-stranded RNA. During maturation, single-stranded copies of these segments are packaged into preformed polymerase complex particles. Only phi6 RNA is packaged, and each particle contains only one copy of each segment. An in vitro packaging and replication assay has been developed for phi6, and the packaging signals (pac sites) have been mapped to the 5' ends of the RNA segments. In this study, we propose secondary structure models for the pac sites of phi6 single-stranded RNA segments. Our models accommodate data from structure-specific chemical modifications, free energy minimizations, and phylogenetic comparisons. Previously reported pac site deletion studies are also discussed. Each pac site possesses a unique architecture, that, however, contains common structural elements.

  3. Cooperativity Leads to Temporally-Correlated Fluctuations in the Bacteriophage Lambda Genetic Switch

    Directory of Open Access Journals (Sweden)

    Jacob Quinn Shenker

    2015-04-01

    Full Text Available Cooperative interactions are widespread in biochemical networks, providing the nonlinear response that underlies behavior such as ultrasensitivity and robust switching. We introduce a temporal correlation function—the conditional activity—to study the behavior of these phenomena. Applying it to the bistable genetic switch in bacteriophage lambda, we find that cooperative binding between binding sites on the prophage DNA lead to non-Markovian behavior, as quantified by the conditional activity. Previously, the conditional activity has been used to predict allosteric pathways in proteins; here, we show that it identifies the rare unbinding events which underlie induction from lysogeny to lysis.

  4. Bacteriophages safely reduce Salmonella contamination in pet food and raw pet food ingredients.

    Science.gov (United States)

    Soffer, Nitzan; Abuladze, Tamar; Woolston, Joelle; Li, Manrong; Hanna, Leigh Farris; Heyse, Serena; Charbonneau, Duane; Sulakvelidze, Alexander

    2016-01-01

    Contamination of pet food with Salmonella is a serious public health concern, and several disease outbreaks have recently occurred due to human exposure to Salmonella tainted pet food. The problem is especially challenging for raw pet foods (which include raw meats, seafood, fruits, and vegetables). These foods are becoming increasingly popular because of their nutritional qualities, but they are also more difficult to maintain Salmonella -free because they lack heat-treatment. Among various methods examined to improve the safety of pet foods (including raw pet food), one intriguing approach is to use bacteriophages to specifically kill Salmonella serotypes. At least 2 phage preparations (SalmoFresh® and Salmonelex™) targeting Salmonella are already FDA cleared for commercial applications to improve the safety of human foods. However, similar preparations are not yet available for pet food applications. Here, we report the results of evaluating one such preparation (SalmoLyse®) in reducing Salmonella levels in various raw pet food ingredients (chicken, tuna, turkey, cantaloupe, and lettuce). Application of SalmoLyse® in low (ca. 2-4×10 6 PFU/g) and standard (ca. 9×10 6 PFU/g) concentrations significantly ( P contamination in all raw foods examined compared to control treatments. When SalmoLyse®-treated (ca. 2×10 7 PFU/g) dry pet food was fed to cats and dogs, it did not trigger any deleterious side effects in the pets. Our data suggest that the bacteriophage cocktail lytic for Salmonella can significantly and safely reduce Salmonella contamination in various raw pet food ingredients.

  5. Optimizing Propagation of Staphylococcus aureus Infecting Bacteriophage vB_SauM-phiIPLA-RODI on Staphylococcus xylosus Using Response Surface Methodology.

