WorldWideScience

Sample records for human feces samples

  1. Specificity of a Bacteroides thetaiotaomicron marker for human feces

    Science.gov (United States)

    Carson, C.A.; Christiansen, J.M.; Yampara-Iquise, H.; Benson, V.W.; Baffaut, C.; Davis, J.V.; Broz, R.R.; Kurtz, W.B.; Rogers, W.M.; Fales, W.H.

    2005-01-01

    A bacterial primer set, known to produce a 542-bp amplicon specific for Bacteroides thetaiotaomicron, generated this product in PCR with 1 ng of extracted DNA from 92% of 25 human fecal samples, 100% of 20 sewage samples, and 16% of 31 dog fecal samples. The marker was not detected in 1 ng of fecal DNA from 61 cows, 35 horses, 44 pigs, 24 chickens, 29 turkeys, and 17 geese. Copyright ?? 2005, American Society for Microbiology. All Rights Reserved.

  2. Impact of consumption of probiotic lactobacilli-containing yogurt on microbial composition in human feces.

    Science.gov (United States)

    Uyeno, Yutaka; Sekiguchi, Yuji; Kamagata, Yoichi

    2008-02-29

    An in vivo study was carried out to determine the effect of consuming probiotic lactobacilli-containing yogurt on the composition of microbiota in the human gut. Fifteen healthy adults ingested a daily serving of one of three commercial yogurts (two of the products contained a probiotic lactobacilli strain) for 20 days. Fecal samples at defined time points before, during, and after the period of yogurt ingestion were collected and analyzed. The fecal population of lactobacilli was determined by a culture-based method and subsequent colony PCR for the identification of species. Six predominant bacterial groups in the fecal samples were quantitatively determined based on a sequence-specific SSU rRNA cleavage method coupled with a suite of oligonucleotide probes, which was optimized for the target-specific detection of bacterial groups inhabiting human feces. In the ingestion period, one probiotic strain was detected in the feces of all five subjects who consumed the yogurt containing the strain, while the other strain was detected in three of another five subjects. The population levels of the two major groups (Bacteroides and Prevotella, and the Clostridium coccoides-Eubacterium rectale group) in the fecal samples tended to change in response to the ingestion but the change did not seem to be dependent on the product-specific property of each yogurt. These results suggest that the human fecal bacterial community could be altered by ingesting yogurt, although whether probiotic lactobacilli are present or absent in the yogurt does not seem to be a factor in this change.

  3. Use of pooled samples for the detection of Salmonella in feces by polymerase chain reaction.

    Science.gov (United States)

    Singer, Randall S; Cooke, Cara L; Maddox, Carol W; Isaacson, Richard E; Wallace, Richard L

    2006-07-01

    Many epidemiological studies of Salmonella rely on conventional bacteriological culture methods to detect Salmonella in fecal samples. These culture-based methods are inefficient for epidemiological studies in populations with a low prevalence of Salmonella. The objective of this study was to optimize a protocol that uses pooled Salmonella enrichment broth cultures of bovine feces and polymerase chain reaction (PCR) for the detection of the invA gene of Salmonella in feces. In one field trial, 196 animals were sampled, and all samples were tested by culture, invA PCR on individual samples, invA PCR on pools of 5 samples, and BAX PCR on individual samples. All assays showed a high agreement on individual samples (kappa > or = 0.75). The invA PCR was run on each of 40 pools and detected 19 of 22 culture-positive pools. In another field trial, 152 samples were taken from 4 dairies, and the invA PCR was performed on pools of 5 samples in addition to bacteriological culture of individual samples. Salmonella was detected in 5 of the 32 pools (7 total positive samples) by both PCR and culture. One pool was PCR-positive but culture-negative. Pooling did not dramatically affect the performance of the invA PCR; most of the culture-positive samples were detected, including all of the samples when there were 4 or more Salmonella colonies on the agar plate. Based on these field trials, invA PCR on pooled samples appears to be an efficient method of Salmonella detection as long as Salmonella loads are not extremely low.

  4. Electricity generation directly using human feces wastewater for life support system

    Science.gov (United States)

    Fangzhou, Du; Zhenglong, Li; Shaoqiang, Yang; Beizhen, Xie; Hong, Liu

    2011-05-01

    Wastewater reuse and power regeneration are key issues in the research of bioregeneration life support system (BLSS). Microbial fuel cell (MFC) can generate electricity during the process of wastewater treatment, which might be promising to solve the two problems simultaneously. We used human feces wastewater containing abundant organic compounds as the substrate of MFC to generate electricity, and the factors concerning electricity generation capacity were investigated. The removal efficiency of total chemical oxygen demand (TCOD), Soluble chemical oxygen demand (SCOD) and NH4+ reached 71%, 88% and 44%, respectively with two-chamber MFC when it was fed with the actual human feces wastewater and operated for 190 h. And the maximum power density reached 70.8 mW/m 2, which implicated that MFC technology was feasible and appropriate for treating human feces wastewater. In order to improve the power generation of MFC further, human feces wastewater were fermented before poured into MFC, and the result showed that fermentation pretreatment could improve the MFC output obviously. The maximum power density of MFC fed with pretreated human feces wastewater was 22 mW/m 2, which was 47% higher than that of the control without pretreatment (15 mW/m 2). Furthermore, the structure of MFC was studied and it was found that both enlarging the area of electrodes and shortening the distance between electrodes could increase the electricity generation capacity. Finally, an automatic system, controlled by time switches and electromagnetic valves, was established to process one person's feces wastewater (1 L/d) while generating electricity. The main parts of this system comprised a pretreatment device and 3 one-chamber air-cathode MFCs. The total power could reach 787.1 mW and power density could reach the maximum of about 240 mW/m 2.

  5. Characterization of Enterococcus spp. from human and animal feces using 16S rRNA sequences, the esp gene, and PFGE for microbial source tracking in Korea.

    Science.gov (United States)

    Kim, Sei-Yoon; Lee, Jung Eun; Lee, Sunghee; Lee, Hee Tae; Hur, Ho-Gil; Ko, Gwangpyo

    2010-05-01

    Contamination from human and animal fecal waste is a primary cause of water pollution. Microbial source tracking (MST) may be a useful tool for high-quality environmental management and for assessing human health risks associated with water pollution. The goal of this study was to evaluate Enterococcus spp. as a target organism for MST. Thirty-four fecal samples were collected from five different sources (human, chicken, pig, cow, and goose) in South Korea. In total, 237 Enterococcus spp. were isolated from feces using membrane- Enterococcus indoxyl-beta-d-glucoside agar. The 16S rRNA gene and the whole genome were analyzed using nucleic acid sequencing and pulsed-field gel electrophoresis (PFGE), respectively. Both phylogenetic analysis and principal coordinate analysis using UniFrac were performed on the nucleic acid sequences of the 16S rRNA gene. According to P-tests from UniFrac, significant differences existed between Enterococcus spp. isolated from human feces and those from animal feces. In addition, we evaluated whether the esp gene of Enterococcus faecium could be a specific target for Enterococcus spp. isolated from human feces. Of 58 E. faecium isolates tested, only three were esp-positive. The specificity of the esp gene of E. faecium isolated from human feces was 100%, but the sensitivity was esp gene and 16S rRNA sequences, whereas PFGE provides limited information on the fecal sources of Enterococcus spp.

  6. Evaluation of microbial contamination of feces and soil on a laying-hen farm depending on sampling site and season

    Directory of Open Access Journals (Sweden)

    Beata Trawińska

    2016-04-01

    Full Text Available ABSTRACT The objective of the present study was to evaluate soil collected from a laying-hen farm and bird manure according to the season of the year and sampling site. Soil samples were taken at the poultry facility wall and at the distances of 15 m and 45 m from the building. Bird feces samples were collected inside the poultry house at the entrance and at 1/4 and 1/2 length of the building. Soil and bird feces samples were evaluated by bacteriological qualitative and quantitative analyses. The largest bacterial load was determined in the samples taken at the poultry facility wall in December/January. Soil microbial contamination degree was low. The highest bacterial count in bird manure was found in the samples collected at 1/2 length of the hen house at the end of December/January. The qualitative study of bird feces showed the presence of E. coli bacteria all through the research period and Enterobacter spp. in the samples taken from July until September. Microbial contamination of soil environment and bird feces is most likely to be affected by winter period as at that time the highest microbial population can be determined. This fact may be linked to the prevailing climatic and microclimatic conditions.

  7. Pheromone-responsive conjugative vancomycin resistance plasmids in Enterococcus faecalis isolates from humans and chicken feces.

    Science.gov (United States)

    Lim, Suk-Kyung; Tanimoto, Koichi; Tomita, Haruyoshi; Ike, Yasuyoshi

    2006-10-01

    The drug resistances and plasmid contents of a total of 85 vancomycin-resistant enterococcus (VRE) strains that had been isolated in Korea were examined. Fifty-four of the strains originated from samples of chicken feces, and 31 were isolated from hospital patients in Korea. Enterococcus faecalis KV1 and KV2, which had been isolated from a patient and a sample of chicken feces, respectively, were found to carry the plasmids pSL1 and pSL2, respectively. The plasmids transferred resistances to vancomycin, gentamicin, kanamycin, streptomycin, and erythromycin to E. faecalis strains at a high frequency of about 10(-3) per donor cell during 4 hours of broth mating. E. faecalis strains containing each of the pSL plasmids formed clumps after 2 hours of incubation in broth containing E. faecalis FA2-2 culture filtrate (i.e., the E. faecalis sex pheromone), and the plasmid subsequently transferred to the recipient strain in a 10-min short mating in broth, indicating that the plasmids are responsive to E. faecalis pheromones. The pSL plasmids did not respond to any of synthetic pheromones for the previously characterized plasmids. The pheromone specific for pSL plasmids has been designated cSL1. Southern hybridization analysis showed that specific FspI fragments from each of the pSL plasmids hybridized with the aggregation substance gene (asa1) of the pheromone-responsive plasmid pAD1, indicating that the plasmids had a gene homologous to asa1. The restriction maps of the plasmids were identical, and the size of the plasmids was estimated to be 128.1 kb. The plasmids carried five drug resistance determinants for vanA, ermB, aph(3'), aph(6'), and aac(6')/aph(2'), which encode resistance to vancomycin, erythromycin, kanamycin, streptomycin, and gentamicin/kanamycin, respectively. Nucleotide sequence analyses of the drug resistance determinants and their flanking regions are described in this report. The results described provide evidence for the exchange of genetic information

  8. Degradation of typical antibiotics during human feces aerobic composting under different temperatures.

    Science.gov (United States)

    Shi, Honglei; Wang, Xiaochang C; Li, Qian; Jiang, Shanqing

    2016-08-01

    Four typical antibiotics were added to human feces for aerobic composting using batch reactors with sawdust as the bulk matrix. Under three composting temperatures (room temperature, 35 ± 2 °C and 55 ± 2 °C), decreases in the extractable concentrations of antibiotics in the compost were monitored for 20 days. As a result, the removals of extractable tetracycline and chlortetracycline were found to be more temperature-dependent than the removals of sulfadiazine and ciprofloxacin. However, more than 90 % of all of the extractable antibiotics were removed at 55 ± 2 °C. Three specific experiments were further conducted to identify the possible actions for antibiotic removal, including self-degradation in aqueous solution, composting with a moist sterile sawdust matrix without adding feces and composting with human feces and moist sterile sawdust. As a result, it was found that the removal of tetracycline and chlortetracycline was mainly due to chemical degradation in water, whereas the removal of sulfadiazine was mainly attributed to adsorption onto sawdust particles. The microbial activity of compost varied with temperature to a certain extent, but the differences were insignificant among different antibiotics. Although microbial action is important for organic matter decomposition, its contribution to antibiotic degradation was small for the investigated antibiotics, except for ciprofloxacin, which was degraded by up to 20 % due to microbial action.

  9. Effect of Raw-Milk Cheese Consumption on the Enterococcal Flora of Human Feces

    OpenAIRE

    Gelsomino, Roberto; Vancanneyt, Marc; Cogan, Timothy M.; Swings, Jean

    2003-01-01

    Enterococci are one of the major facultative anaerobic bacterial groups that reside in the human gastrointestinal tract. In the present study, the composition of the enterococcal fecal flora in three healthy humans was analyzed before, during, and after the daily consumption of ∼125 g of a raw-milk Cheddar-type cheese containing 3.2 × 104 enterococci/g of cheese. Enterococcal counts ranged between 1.4 × 102 and 2.5 × 108 CFU/g of feces and differed from subject to subject and from week to wee...

  10. Fecal collection, ambient preservation, and DNA extraction for PCR amplification of bacterial and human markers from human feces.

    Science.gov (United States)

    Nechvatal, Jordan M; Ram, Jeffrey L; Basson, Marc D; Namprachan, Phanramphoei; Niec, Stephanie R; Badsha, Kawsar Z; Matherly, Larry H; Majumdar, Adhip P N; Kato, Ikuko

    2008-02-01

    Feces contain intestinal bacteria and exfoliated epithelial cells that may provide useful information concerning gastrointestinal tract health. Intestinal bacteria that synthesize or metabolize potential carcinogens and produce anti-tumorigenic products may have relevance to colorectal cancer, the second most common cause of cancer deaths in the USA. To facilitate epidemiological studies relating bacterial and epithelial cell DNA and RNA markers, preservative/extraction methods suitable for self-collection and shipping of fecal samples at room temperature were tested. Purification and PCR amplification of fecal DNA were compared after preservation of stool samples in RNAlater (R) or Paxgene (P), or after drying over silica gel (S) or on Whatman FTA cards (W). Comparisons were made to samples frozen in liquid nitrogen (N2). DNA purification methods included Whatman (accompanying FTA cards), Mo-Bio Fecal (MB), Qiagen Stool (QS), and others. Extraction methods were compared for amount of DNA extracted, DNA amplifiable in a real-time SYBR-Green quantitative PCR format, and the presence of PCR inhibitors. DNA can be extracted after room temperature storage for five days from W, R, S and P, and from N2 frozen samples. High amounts of total DNA and PCR-amplifiable Bacteroides spp. DNA (34%+/-9% of total DNA) with relatively little PCR inhibition were especially obtained with QS extraction applied to R preserved samples (method QS-R). DNA for human reduced folate carrier (SLC19A1) genomic sequence was also detected in 90% of the QS-R extracts. Thus, fecal DNA is well preserved by methods suitable for self-collection that may be useful in future molecular epidemiological studies of intestinal bacteria and human cancer markers.

  11. Dung beetles (Coleoptera: Scarabaeidae) attracted to dung of the largest herbivorous rodent on earth: a comparison with human feces.

    Science.gov (United States)

    Puker, Anderson; Correa, César M A; Korasaki, Vanesca; Ferreira, Kleyton R; Oliveira, Naiara G

    2013-12-01

    The capybara, Hydrochoerus hydrochaeris (L.) (Rodentia: Caviidae), is the largest herbivorous rodent on Earth and abundant in the Neotropical region, which can provide a stable food source of dung for dung beetle communities (Coleoptera: Scarabaeidae: Scarabaeinae). However, the use of capybara dung by dung beetles is poorly known. Here, we present data on the structure of the dung beetle community attracted to capybara dung and compare with the community attracted to human feces. Dung beetles were captured with pitfall traps baited with fresh capybara dung and human feces in pastures with exotic grass (Brachiaria spp.), patches of Brazilian savanna (Cerrado), and points of degraded riparian vegetation along the Aquidauana river in Anastácio and Aquidauana, Mato Grosso do Sul, Brazil. In traps baited with human feces, 13,809 individuals of 31 species were captured, and in those baited with capybara dung 1,027 individuals belonging to 26 species were captured. The average number of individuals and species captured by the traps baited with human feces was greater than for capybara dung in all habitats studied. Composition of the communities attracted to human feces and capybara dung formed distinct groups in all habitats. Despite the smaller number of species and individuals captured in capybara dung when compared with human feces, capybara dung was attractive to dung beetles. In Brazil, the legalization of hunting these rodents has been debated, which would potentially affect the community and consequently the ecological functions performed by dung beetles that use the feces of these animals as a resource. In addition, the knowledge of the communities associated with capybaras may be important in predicting the consequences of future management of their populations.