    Science.gov (United States)

    González-Menéndez, Eva; Arroyo-López, Francisco Noé; Martínez, Beatriz; García, Pilar; Garrido-Fernández, Antonio; Rodríguez, Ana

    2018-03-27

    The use of bacteriophages for killing pathogenic bacteria is a feasible alternative to antibiotics and disinfectants. To obtain the large quantities of phages required for this application, large-scale production of bacteriophages must be optimized. This study aims to define conditions that maximize the phage yield of the virulent and polyvalent staphylococcal bacteriophage vB_SauM-phiIPLA-RODI in broth culture, using the food-grade species Staphylococcus xylosus as the host strain to reduce the risk of growing massive quantities of pathogenic bacteria and therefore, to ensure the safety of the final phage stock. The effect of four variables, namely initial bacterial concentration (5.66-8.40 log 10 colony-forming unit (CFU)/mL), initial phage concentration (5-8 log 10 plaque-forming unit (PFU)/mL), temperature (21-40 °C) and agitation (20-250 rpm), on phage yield (response) was studied by using response surface methodology (RSM). Successive experimental designs showed that agitation did not significantly impact phage yield, while temperature did have a significant effect, with 38 °C being the optimum for phage propagation. The results allowed the design of a model to describe phage yield as a function of the initial bacterial and phage concentrations at fixed agitation (135 rpm), and optimum temperature (38 °C). The maximum experimental phage yield obtained was 9.3 log 10 PFU/mL, while that predicted by the model under the optimized conditions (7.07 log 10 CFU/mL initial bacterial population and 6.00 log 10 PFU/mL initial phage titer) was 9.25 ± 0.30 log 10 PFU/mL, with the desirability of 0.96. This yield is comparable to that obtained when the phage was propagated on the original host, Staphylococcus aureus. Bacteriophage phiIPLA-RODI showed the same host range and very similar biofilm removal ability regardless of the staphylococcal species used for its propagation. The results presented in this study show the suitability of using a food-grade strain of S

  6. Optimizing Propagation of Staphylococcus aureus Infecting Bacteriophage vB_SauM-phiIPLA-RODI on Staphylococcus xylosus Using Response Surface Methodology

    Directory of Open Access Journals (Sweden)

    Eva González-Menéndez

    2018-03-01

    Full Text Available The use of bacteriophages for killing pathogenic bacteria is a feasible alternative to antibiotics and disinfectants. To obtain the large quantities of phages required for this application, large-scale production of bacteriophages must be optimized. This study aims to define conditions that maximize the phage yield of the virulent and polyvalent staphylococcal bacteriophage vB_SauM-phiIPLA-RODI in broth culture, using the food-grade species Staphylococcus xylosus as the host strain to reduce the risk of growing massive quantities of pathogenic bacteria and therefore, to ensure the safety of the final phage stock. The effect of four variables, namely initial bacterial concentration (5.66–8.40 log10 colony-forming unit (CFU/mL, initial phage concentration (5–8 log10 plaque-forming unit (PFU/mL, temperature (21–40 °C and agitation (20–250 rpm, on phage yield (response was studied by using response surface methodology (RSM. Successive experimental designs showed that agitation did not significantly impact phage yield, while temperature did have a significant effect, with 38 °C being the optimum for phage propagation. The results allowed the design of a model to describe phage yield as a function of the initial bacterial and phage concentrations at fixed agitation (135 rpm, and optimum temperature (38 °C. The maximum experimental phage yield obtained was 9.3 log10 PFU/mL, while that predicted by the model under the optimized conditions (7.07 log10 CFU/mL initial bacterial population and 6.00 log10 PFU/mL initial phage titer was 9.25 ± 0.30 log10 PFU/mL, with the desirability of 0.96. This yield is comparable to that obtained when the phage was propagated on the original host, Staphylococcus aureus. Bacteriophage phiIPLA-RODI showed the same host range and very similar biofilm removal ability regardless of the staphylococcal species used for its propagation. The results presented in this study show the suitability of using a food

  7. A cII-dependent promoter is located within the Q gene of bacteriophage lambda.

    OpenAIRE

    Hoopes, B C; McClure, W R

    1985-01-01

    We have found a cII-dependent promoter, PaQ, within the Q gene of bacteriophage lambda. Transcription experiments and abortive initiation assays performed in vitro showed that the promoter strength and the cII affinity of PaQ were comparable to the other cII-dependent lambda promoters, PE and PI. The location and leftward direction of PaQ suggests a possible role in the delay of lambda late-gene expression by cII protein, a phenomenon that has been called cII-dependent inhibition. We have con...