  12. Antibiotic resistance in Escherichia coli strains isolated from Antarctic bird feces, water from inside a wastewater treatment plant, and seawater samples collected in the Antarctic Treaty area

    Science.gov (United States)

    Rabbia, Virginia; Bello-Toledo, Helia; Jiménez, Sebastián; Quezada, Mario; Domínguez, Mariana; Vergara, Luis; Gómez-Fuentes, Claudio; Calisto-Ulloa, Nancy; González-Acuña, Daniel; López, Juana; González-Rocha, Gerardo

    2016-06-01

    Antibiotic resistance is a problem of global concern and is frequently associated with human activity. Studying antibiotic resistance in bacteria isolated from pristine environments, such as Antarctica, extends our understanding of these fragile ecosystems. Escherichia coli strains, important fecal indicator bacteria, were isolated on the Fildes Peninsula (which has the strongest human influence in Antarctica), from seawater, bird droppings, and water samples from inside a local wastewater treatment plant. The strains were subjected to molecular typing with pulsed-field gel electrophoresis to determine their genetic relationships, and tested for antibiotic susceptibility with disk diffusion tests for several antibiotic families: β-lactams, quinolones, aminoglycosides, tetracyclines, phenicols, and trimethoprim-sulfonamide. The highest E. coli count in seawater samples was 2400 cfu/100 mL. Only strains isolated from seawater and the wastewater treatment plant showed any genetic relatedness between groups. Strains of both these groups were resistant to β-lactams, aminoglycosides, tetracycline, and trimethoprim-sulfonamide.In contrast, strains from bird feces were susceptible to all the antibiotics tested. We conclude that naturally occurring antibiotic resistance in E. coli strains isolated from Antarctic bird feces is rare and the bacterial antibiotic resistance found in seawater is probably associated with discharged treated wastewater originating from Fildes Peninsula treatment plants.

  13. Characterizing Properties of Biochar Produced from Simulated Human Feces and Its Potential Applications.

    Science.gov (United States)

    Ilango, Ajaykannan; Lefebvre, Olivier

    2016-03-01

    This study presents a comprehensive characterization of biochar obtained from simulated human feces (SHF) with a view to improve human waste sanitization and stabilization before usage as a resource. The possible applications of SHF are as a fuel, as a soil amendment, or for emerging applications (e.g., activated carbon precursor and odor control), depending on the charring conditions. Simulated human feces were charred under different conditions of peak temperature (200-800°C), heating rate (2-50°C min), and holding time (0.5-6.0 h); these parameters have been shown to have the largest influence on the thermal and physicochemical characteristics of the final product. The peak temperature was shown to have a higher impact than the heating rate or the holding time. At 200°C, the very mild structural changes of the product were characteristic of dry torrefaction, a process useful to remove moisture and sterilize the product. At 400°C the carbon content (76.2 ± 0.4) and the calorific heat value (30.6 ± 0.4 MJ kg) of the product increased by 60%. From 600°C onward, the improved degree of aromatization verified by Fourier transform infrared spectrometry (alkene [C=C] stretching around 1680-1450 cm) and C nuclear magnetic resonance (C=C stretching at 140-110 ppm) made the biochar increasingly suitable for carbon sequestration or commercial fabrication of briquettes of charcoal. In conclusion, SHF proved to be a suitable feedstock to produce a biochar whose characteristics depended mostly on the peak charring temperature. Ultimately, the selection of a suitable application may depend on local and sociological considerations.

  14. Identification and quantification of Lactobacillus casei strain Shirota in human feces with strain-specific primers derived from randomly amplified polymorphic DNA.

    Science.gov (United States)

    Fujimoto, Junji; Matsuki, Takahiro; Sasamoto, Masae; Tomii, Yasuaki; Watanabe, Koichi

    2008-08-15

    Lactobacillus casei strain Shirota (LcS) has been used in the production of fermented milk products for many years and is one of the most intensively studied probiotics. To evaluate the ability of LcS to proliferate in human intestines after it has been ingested, we developed a PCR-based method to identify and quantify LcS using an LcS-specific primer set (pLcS) derived from a randomly amplified polymorphic DNA (RAPD) analysis. We confirmed the high specificity of the pLcS primer set in 167 bacterial strains (57 strains of L. casei and 110 other strains of bacteria commonly isolated from human feces). The method's ability to identify LcS matched that of an ELISA using a monoclonal antibody and a RAPD analysis in a representative sample of colonies cultured from human feces. The detection limit of quantitative PCR (qPCR) using pLcS was 10(4.6) per gram of feces. The number of LcS in feces detected with qPCR was highly and significantly correlated with the number of LcS added to fecal samples within the range of 10(4.6) to 10(9.6) per gram feces (r(2)=0.999, P<0.001). After 14 healthy subjects ingested 10(11.0) CFU of LcS daily for 7 days, 10(9.1+/-0.5) LcS g(-1) (mean+/-S.D.) was detected in the fecal samples of all subjects by qPCR, and 10(8.0+/-0.9) CFU g(-1) was detected by culture; these values were significantly different (P<0.001, paired t-test). After the subjects stopped ingesting LcS, fecal LcS counts obtained with both methods decreased daily. The values produced by the 2 methods might have differed because of an overestimation in the PCR analysis due to the presence of dead LcS cells or an underestimation in the culture system due to the use of selective culture media; however, dead LcS cells can also be beneficial as immunomodulators. We confirmed that qPCR with an LcS-specific primer set was a rapid and accurate method for determining the total amount of LcS in feces including dead or less active cells which could not be detected by culture method.

  15. Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches.

    Directory of Open Access Journals (Sweden)

    Gunilla Veslemøy Schmidt

    Full Text Available The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same samples, and therefore differences in antibiotic resistance levels between samples were more readily detected. To our knowledge, this is the first study to describe sampling and pooling methods for qPCR quantification of antibiotic resistance genes in total DNA extracted from swine feces.

  16. Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

    Science.gov (United States)

    Mellerup, Anders; Ståhl, Marie

    2015-01-01

    The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same samples, and therefore differences in antibiotic resistance levels between samples were more readily detected. To our knowledge, this is the first study to describe sampling and pooling methods for qPCR quantification of antibiotic resistance genes in total DNA extracted from swine feces. PMID:26114765

  17. Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches.

    Science.gov (United States)

    Schmidt, Gunilla Veslemøy; Mellerup, Anders; Christiansen, Lasse Engbo; Ståhl, Marie; Olsen, John Elmerdahl; Angen, Øystein

    2015-01-01

    The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same samples, and therefore differences in antibiotic resistance levels between samples were more readily detected. To our knowledge, this is the first study to describe sampling and pooling methods for qPCR quantification of antibiotic resistance genes in total DNA extracted from swine feces.

  18. Species From Feces: Order-Wide Identification of Chiroptera From Guano and Other Non-Invasive Genetic Samples

    Science.gov (United States)

    Williamson, Charles H. D.; Sanchez, Daniel E.; Sobek, Colin J.; Chambers, Carol L.

    2016-01-01

    Bat guano is a relatively untapped reservoir of information, having great utility as a DNA source because it is often available at roosts even when bats are not and is an easy type of sample to collect from a difficult-to-study mammalian order. Recent advances from microbial community studies in primer design, sequencing, and analysis enable fast, accurate, and cost-effective species identification. Here, we borrow from this discipline to develop an order-wide DNA mini-barcode assay (Species from Feces) based on a segment of the mitochondrial gene cytochrome c oxidase I (COI). The assay works effectively with fecal DNA and is conveniently transferable to low-cost, high-throughput Illumina MiSeq technology that also allows simultaneous pairing with other markers. Our PCR primers target a region of COI that is highly discriminatory among Chiroptera (92% species-level identification of barcoded species), and are sufficiently degenerate to allow hybridization across diverse bat taxa. We successfully validated our system with 54 bat species across both suborders. Despite abundant arthropod prey DNA in guano, our primers were highly specific to bats; no arthropod DNA was detected in thousands of feces run on Sanger and Illumina platforms. The assay is extendable to fecal pellets of unknown age as well as individual and pooled guano, to allow for individual (using singular fecal pellets) and community (using combined pellets collected from across long-term roost sites) analyses. We developed a searchable database (http://nau.edu/CEFNS/Forestry/Research/Bats/Search-Tool/) that allows users to determine the discriminatory capability of our markers for bat species of interest. Our assay has applications worldwide for examining disease impacts on vulnerable species, determining species assemblages within roosts, and assessing the presence of bat species that are vulnerable or facing extinction. The development and analytical pathways are rapid, reliable, and inexpensive, and

  19. Sample handling factors affecting the enumeration of lactobacilli and cellulolytic bacteria in equine feces

    Science.gov (United States)

    The objectives were to compare media types and evaluate the effects of fecal storage time and temperature on the enumeration of cellulolytic bacteria and lactobacilli from horses. Fecal samples were collected from horses (n = 3) and transported to the lab (CO2, 37 ºC, 0.5 h). The samples were assign...

  20. DETECTION OF INTRINSIC VANCOMYCIN RESISTANT ENTEROCOCCI IN ANIMAL AND HUMAN FECES

    Science.gov (United States)

    A survey was conducted to determine the occurrence of vancomycin resistant enterococci (VRE) in animal and human fecal samples. Fecal samples from 14 animal species and humans were analyzed by quantitative culture for enterococci and VRE. Over 800 VRE isolates were characterize...

  1. Mechanism of Human Influenza Virus RNA Persistence and Virion Survival in Feces: Mucus Protects Virions From Acid and Digestive Juices.

    Science.gov (United States)

    Hirose, Ryohei; Nakaya, Takaaki; Naito, Yuji; Daidoji, Tomo; Watanabe, Yohei; Yasuda, Hiroaki; Konishi, Hideyuki; Itoh, Yoshito

    2017-07-01

    Although viral RNA or infectious virions have been detected in the feces of individuals infected with human influenza A and B viruses (IAV/IBV), the mechanism of viral survival in the gastrointestinal tract remains unclear. We developed a model that attempts to recapitulate the conditions encountered by a swallowed virus. While IAV/IBV are vulnerable to simulated digestive juices (gastric acid and bile/pancreatic juice), highly viscous mucus protects viral RNA and virions, allowing the virus to retain its infectivity. Our results suggest that virions and RNA present in swallowed mucus are not inactivated or degraded by the gastrointestinal environment, allowing their detection in feces. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

  2. Comparison between immunomagnetic separation, coupled with immunofluorescence, and the techniques of Faust et al. and of Lutz for the diagnosis of Giardia lamblia cysts in human feces

    Directory of Open Access Journals (Sweden)

    Souza Doris Sobral Marques

    2003-01-01

    Full Text Available In the present study, the performance of Immunomagnetic Separation technique, coupled with Immunofluorescence (IMS-IFA, was compared with the FAUST et al. and Lutz parasitological techniques for the detection of Giardia lamblia cysts in human feces. One hundred and twenty-seven samples were evaluated by the three techniques at the same time showing a rate of cyst detection of 27.5% by IMS-IFA and 15.7% by both Faust et al. and Lutz techniques. Data analysis showed a higher sensitivity of IMS-IFA for the detection of G. lamblia cysts in comparison with the techniques of FAUST et al. and Lutz. The use of this methodology as a routine procedure enables the processing of many samples simultaneously, in order to increase recovery rate of G. lamblia cysts and reduce the time of sample storage.

  3. Occurrence of microbial indicators and Clostridium perfringens in wastewater, water column samples, sediments, drinking water, and Weddell seal feces collected at McMurdo Station, Antarctica.

    Science.gov (United States)

    Lisle, John T; Smith, James J; Edwards, Diane D; McFeters, Gordon A

    2004-12-01

    McMurdo Station, Antarctica, has discharged untreated sewage into McMurdo Sound for decades. Previous studies delineated the impacted area, which included the drinking water intake, by using total coliform and Clostridium perfringens concentrations. The estimation of risk to humans in contact with the impacted and potable waters may be greater than presumed, as these microbial indicators may not be the most appropriate for this environment. To address these concerns, concentrations of these and additional indicators (fecal coliforms, Escherichia coli, enterococci, coliphage, and enteroviruses) in the untreated wastewater, water column, and sediments of the impacted area and drinking water treatment facility and distribution system at McMurdo Station were determined. Fecal samples from Weddell seals in this area were also collected and analyzed for indicators. All drinking water samples were negative for indicators except for a single total coliform-positive sample. Total coliforms were present in water column samples at higher concentrations than other indicators. Fecal coliform and enterococcus concentrations were similar to each other and greater than those of other indicators in sediment samples closer to the discharge site. C. perfringens concentrations were higher in sediments at greater distances from the discharge site. Seal fecal samples contained concentrations of fecal coliforms, E. coli, enterococci, and C. perfringens similar to those found in untreated sewage. All samples were negative for enteroviruses. A wastewater treatment facility at McMurdo Station has started operation, and these data provide a baseline data set for monitoring the recovery of the impacted area. The contribution of seal feces to indicator concentrations in this area should be considered.

  4. Strategy for nuclear-magnetic-resonance-based metabolomics of human feces

    DEFF Research Database (Denmark)

    Lamichhane, Santosh; Yde, Christian Clement; Schmedes, Mette Søndergaard

    2015-01-01

    Metabolomic analyses of fecal material are gaining increasing attention because the gut microbial ecology and activity have an impact on the human phenotype and regulate host metabolism. Sample preparation is a crucial step, and in this study we recommend a methodology for extraction and analysis...

  5. [Fertility and Environmental Impacts of Urban Scattered Human Feces Used as Organic Granular Fertilizer for Leaf Vegetables].

    Science.gov (United States)

    Lü, Wen-zhou; Qiao, Yu-xiang; Yu, Ning; Shi, Rong-hua; Wang, Guang-ming

    2015-09-01

    The disposal of urban scattered human feces has become a difficult problem for the management of modern city. In present study, the scattered human feces underwent the collection, scum removal, flocculation and dehydration, finally became the granular fertilizer; the effects of the ratio of fertilizer to soil on the growth of the pakchoi and the quality of soil and leaching water were evaluated, and the feasibility of granular fertilizer manuring the pakchoi was discussed by pot experiments. The results showed that the granular fertilizer significantly enhanced the production of the pakchoi which were not polluted by the intestinal microorganisms under the experiment conditions; meanwhile, at the proper ratio of fertilizer to soil, the concentration of these microorganisms in the leaching water was lower than that in the control check. Chemical analyses of soil revealed that the nutrient content of nitrogen, phosphorus, potassium and organic matters in soil became much richer in all treatments. In addition, the granular fertilizer improved the physical- chemical properties of soil, including raising the level of soil porosity and reducing the volume weight of soil. Application of granular fertilizer won't pollute the soil or leaching water; instead, it can also prevent nitrogen, potassium and intestinal microorganisms from leaching inio ground water at the proper ratio of granular fertilizer to soil.

  6. Comparative study of microbial-derived phenolic metabolites in human feces after intake of gin, red wine, and dealcoholized red wine.

    Science.gov (United States)

    Jiménez-Girón, Ana; Queipo-Ortuño, María Isabel; Boto-Ordóñez, Maria; Muñoz-González, Irene; Sánchez-Patán, Fernando; Monagas, Maria; Martín-Álvarez, Pedro J; Murri, Mora; Tinahones, Francisco J; Andrés-Lacueva, Cristina; Bartolomé, Begoña; Moreno-Arribas, M Victoria

    2013-04-24

    The analysis of microbial phenolic metabolites in fecal samples from in vivo studies is crucial to understanding the potential modulatory effects derived from polyphenol consumption and its overall health effects, particularly at the gut level. In this study, the composition of microbial phenolic metabolites in human feces collected after regular consumption of either red wine, dealcoholized red wine, or gin was analyzed by UPLC-ESI-MS/MS. Red wine interventions produce a change in the content of eight phenolic acids, which are probably derived from the catabolism of flavan-3-ols and anthocyanins, the main flavonoids in red wine. Moreover, alcohol seemed not to influence the formation of phenolic metabolites by the gut microbiota. A principal component analysis revealed large interindividual differences in the formation of microbial metabolites after each red wine polyphenol intervention, but not after the gin intervention, indicating differences in the gut microbial composition among subjects.

  7. Aciduric Strains of Lactobacillus reuteri and Lactobacillus rhamnosus, Isolated from Human Feces, Have Strong Adhesion and Aggregation Properties.