  8. Isolation, in vitro culture, ultrastructure study, and characterization by lectin-agglutination tests of Phytomonas isolated from tomatoes (Lycopersicon esculentum) and cherimoyas (Anona cherimolia) in southeastern Spain.

    Science.gov (United States)

    Sanchez-Moreno, M; Fernandez-Becerra, C; Mascaro, C; Rosales, M J; Dollet, M; Osuna, A

    1995-01-01

    Plants of Lycopersicon esculentum (grown in greenhouses) and Anona cherimolia cultivated in southeastern Spain were examined for the presence of trypanosomatid flagellates. Kinetoplastid protozoa were found in the fruits but not in the phloem or other plant tissues. Parasites were detected from the onset of fruiting. Isolates were detected from the onset of fruiting. Isolates were adapted to in vitro culturing in monophase media. The form and the structural organization was studied by scanning and transmission electron microscopy. The parasites showed an ultrastructural pattern similar to that of other species of the genus Phytomonas. In tomatoes experimentally inoculated with flagellates cultivated in vitro, we observed that the parasites did not lose their infectious capacity. Three strains of trypanosomatids of the genus Phytomonas, isolated from different species of Euphorbia (E. characias and E. hyssopifolia) and from Cocos nucifera, were compared with our isolates by lectin-agglutination tests. Our isolates were different from the two strains isolated from Euphorbia, but with this technique we could not differentiate our isolates from those of the coconut, nor could we differentiate between the isolates, their ultrastructural similarity together with their similar behavior in the lectin-agglutination test suggesting that these isolates have a common origin.

  9. HSP70 gene expression in Mytilus galloprovincialis hemocytes is triggered by moderate heat shock and Vibrio anguillarum, but not by V. splendidus or Micrococcus lysodeikticus.

    Science.gov (United States)

    Cellura, Cinzia; Toubiana, Mylène; Parrinello, Nicolo; Roch, Philippe

    2006-01-01

    Complete sequence of HSP70 cDNA from the mussel, Mytilus galloprovincialis was established before quantifying its expression following moderate heat shock or injection of heat-killed bacteria. HSP70 cDNA is comprised of 2378 bp including one ORF of 654 aa, with a predicted 70 bp 5'-UTR and a 343 bp 3'-UTR (GenBank, 18 Jan 05, AY861684). Alignment identity ranged from 89% for Crassostrea ariakensis to 72% for C. virginica. Curiously, HSP70 gene and cDNA sequences from M. galloprovincialis, deposited later (03 and 27 May), show only 73% identity with the present sequence. Meanwhile, characteristic motifs of the HSP70 family were located in conserved positions. Expression of HSP70 gene was quantified on circulating hemocyte mRNA using Q-PCR after RT using random hexaprimers. Housekeeping gene was 28S rRNA. Four stresses were applied: heat shock that consisted of immersing mussels for 90 min at 30 degrees C and returning them to 20 degrees C sea water, one injection of heat-killed Gram-negative bacteria, Vibrio splendidus LGP32, one injection of heat-killed Gram-negative bacteria Vibrio anguillarum, one injection of heat-killed Gram-positive bacteria Micrococcus lysodeikticus. We found no significant modification of 28S rRNA gene expression. Significant increase of 5.2 +/- 0.4 fold the ratio HSP70/28S rRNA was observed 6 h after heat shock and was maximum at 15 h (6.1 +/- 1.1), and still significant after 24 h (1.7 +/- 0.03). Similarly, injecting V. anguillarum resulted in a significant increase of 2.7 +/- 0.1 after 12 h. Expression was maximum after 48 h (5.2 +/- 0.05) and returned to baseline after 72 h. In contrast, injecting V. splendidus or M. lysodeikticus failed to significantly modulate HSP70 gene expression at least during the first 3 days post-injection. Consequently, mussel hemocytes appeared to discriminate between pathogenic and non-pathogenic Vibrios, as well as between Gram-negative and Gram-positive bacteria.