    Science.gov (United States)

    Klopper, Kyle B; Deane, Shelly M; Dicks, Leon M T

    2017-07-29

    Human feces were streaked onto MRS Agar adjusted to pH 2.5, 3.0, and 6.4, respectively, and medium supplemented with 1.0% (w/v) bile salts. Two aciduric strains, identified as Lactobacillus reuteri HFI-LD5 and Lactobacillus rhamnosus HFI-K2 (based on 16S rDNA and recA sequences), were non-hemolytic and did not hydrolyze mucin. The surface of Lactobacillus reuteri HFI-LD5 cells has a weak negative charge, whereas Lactobacillus rhamnosus HFI-K2 has acidic and basic properties, and produces exopolysaccharides (EPS). None of the strains produce bacteriocins. Both strains are resistant to several antibiotics, including sulfamethoxazole-trimethoprim and sulphonamides. The ability of Lactobacillus reuteri HFI-LD5 and Lactobacillus rhamnosus HFI-K2 to grow at pH 2.5 suggests that they will survive passage through the stomach. EPS production may assist in binding to intestinal mucus, especially in the small intestinal tract, protect epithelial cells, and stimulate the immune system. Lactobacillus reuteri HFI-LD5 and Lactobacillus rhamnosus HFI-K2 may be used as probiotics, especially in the treatment of small intestinal bacterial overgrowth (SIBO).

  8. DIRECT FLOW-CYTOMETRY OF ANAEROBIC-BACTERIA IN HUMAN FECES

    NARCIS (Netherlands)

    VANDERWAAIJ, LA; MESANDER, G; LIMBURG, PC; VANDERWAAIJ, D

    1994-01-01

    We describe a flow cytometry method for analysis of noncultured anaerobic bacteria present in human fecal suspensions. Nonbacterial fecal compounds, bacterial fragments, and large aggregates could be discriminated from bacteria by staining with propidium iodide (PI) and setting a discriminator on PI

  9. Carbon isotopes profiles of human whole blood, plasma, red blood cells, urine and feces for biological/biomedical 14C-accelerator mass spectrometry applications.

    Science.gov (United States)

    Kim, Seung-Hyun; Chuang, Jennifer C; Kelly, Peter B; Clifford, Andrew J

    2011-05-01

    Radiocarbon ((14)C) is an ideal tracer for in vivo human ADME (absorption, distribution, metabolism, elimination) and PBPK (physiological-based pharmacokinetic) studies. Living plants peferentially incorporate atmospheric (14)CO(2) versus (13)CO(2) versus (12)CO(2), which result in unique signature. Furthermore, plants and the food chains they support also have unique carbon isotope signatures. Humans, at the top of the food chain, consequently acquire isotopic concentrations in the tissues and body fluids depending on their dietary habits. In preparation of ADME and PBPK studies, 12 healthy subjects were recruited. The human baseline (specific to each individual and their diet) total carbon (TC) and carbon isotope (13)C (δ(13)C) and (14)C (F(m)) were quantified in whole blood (WB), plasma, washed red blood cell (RBC), urine, and feces. TC (mg of C/100 μL) in WB, plasma, RBC, urine, and feces were 11.0, 4.37, 7.57, 0.53, and 1.90, respectively. TC in WB, RBC, and feces was higher in men over women, P < 0.05. Mean δ(13)C were ranked low to high as follows: feces < WB = plasma = RBC = urine, P < 0.0001. δ(13)C was not affected by gender. Our analytic method shifted δ(13)C by only ±1.0 ‰ ensuring our F(m) measurements were accurate and precise. Mean F(m) were ranked low to high as follows: plasma = urine < WB = RBC = feces, P < 0.05. F(m) in feces was higher for men over women, P < 0.05. Only in WB, (14)C levels (F(m)) and TC were correlated with one another (r = 0.746, P < 0.01). Considering the lag time to incorporate atmospheric (14)C into plant foods (vegetarian) and or then into animal foods (nonvegetarian), the measured F(m) of WB in our population (recruited April 2009) was 1.0468 ± 0.0022 (mean ± SD), and the F(m) of WB matched the (extrapolated) atmospheric F(m) of 1.0477 in 2008. This study is important in presenting a procedure to determine a baseline for a study group for human ADME and PBPK studies using (14)C as a tracer.

  10. Isolation of Klebsiella terrigena from human feces: biochemical reactions, capsule types, and antibiotic sensitivity.

    Science.gov (United States)

    Podschun, R

    1991-04-01

    Colonization of the human intestinal tract by a newly proposed species, K. terrigena, was investigated. 5377 different stool specimens from healthy persons (food handlers) yielded 50 isolates (0.9%). Biochemically, low frequencies in the degradation of urea, dulcitol, and utilization of citrate at 37 degrees C were found when compared to K. pneumoniae. At 30 degrees C, urea hydrolysis was observed twice as often as at 37 degrees C. Apart from ampicillin, K. terrigena was susceptible to 12 other antimicrobial drugs tested. Multiple drug resistance was rare, few isolates being resistant against 2-4 antibiotic agents. Capsule typing revealed 30 different serotypes, K 70 and K 14 were the most frequent. Six strains expressed capsule types K 2 and K 5, which have been reported to be associated with virulence in K. pneumoniae. A possible pathogenic role of K. terrigena is discussed.

  11. An isotope-labeled chemical derivatization method for the quantitation of short-chain fatty acids in human feces by liquid chromatography–tandem mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Han, Jun; Lin, Karen; Sequeira, Carita [University of Victoria – Genome BC Proteomics Centre, University of Victoria, Vancouver Island Technology Park, 3101–4464 Markham Street, Victoria, BC V8Z 7X8 (Canada); Borchers, Christoph H., E-mail: christoph@proteincentre.com [University of Victoria – Genome BC Proteomics Centre, University of Victoria, Vancouver Island Technology Park, 3101–4464 Markham Street, Victoria, BC V8Z 7X8 (Canada); Department of Biochemistry and Microbiology, University of Victoria, Petch Building Room 207, 3800 Finnerty Road, Victoria, BC V8P 5C2 (Canada)

    2015-01-07

    Highlights: • 3-Nitrophenylhydrazine was used to derivatize short-chain fatty acids (SCFAs) for LC-MS/MS. • {sup 13}C{sub 6} analogues were produced for use as isotope-labeled internal standards. • Isotope-labeled standards compensate for ESI matrix effects in LC-MS/MS. • Femtomolar sensitivities and 93–108% quantitation accuracy were achieved for human fecal SCFAs. - Abstract: Short-chain fatty acids (SCFAs) are produced by anaerobic gut microbiota in the large bowel. Qualitative and quantitative measurements of SCFAs in the intestinal tract and the fecal samples are important to understand the complex interplay between diet, gut microbiota and host metabolism homeostasis. To develop a new LC-MS/MS method for sensitive and reliable analysis of SCFAs in human fecal samples, 3-nitrophenylhydrazine (3NPH) was employed for pre-analytical derivatization to convert ten C{sub 2}–C{sub 6} SCFAs to their 3-nitrophenylhydrazones under a single set of optimized reaction conditions and without the need of reaction quenching. The derivatives showed excellent in-solution chemical stability. They were separated on a reversed-phase C{sub 18} column and quantitated by negative-ion electrospray ionization – multiple-reaction monitoring (MRM)/MS. To achieve accurate quantitation, the stable isotope-labeled versions of the derivatives were synthesized in a single reaction vessel from {sup 13}C{sub 6}-3NPH, and were used as internal standard to compensate for the matrix effects in ESI. Method validation showed on-column limits of detection and quantitation over the range from low to high femtomoles for the ten SCFAs, and the intra-day and inter-day precision for determination of nine of the ten SCFAs in human fecal samples was ≤8.8% (n = 6). The quantitation accuracy ranged from 93.1% to 108.4% (CVs ≤ 4.6%, n = 6). This method was used to determine the SCFA concentrations and compositions in six human fecal samples. One of the six samples, which was collected from a

  12. Structure of the gut microbiome following colonization with human feces determines colonic tumor burden.

    Science.gov (United States)

    Baxter, Nielson T; Zackular, Joseph P; Chen, Grace Y; Schloss, Patrick D

    2014-01-01

    A growing body of evidence indicates that the gut microbiome plays a role in the development of colorectal cancer (CRC). Patients with CRC harbor gut microbiomes that are structurally distinct from those of healthy individuals; however, without the ability to track individuals during disease progression, it has not been possible to observe changes in the microbiome over the course of tumorigenesis. Mouse models have demonstrated that these changes can further promote colonic tumorigenesis. However, these models have relied upon mouse-adapted bacterial populations and so it remains unclear which human-adapted bacterial populations are responsible for modulating tumorigenesis. We transplanted fecal microbiota from three CRC patients and three healthy individuals into germ-free mice, resulting in six structurally distinct microbial communities. Subjecting these mice to a chemically induced model of CRC resulted in different levels of tumorigenesis between mice. Differences in the number of tumors were strongly associated with the baseline microbiome structure in mice, but not with the cancer status of the human donors. Partitioning of baseline communities into enterotypes by Dirichlet multinomial mixture modeling resulted in three enterotypes that corresponded with tumor burden. The taxa most strongly positively correlated with increased tumor burden were members of the Bacteroides, Parabacteroides, Alistipes, and Akkermansia, all of which are Gram-negative. Members of the Gram-positive Clostridiales, including multiple members of Clostridium Group XIVa, were strongly negatively correlated with tumors. Analysis of the inferred metagenome of each community revealed a negative correlation between tumor count and the potential for butyrate production, and a positive correlation between tumor count and the capacity for host glycan degradation. Despite harboring distinct gut communities, all mice underwent conserved structural changes over the course of the model. The

  13. Investigation of intestinal parasites in pig feces that are also human pathogens.

    Science.gov (United States)

    Uysal, Hayriye Kirkoyun; Boral, Ozden; Metiner, Kemal; Ilgaz, Atilla

    2009-01-01

    A total of 238 pig fecal specimens were collected from pig farms in Corlu (Tekirdağ), Ayazma, and Arnavutköy (Istanbul) during the summer. Out of the 238 pig specimens, 105 were from pigs younger than 6 months and 133 from pigs older than 6 months. These were investigated for intestine parasites in particular the ones that are human pathogens. Cryptosporidium spp. was detected In 21 fecal specimens (8.8%), Giardia spp. in 9 (3.7%), Balantidium coli cysts in 4 (1.6%) and Ascaris suum eggs in 9 (4.1%). Giardia lamblia were found in 8 (7.6%) of 105 pigs younger than 6 months, Cryptosporidium spp. in 12 (11.4%), Balantidium coli cysts in 2 (1.5%). In the pigs older than 6 months Giardia lamblia were found in 1 (0.7%), Cryptosporidium spp. in 9 (6.7%), Balantidium coli cysts in 2 (1.5%). and Ascaris suum eggs in 9 (6.7%). The difference in the rate of G. lamblia (p=0.01) in pigs less than 6 months and of A. suum in those over 6 months was found to be statistically significant (p=0.005). Our results revealed that pigs are important sources of these parasites.

  14. Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

    OpenAIRE

    Gunilla Veslemøy Schmidt; Anders Mellerup; Lasse Engbo Christiansen; Marie Ståhl; John Elmerdahl Olsen; Øystein Angen

    2015-01-01

    The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling an...

  15. Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

    DEFF Research Database (Denmark)

    Schmidt, Gunilla Veslemøy; Mellerup, Anders; Christiansen, Lasse Engbo;

    2015-01-01

    in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes...... when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same...

  16. Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

    DEFF Research Database (Denmark)

    Schmidt, Gunilla Veslemøy; Mellerup, Anders; Christiansen, Lasse Engbo;

    2015-01-01

    for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined...... in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes......The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays...

  17. Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

    DEFF Research Database (Denmark)

    Schmidt, Gunilla Veslemøy; Mellerup, Anders; Christiansen, Lasse Engbo

    2015-01-01

    The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays...... for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined...... in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes...

  18. 应用real-time PCR法快速定量人类粪便中双歧杆菌的研究%A fast quantification for Bifidobacteria in human feces by real-time PCR

    Institute of Scientific and Technical Information of China (English)

    于静; 文姝; 唐立

    2009-01-01

    Objective To build a fast and accurate method of quantification for Bifidobacteria in human feces. Method Traditional culture, common PCR, real-time PCR were used. Result (1) The inhibitors in the feces samples were removed when the samples were centrifuged, washed and diluted,so that the Bifidobacteria in the feces samples could be directly quantitated by PCR and real-time PCR. (2) The direct semiquantitation by PCR established in this study showed a satisfactory distinguishability when Bifidobacteria in feces samples was between 10~3 ~10~7CFU /ml. There was no obvious difference between the result from RT-PCR quantification and that from cultured method (P > 0.05); The direct quantita-tion by PCR showed a satisfactory distinguishability when Bifidobacteria in feces samples was between 10~1 ~ 10~7CFU /ml, without obvious difference between the result from RT-PCR and that from cultured method (P > 0. 05). Conclusion Bifidobacteria in feces samples can be directly quantitated or semiquantitated by real-time PCR method or common PCR method.%目的 建立快速、准确从粪便标本中定量双歧杆菌的RT-PCR技术.方法 传统培养定量法,普通PCR定量法,real-time PCR 比较测量.结果 (1)粪便标本前处理采取简单的离心和清洗、稀释步骤能去除粪便标本中的抑制物,实现不提取 DNA直接进行PCR、real-time定量粪便中双歧杆菌.(2)本实验建立的PCR方法直接半定量粪便双歧杆菌技术在双歧杆菌值介于10~3~10~7CFU/ml时具有较好的分辨率,粪便标本普通PCR得理论菌数与培养得菌数值之间差异无显著性(P>0.05);real-time PCR直接定量双歧杆菌技术在双歧杆菌值介于10~1~10~7CFU /ml时具有较好的分辨率,粪便标本RT-PCR得理论菌数与培养得菌数值之间差异无显著性(P>0.05).结论 利用PCR、real-time PCR直接半定量和定量粪便中的双歧杆菌可行.

  19. Post-operative corticosterone levels in plasma and feces of mice subjected to permanent catheterization and automated blood sampling.

    Science.gov (United States)

    Sundbom, Renée; Jacobsen, Kirsten R; Kalliokoski, Otto; Hau, Jann; Abelson, Klas S P

    2011-01-01

    This study investigated the effects of surgical placement of permanent arterial catheters on plasma corticosterone levels, fecal corticosterone excretion and body weight in male BALB/c/Sca mice. In addition, the effects of voluntarily ingested buprenorphine in doses of 0.5 and 1.0 mg/kg body weight on these parameters were studied. A catheter was placed in the carotid artery during isoflurane anesthesia. Immediately after surgery, the mice were connected to an AccuSampler® μ and blood samples for plasma corticosterone quantification were collected automatically during the first 24 h postoperatively. All fecal boli produced 24 h before and 24 h after surgery were collected for fecal corticosterone excretion measures and the pre- and post-operative body weights were registered. Plasma corticosterone levels were in the range of 150-300 ng/ml after the surgical procedure and the body weight was significantly lower 24 h after surgery compared to its pre-operative value. Contrary to what was expected, the total fecal corticosterone excretion was significantly reduced 24 h after surgery, as was the defecation. Buprenorphine treatment significantly lowered the plasma corticosterone levels, but had no effect on fecal corticosterone excretion or body weight change. It was concluded that surgical placement of an arterial catheter induces a significant stress response, as judged by its effect on plasma corticosterone and body weight. Voluntary ingestion of buprenorphine improved postoperative recovery by lowering plasma corticosterone concentrations. Neither fecal corticosterone excretion nor body weight change seems suitable for postoperative stress assessment in mice in the present experimental setup.

  20. Seasonal concentrations of cesium-137 in rumen content, skeletal muscles and feces of caribou from the Porcupine herd: lichen ingestion rates and implications for human consumption

    Directory of Open Access Journals (Sweden)

    A. C. Allaye-Chan

    1990-09-01

    Full Text Available The Porcupine caribou herd was monitored for cesium-137 during 1987 to address human health concerns over potential meat contamination by radioactive fallout from the Chernobyl accident, and to determine lichen intake rates based on body burdens of radiocesium. A total of 36 caribou were collected from northwestern Alaska and the Yukon Territories in March, June, September, and November. Mean radiocesium concentrations in skeletal muscle peaked in March at 133 Bq/kg fresh weight. This value should not prove hazardous to human health. Radiocesium concentrations in skeletal muscle (wet weight ranged from approximately 22 to 50% of radiocesium concentrations in rumen contents (dry weight, and from approximately 15 to 37% of radiocesium concentrations in feces (dry weight. Radioactivity in feces was significantly correlated with radioactivity in rumen contents. Computer simulations relating lichen intake rates to radiocesium body burdens are presented for 3 scenarios: (1 when seasonal intakes were adjusted to provide the optimum fit between simulated and observed radiocesium body burdens (2 when seasonal intakes were based on empirical data, and (3 when seasonal intakes were adjusted to yield a "conventional" radiocesium curve of a slow fall build-up prior to a late winter plateau.

  1. Bacteroides uniformis is a putative bacterial species associated with the degradation of the isoflavone genistein in human feces.