  10. Evaluation of latex agglutination test (KAtex) for early diagnosis of kala-azar.

    Science.gov (United States)

    Ahsan, M M; Islam, M N; Mollah, A H; Hoque, M A; Hossain, M A; Begum, Z; Islam, M T

    2010-07-01

    Kala-azar is one of the major public health problem in Bangladesh. But the diagnosis of the problem often is difficult, unusual and time consuming, a simple, noninvasive, easy to perform, reliable and rapid diagnostic test has been a long-felt need of the clinicians. Therefore, the present study was conducted to see the sensitivity and specificity of Latex Agglutination test (KAtex) to detect leishmanial antigen from urine of kala-azar cases. The study was carried out in the department of Paediatrics, Mymensingh Medical College and Hospital, Bangladesh during July to December, 2008. A total of 100 urine samples were collected of which 50 were confirmed kala-azar cases and 50 were age and sex matched controls. Out of 50 kala-azar cases 47 showed positive result of KAtex. The test was also positive in 01 out of 30 healthy controls. None of the febrile controls was positive by KAtex. The sensitivity, specificity, positive predictive value and negative predictive value of the test using presence of LD bodies in splenic and/or bone marrow aspirate as gold standard were 94%, 98%, 97.91% and 94.23% respectively. KAtex is simple, noninvasive, easy to perform, rapid and reliable test for diagnosing kala-azar in endemic area and useful for small, less equipped laboratories as well as for the laboratories with better facilities.

  11. Sequence and structural characterization of great salt lake bacteriophage CW02, a member of the T7-like supergroup.

    Science.gov (United States)

    Shen, Peter S; Domek, Matthew J; Sanz-García, Eduardo; Makaju, Aman; Taylor, Ryan M; Hoggan, Ryan; Culumber, Michele D; Oberg, Craig J; Breakwell, Donald P; Prince, John T; Belnap, David M

    2012-08-01

    Halophage CW02 infects a Salinivibrio costicola-like bacterium, SA50, isolated from the Great Salt Lake. Following isolation, cultivation, and purification, CW02 was characterized by DNA sequencing, mass spectrometry, and electron microscopy. A conserved module of structural genes places CW02 in the T7 supergroup, members of which are found in diverse aquatic environments, including marine and freshwater ecosystems. CW02 has morphological similarities to viruses of the Podoviridae family. The structure of CW02, solved by cryogenic electron microscopy and three-dimensional reconstruction, enabled the fitting of a portion of the bacteriophage HK97 capsid protein into CW02 capsid density, thereby providing additional evidence that capsid proteins of tailed double-stranded DNA phages have a conserved fold. The CW02 capsid consists of bacteriophage lambda gpD-like densities that likely contribute to particle stability. Turret-like densities were found on icosahedral vertices and may represent a unique adaptation similar to what has been seen in other extremophilic viruses that infect archaea, such as Sulfolobus turreted icosahedral virus and halophage SH1.

  12. Role of Temperature and Suwannee River Natural Organic Matter on Inactivation Kinetics of Rotavirus and Bacteriophage MS2 by Solar Irradiation

    KAUST Repository

    Romero, Ofelia C.