    Science.gov (United States)

    Renouf, Mathieu; Hendrich, Suzanne

    2011-06-01

    Inter-individual variation in isoflavone absorption depends on gut microbial degradation and affects the efficacy of these compounds. We hypothesized that inter-individual variation in fecal isoflavone disappearance coincided with variation in bacterial species. In vitro anaerobic fecal disappearance of isoflavones was measured from 33 participants by HPLC. Fecal microbial 16S rRNA variable region PCR products were obtained from 4 participants with the greatest and least genistein or glycitein degradation and were subjected to denaturing gradient gel electrophoresis. DNA bands with a homology of 90-95% to Bacteroides uniformis and Faecalibacterium prausnitzii were present in greater intensities in fecal samples showing a genistein disappearance rate constant of 1.47 ± 0.14 h(-1) compared with those with a genistein disappearance rate constant of 0.15 ± 0.03 h(-1) (P < 0.05). Human fecal bacterial species with DNA sequences 90-100% homologous to Tannerella forsythensis and 4 other species were present in greater intensities in fecal samples showing a glycitein disappearance rate constant of 0.57 ± 0.30 h(-1) compared with fecal samples with a glycitein disappearance rate constant of 0.08 ± 0.03 h(-1) (P < 0.05). In high degraders, B. uniformis may be a candidate for genistein degradation and T. forsythensis for glycitein degradation, based on fecal isoflavone degradation in the presence of these species. Bacteroides acidifaciens increased isoflavone disappearance in anaerobic human fecal incubations under nutrient-rich and -depleted conditions, suggesting this species as one responsible for the generally high degradation of isoflavones by humans. These fecal microbes are candidate biomarkers for interindividual variation in isoflavone uptake and efficacy.

  2. Comparison of individual and pooled samples for quantification of antimicrobial resistance genes in swine feces by high-throughput qPCR

    DEFF Research Database (Denmark)

    Clasen, Julie; Mellerup, Anders; Olsen, John Elmerdahl

    2015-01-01

    There is a considerable societal interest in the careful monitoring of antimicrobial resistance (AMR) levels in human and animal populations. Sampling and data analysis can be both costly and time consuming. Optimization of sample pooling procedures is therefore important to reduce costs and anal......There is a considerable societal interest in the careful monitoring of antimicrobial resistance (AMR) levels in human and animal populations. Sampling and data analysis can be both costly and time consuming. Optimization of sample pooling procedures is therefore important to reduce costs...... samples were taken from each pen with respect to the number of pigs in the pen. A total of 48 pools were made of increasing number of individual samples. The levels of 9 different AMR-genes were quantified using dynamic qPCR arrays on the BioMark HD system(Fluidigm®).DNA was extracted using the Maxwell...

  3. Performance of VIDISCA-454 in Feces-Suspensions and Serum

    Directory of Open Access Journals (Sweden)

    Lia van der Hoek

    2012-08-01

    Full Text Available Virus discovery combining sequence unbiased amplification with next generation sequencing is now state-of-the-art. We have previously determined that the performance of the unbiased amplification technique which is operational at our institute, VIDISCA-454, is efficient when respiratory samples are used as input. The performance of the assay is, however, not known for other clinical materials like blood or stool samples. Here, we investigated the sensitivity of VIDISCA-454 with feces-suspensions and serum samples that are positive and that have been quantified for norovirus and human immunodeficiency virus type 1, respectively. The performance of VIDISCA-454 in serum samples was equal to its performance in respiratory material, with an estimated lower threshold of 1,000 viral genome copies. The estimated threshold in feces-suspension is around 200,000 viral genome copies. The decreased sensitivity in feces suspension is mainly due to sequences that share no recognizable identity with known sequences. Most likely these sequences originate from bacteria and phages which are not completely sequenced.

  4. Metabolomic analysis of human fecal microbiota: a comparison of feces-derived communities and defined mixed communities.

    Science.gov (United States)

    Yen, Sandi; McDonald, Julie A K; Schroeter, Kathleen; Oliphant, Kaitlyn; Sokolenko, Stanislav; Blondeel, Eric J M; Allen-Vercoe, Emma; Aucoin, Marc G

    2015-03-01

    The extensive impact of the human gut microbiota on its human host calls for a need to understand the types of communication that occur among the bacteria and their host. A metabolomics approach can provide a snapshot of the microbe-microbe interactions occurring as well as variations in the microbes from different hosts. In this study, metabolite profiles from an anaerobic continuous stirred-tank reactors (CSTR) system supporting the growth of several consortia of bacteria representative of the human gut were established and compared. Cell-free supernatant samples were analyzed by 1D (1)H nuclear magnetic resonance (NMR) spectroscopy, producing spectra representative of the metabolic activity of a particular community at a given time. Using targeted profiling, specific metabolites were identified and quantified on the basis of NMR analyses. Metabolite profiles discriminated each bacterial community examined, demonstrating that there are significant differences in the microbiota metabolome between each cultured community. We also found unique compounds that were identifying features of individual bacterial consortia. These findings are important because they demonstrate that metabolite profiles of gut microbial ecosystems can be constructed by targeted profiling of NMR spectra. Moreover, examination of these profiles sheds light on the type of microbes present in the gut and their metabolic interactions.

  5. Escherichia coli isolated from feces of brown bears (Ursus arctos) have a lower prevalence of human extraintestinal pathogenic E. coli virulence-associated genes.

    Science.gov (United States)

    Vadnov, Maruša; Barbič, Damjana; Žgur-Bertok, Darja; Erjavec, Marjanca Starčič

    2017-01-01

    Eighty-six Escherichia coli strains from feces of either wild brown bears or those living in a zoo were screened for phylogenetic groups using the revisited Clermont phylotyping method and the prevalence of 24 virulence-associated genes (VAGs) of extraintestinal pathogenic E. coli (ExPEC). Our results showed that most strains of E. coli in bears belonged to phylogenetic groups III/IV/V (29%) and B1 (26%). Only half of the tested VAGs were found in the E. coli bear strains, with fimH present in 72%, ompT in 63%, and kpsMT in 43% of the strains. When the data obtained on the fecal E. coli strains from brown bears were compared with the data obtained on 90 fecal E. coli strains from healthy humans, there were significant differences in E. coli population structures between both hosts.

  6. Increased Butyrate Production During Long-Term Fermentation of In Vitro-Digested High Amylose Cornstarch Residues with Human Feces.

    Science.gov (United States)

    Li, Li; Jiang, Hongxin; Kim, Hyun-Jung; Yum, Man-Yu; Campbell, Mark R; Jane, Jay-Lin; White, Pamela J; Hendrich, Suzanne

    2015-09-01

    An in vitro semi-continuous long-term (3 wk) anaerobic incubation system simulating lower gut fermentation was used to determine variability in gut microbial metabolism between 4 predigested high amylose-resistant starch residues (SR): SRV, SRVI, SRVII, and SRGEMS in human fecal samples. Subjects participated twice, 5 mo apart: 30 in Phase I (15 lean, 9 overweight and 6 obese), 29 in Phase II (15 lean, 9 overweight, 5 obese); 13 of 15 lean subjects participated in both phases. Of the 4 SRs, SRV displayed the highest gelatinization temperature, peak temperature, enthalpy changes, and the least digestibility compared with the other SRs. In both phases, compared with blank controls, all SRs increased butyrate ∼2-fold which stabilized at week 2 and only SRV caused greater propionate concentration (∼30%) after 3 wk which might have been partly mediated by its lesser digestibility. Fecal samples from lean and overweight/obese subjects incubated with SRs showed similar short-chain fatty acid production across both time points, which suggests that resistant starch may benefit individuals across BMIs.

  7. Pervasive Environmental Contamination with Human Feces Results in High Prevalence of Zoonotic Sarcocystis Infection in Pigs in the Punjab, India.