    2011-12-15

    Although the sunlight-mediated inactivation of viruses has been recognized as an important process that controls surface water quality, the mechanisms of virus inactivation by sunlight are not yet clearly understood. We investigated the synergistic role of temperature and Suwannee River natural organic matter (SRNOM), an exogenous sensitizer, for sunlight-mediated inactivation of porcine rotavirus and MS2 bacteriophage. Upon irradiation by a full spectrum of simulated sunlight in the absence of SRNOM and in the temperature range of 14-42 °C, high inactivation rate constants, kobs, of MS2 (k obs ≤ 3.8 h-1 or 1-log10 over 0.6 h) and rotavirus (kobs ≤ 11.8 h-1 or ∼1-log10 over 0.2 h) were measured. A weak temperature (14-42 °C) dependence of kobs values was observed for both viruses irradiated by the full sunlight spectrum. Under the same irradiation condition, the presence of SRNOM reduced the inactivation of both viruses due to attenuation of lower wavelengths of the simulated sunlight. For rotavirus and MS2 solutions irradiated by only UVA and visible light in the absence of SRNOM, inactivation kinetics were slow (kobs < 0.3 h-1 or <1-log10 unit reduction over 7 h) and temperature-independent for the range considered. Conversely, under UVA and visible light irradiation and in the presence of SRNOM, temperature-dependent inactivation of MS2 was observed. For rotavirus, the SRNOM-mediated exogenous inactivation was only important at temperatures >33 °C, with low rotavirus kobs values (kobs ≈ 0.2 h-1; 1-log10 unit reduction over 12 h) for the temperature range of 14-33 °C. These kobs values increased to 0.5 h-1 at 43 °C and 1.5 h-1 (1-log10 reduction over 1.6 h) at 50 °C. While SRNOM-mediated exogenous inactivation of MS2 was triggered by singlet oxygen, the presence of hydrogen peroxide was important for rotavirus inactivation in the 40-50 °C range. © 2011 American Chemical Society.

  13. BENEFICIAL FACE OF BACTERIOPHAGES: APPLICATIONS IN FOOD PROCESSING

    Directory of Open Access Journals (Sweden)

    H. V. Raghu

    2012-06-01

    Full Text Available Foods are processed to make them available at all places; consequently, our awareness regarding hygiene measures in food production has also increased dramatically over the last decades. In many countries cases associated with foodborne infectious are increased. However, available techniques are unable to effectively control the problem. Further, exploring novel methods and technologies for ensuring the safety of food with effective quality control approaches are under research. Phages are the natural enemies of bacteria, and are more specific to host renders them ideal candidates for applications designed to increase food safety during the production process. Scientific findings are available showing the possibility to use as biocontrol agents against various pathogens with out interfering with the natural microflora or the cultures in fermented products. Furthermore, phages or phage derived proteins can also be used to detect the presence of unwanted pathogens in food or the production environments, which allows quick and sp ecific identification of viable cells. Bacteriophages are natural, found in various environments including water; foods etc. and are not found significantly influence the human cells.

  14. Biodiversity of Lactobacillus helveticus bacteriophages isolated from cheese whey starters.

    Science.gov (United States)

    Zago, Miriam; Bonvini, Barbara; Rossetti, Lia; Meucci, Aurora; Giraffa, Giorgio; Carminati, Domenico

    2015-05-01

    Twenty-one Lactobacillus helveticus bacteriophages, 18 isolated from different cheese whey starters and three from CNRZ collection, were phenotypically and genetically characterised. A biodiversity between phages was evidenced both by host range and molecular (RAPD-PCR) typing. A more detailed characterisation of six phages showed similar structural protein profiles and a relevant genetic biodiversity, as shown by restriction enzyme analysis of total DNA. Latent period, burst time and burst size data evidenced that phages were active and virulent. Overall, data highlighted the biodiversity of Lb. helveticus phages isolated from cheese whey starters, which were confirmed to be one of the most common phage contamination source in dairy factories. More research is required to further unravel the ecological role of Lb. helveticus phages and to evaluate their impact on the dairy fermentation processes where whey starter cultures are used.

  15. DNA Packaging Specificity of Bacteriophage N15 with an Excursion into the Genetics of a Cohesive End Mismatch.

    Directory of Open Access Journals (Sweden)

    Michael Feiss

    Full Text Available During DNA replication by the λ-like bacteriophages, immature concatemeric DNA is produced by rolling circle replication. The concatemers are processed into mature chromosomes with cohesive ends, and packaged into prohead shells, during virion assembly. Cohesive ends are generated by the viral enzyme terminase, which introduces staggered nicks at cos, an approx. 200 bp-long sequence containing subsites cosQ, cosN and cosB. Interactions of cos subsites of immature concatemeric DNA with terminase orchestrate DNA processing and packaging. To initiate DNA packaging, terminase interacts with cosB and nicks cosN. The cohesive ends of N15 DNA differ from those of λ at 2/12 positions. Genetic experiments show that phages with chromosomes containing mismatched cohesive ends are functional. In at least some infections, the cohesive end mismatch persists through cyclization and replication, so that progeny phages of both allelic types are produced in the infected cell. N15 possesses an asymmetric packaging specificity: N15 DNA is not packaged by phages λ or 21, but surprisingly, N15-specific terminase packages λ DNA. Implications for genetic interactions among λ-like bacteriophages are discussed.

  16. Characterization of bacteriophage KVP40 and T4 RNA ligase 2

    International Nuclear Information System (INIS)

    Yin Shenmin; Kiong Ho, C.; Miller, Eric S.; Shuman, Stewart

    2004-01-01

    Bacteriophage T4 RNA ligase 2 (Rnl2) exemplifies a subfamily of RNA strand-joining enzymes that includes the trypanosome RNA editing ligases. A homolog of T4 Rnl2 is encoded in the 244-kbp DNA genome of vibriophage KVP40. We show that the 335-amino acid KVP40 Rnl2 is a monomeric protein that catalyzes RNA end-joining through ligase-adenylate and RNA-adenylate (AppRNA) intermediates. In the absence of ATP, pre-adenylated KVP40 Rnl2 reacts with an 18-mer 5'-PO 4 single-strand RNA (pRNA) to form an 18-mer RNA circle. In the presence of ATP, Rnl2 generates predominantly AppRNA. Isolated AppRNA can be circularized by KVP40 Rnl2 in the absence of ATP. The reactivity of phage Rnl2 and the distribution of the products are affected by the length of the pRNA substrate. Whereas 18-mer and 15-mer pRNAs undergo intramolecular sealing by T4 Rnl2 to form monomer circles, a 12-mer pRNA is ligated intermolecularly to form dimers, and a 9-mer pRNA is unreactive. In the presence of ATP, the 15-mer and 12-mer pRNAs are converted to AppRNAs, but the 9-mer pRNA is not. A single 5' deoxynucleotide substitution of an 18-mer pRNA substrate has no apparent effect on the 5' adenylation or circularization reactions of T4 Rnl2. In contrast, a single deoxyribonucleoside at the 3' terminus strongly and selectively suppresses the sealing step, thereby resulting in accumulation of high levels of AppRNA in the absence of ATP. The ATP-dependent 'capping' of RNA with AMP by Rnl2 is reminiscent of the capping of eukaryotic mRNA with GMP by GTP:RNA guanylyltransferase and suggests an evolutionary connection between bacteriophage Rnl2 and eukaryotic RNA capping enzymes

  17. Dominant negative umuD mutations decreasing RecA-mediated cleavage suggest roles for intact UmuD in modulation of SOS mutagenesis

    International Nuclear Information System (INIS)

    Battista, J.R.; Ohta, Toshihiro; Nohmi, Takehiko; Sun, W.; Walker, G.C.

    1990-01-01

    The products of the SOS-regulated umuDC operon are required for most UV and chemical mutagenesis in Escherichia coli. The UmuD protein shares homology with a family of proteins that includes LexA and several bacteriophage repressors. UmuD is posttranslationally activated for its role n mutagenesis by a RecA-mediated proteolytic cleavage that yields UmuD'. A set of missense mutants of umuD was isolated and shown to encode mutant UmuD proteins that are deficient in RecA-mediated cleavage in vivo. Most of these mutations are dominant to umuD + with respect to UV mutagenesis yet do not interfere with SOS induction. Although both UmuD and UmuD' form homodimers, the authors provide evidence that they preferentially form heterodimers. The relationship of UmuD to LexA, λ repressor, and other members of the family of proteins is discussed and possible roles intact UmuD in modulating SOS mutagenesis are discussed

  18. Involvement of DNA gyrase in replication and transcription of bacteriophage T7 DNA

    International Nuclear Information System (INIS)

    De Wyngaert, M.A.; Hinkle, D.C.

    1979-01-01

    Growth of bacteriophage T7 is inhibited by the antibiotic coumermycin A 1 , an inhibitor of the Escherichia coli DNA gyrase. Since growth of the phage is insensitive to the antibiotic in strains containing a coumermycin-resistent DNA gyrase, this enzyme appears to be required for phage growth. We have investigated the effect of coumermycin on the kinetics of DNA, RNA, and protein synthesis during T7 infection. DNA synthesis is completely inhibited by the antibiotic. In addition, coumermycin significantly inhibits transcription of late but not early genes. Thus, E. coli DNA gyrase may play an important role in transcription as well as in replication of T7 DNA

  19. The oxygen effect in bacteriophages irradiated in different media. 1

    International Nuclear Information System (INIS)

    Korystov, Yu.N.; Veksler, F.B.

    1983-01-01

    The oxygen effect (OE) on bacteriophage T4 in a salt solution was studied. It is shown that the sign and magnitude of OE depend on the conditions of the postirradiation incubation of the phage in irradiated medium. The direct OE is due to postirradiation lesion of the phage by hydrogen peroxide which is formed in greater amounts after irradiation in oxygen than in anoxia. The addition of catalase is shown to eliminate the postirradiation inactivation of the phage. In this case an opposite OE is observed. The mechanism of this effect is a scavenge of hydrogen atoms which damage the phage by oxygen. In the presence of catalase the OE depends also on pH of the solution. It is suggested that the hydroxyl radical arising from the reaction of H 2 O 2 with Fe 2+ is responsible for the damaging effect of H 2 O 2 . (author)

  20. Lactococcus bacteriophages isolated from whey and their effects on commercial lactic starters

    Directory of Open Access Journals (Sweden)

    Maria Raquel de Godoy Oriani

    2004-08-01

    Full Text Available The incidence of phages of lactic acid bacteria in milk industry and their effects on acidification ability of commercial lactic acid starters were studied. Cheese whey samples (33 samples were collected from 17 factories. A total of 16 bacteriophages were isolated (12 specific for Lactococcus lactis, 3 for L. diacetylactis and one capable of lysing both species. The results showed that 10% reduction in acidification tests was not good indication of phage in the sample. The majority of samples showed reduction higher than 10%, although only 65% were phage positive. The isolated phages were quite stable and showed no reduction in infectivity even after 20 daily replications. A pool of bacteriophages was prepared from isolates and inoculated in 12 commercial lactic starters. After 8 hours of incubation, only 2 showed reduced acidification. Bacterial strains isolated from commercial starters were tested regarding the phage resistance. Considerable difference in phage sensitivity was observed among different starters (BD, D, O and L. diacetylactis. Five bacteriophages showed no infectivity on any isolates but one was infective for most of isolates.Para ampliar conhecimentos sobre a incidência de bacteriófagos de bactérias lácticas na indústria de leite do Estado de São Paulo e a sua influência sobre a capacidade acidificante de fermentos lácticos disponíveis em nosso mercado, o presente trabalho foi conduzido com o intuito de esclarecer a real situação dos laticínios no Estado. Foram coletadas 33 amostras de soro de queijo em 17 laticínios. Foram isolados 16 bacteriófagos, 12 específicos para Lactococcus lactis, 3 para L. diacetylactis e um capaz de lisar ambos os microrganismos. Os experimentos mostraram que, uma diminuição de 10% na acidez em presença de soro suspeito, ao contrário do estabelecido na literatura, não reflete a veracidade da presença de bacteriófagos na amostra, uma vez que a maioria apresentou redução acima