    Science.gov (United States)

    Kaur, M; Singh, B B; Sharma, R; Gill, J P S

    2016-04-01

    Three species of Sarcocystis-S. miescheriana, S. suihominis, and S. porcifelis-have been recorded from pigs ( Sus scrofa ). Among these 3 species, the zoonotic species S. suihominis is of paramount importance and an important food safety issue. Previous studies indicate prevalence of porcine Sarcocystis species in India, but molecular evidence, among other evidence, is required for proper species differentiation. Myocardium from 250 stray and farm pigs destined for slaughter for human consumption were collected from slaughter shops located in urban slums in Punjab, northern India. Tissues were examined for Sarcocystis by using an intact cyst isolation method, pepsin acid digestion, Sarcocystis 18S ribosomal RNA polymerase chain reaction (PCR), and real-time quantitative PCR with melting curve analysis (qPCR-MCA). The combination of primers was used for 18S rRNA PCR amplification followed by sequencing. Ten representative samples were sequenced in both the directions from which 7 readable sequences were obtained for phylogenetic analysis. Sarcocystis cysts/zoites were recorded in 146 (58.4%), 169 (67.6%), 182 (72.8%), and 191 (76.4%) of samples by using intact cyst isolation, pepsin HCl digestion, conventional PCR, and qPCR-MCA, respectively. Molecularly, 1 S. miescheriana isolate and 6 isolates of the zoonotic species S. suihominis were recorded. This is the first study providing molecular identification for the presence of zoonotic species S. suihomonis in India. The prevalence of zoonotic S. suihominis in pork in India is worrisome and warrants intervention policies to stop the practice of rearing pigs under unhygienic conditions.

  8. Isolation of Bacteroides from fish and human fecal samples for identification of unique molecular markers.

    Science.gov (United States)

    Kabiri, Leila; Alum, Absar; Rock, Channah; McLain, Jean E; Abbaszadegan, Morteza

    2013-12-01

    Bacteroides molecular markers have been used to identify human fecal contamination in natural waters, but recent work in our laboratory confirmed cross-amplification of several human-specific Bacteroides spp. assays with fecal DNA from fish. For identification of unique molecular markers, Bacteroides from human (n = 4) and fish (n = 7) fecal samples were cultured and their identities were further confirmed using Rapid ID 32A API strips. The 16S rDNA from multiple isolates from each sample was PCR amplified, cloned, and sequenced to identify unique markers for development of more stringent human-specific assays. In human feces, Bacteroides vulgatus was the dominant species (75% of isolates), whereas in tilapia feces, Bacteroides eggerthii was dominant (66%). Bacteroides from grass carp, channel catfish, and blue catfish may include Bacteroides uniformis, Bacteroides ovatus, or Bacteroides stercoris. Phylogenic analyses of the 16S rRNA gene sequences showed distinct Bacteroides groupings from each fish species, while human sequences clustered with known B. vulgatus. None of the fish isolates showed significant similarity to Bacteroides sequences currently deposited in NCBI (National Center for Biotechnology Information). This study expands the current sequence database of cultured fish Bacteroides. Such data are essential for identification of unique molecular markers in human Bacteroides that can be utilized in differentiating fish and human fecal contamination in water samples.

  9. Genome Sequence of "Candidatus Methanomassiliicoccus intestinalis" Issoire-Mx1, a Third Thermoplasmatales-Related Methanogenic Archaeon from Human Feces.

    Science.gov (United States)

    Borrel, Guillaume; Harris, Hugh M B; Parisot, Nicolas; Gaci, Nadia; Tottey, William; Mihajlovski, Agnès; Deane, Jennifer; Gribaldo, Simonetta; Bardot, Olivier; Peyretaillade, Eric; Peyret, Pierre; O'Toole, Paul W; Brugère, Jean-François

    2013-07-11

    "Candidatus Methanomassiliicoccus intestinalis" Issoire-Mx1 is a methanogenic archaeon found in the human gut and is a representative of the novel order of methanogens related to Thermoplasmatales. Its complete genome sequence is presented here.

  10. CE-LIF-MSn profiling of oligosaccharides in human milk and feces of breast-fed babies

    NARCIS (Netherlands)

    Albrecht, S.A.; Schols, H.A.; Heuvel, van den E.G.H.M.; Voragen, A.G.J.; Gruppen, H.

    2010-01-01

    Mixtures of the complex human milk oligosaccharides (HMOs) are difficult to analyze and gastrointestinal bioconversion products of HMOs may complicate analysis even more. Their analysis, therefore, requires the combination of a sensitive and high-resolution separation technique with a mass identific

  11. Occurrence of bacteriophages infecting Bacteroides host strains (ARABA 84 and GB-124) in fecal samples of human and animal origin.

    Science.gov (United States)

    Diston, David; Wicki, Melanie

    2015-09-01

    Bacteriophage-based microbial source-tracking studies are an economical and simple way of identifying fecal sources in polluted water systems. Recently isolated Bacteroides spp. strains ARABA 84, and GB-124 have been shown to detect bacteriophages exclusively in aquatic systems impacted by human fecal material. To date, limited examination of the occurrence or concentration of phages capable of infecting Bacteroides fragilis strain GB-124 or B. thetaiotaomicron strain ARABA 84 in human and animal feces has been carried out. This study reports the prevalence rates and concentrations of phages infecting ARABA 84 and GB-124 host strains in human and a range of animal feces. Discrete human fecal samples (n=55) and pooled animal samples (n=46, representing the feces of over 230 animals) were examined for phages infecting the host strains ARABA 84, GB-124, and E. coli strain WG5. Both human Bacteroides host strains were highly specific (95% and 100% for ARABA 84 and GB-124, respectively), challenging results from previous studies. This study supports the use of Bacteroides strains GB-124 and ARABA 84 in fecal source tracking studies for the detection of human fecal contamination.

  12. Multiple Zoonotic Parasites Identified in Dog Feces Collected in Ponte de Lima, Portugal—A Potential Threat to Human Health

    Directory of Open Access Journals (Sweden)

    Teresa Letra Mateus

    2014-09-01

    Full Text Available Dogs play many roles and their presence within people’s houses has increased. In rural settings dog faeces are not removed from the streets, representing an environmental pollution factor. Our aim was to evaluate the occurrence of environmental contamination with zoonotic intestinal parasites of three groups of dogs in Ponte de Lima, Portugal, with a particular emphasis on Echinococcus granulosus. We collected 592 dog faecal samples from the environment, farm and hunting dogs. Qualitative flotation coprological analysis was performed and the frequency in the positive samples ranged between 57.44% and 81.19% in different groups. We isolated up to four different parasites in one sample and detected seven intestinal parasitic species, genera or families overall. Ancylostomatidae was the most prevalent parasite, followed by Trichuris spp., Toxocara spp., Isospora spp., Dipylidium caninum, Taeniidae and Toxascaris leonina. Taeniidae eggs were analyzed with the PCR technique and revealed not to be from Echinococcus. The parasite prevalence and the diversity of zoonotic parasites found were high, which calls for a greater awareness of the problem among the population, especially hunters. Promoting research at the local level is important to plan control strategies. Health education should be developed with regard to farmers and hunters, and a closer collaboration between researchers, practitioners and public health authorities is needed.

  13. Multiple zoonotic parasites identified in dog feces collected in Ponte de Lima, Portugal-a potential threat to human health.

    Science.gov (United States)

    Mateus, Teresa Letra; Castro, António; Ribeiro, João Niza; Vieira-Pinto, Madalena

    2014-09-01

    Dogs play many roles and their presence within people's houses has increased. In rural settings dog faeces are not removed from the streets, representing an environmental pollution factor. Our aim was to evaluate the occurrence of environmental contamination with zoonotic intestinal parasites of three groups of dogs in Ponte de Lima, Portugal, with a particular emphasis on Echinococcus granulosus. We collected 592 dog faecal samples from the environment, farm and hunting dogs. Qualitative flotation coprological analysis was performed and the frequency in the positive samples ranged between 57.44% and 81.19% in different groups. We isolated up to four different parasites in one sample and detected seven intestinal parasitic species, genera or families overall. Ancylostomatidae was the most prevalent parasite, followed by Trichuris spp., Toxocara spp., Isospora spp., Dipylidium caninum, Taeniidae and Toxascaris leonina. Taeniidae eggs were analyzed with the PCR technique and revealed not to be from Echinococcus. The parasite prevalence and the diversity of zoonotic parasites found were high, which calls for a greater awareness of the problem among the population, especially hunters. Promoting research at the local level is important to plan control strategies. Health education should be developed with regard to farmers and hunters, and a closer collaboration between researchers, practitioners and public health authorities is needed.

  14. Comparative fermentation of insoluble carbohydrates in an in vitro human feces model spiked with Lactobacillus acidophilus NCFM

    DEFF Research Database (Denmark)

    Knudsen, Anne; van Zanten, Gabriella C.; Jensen, Susanne L.

    2013-01-01

    in lactic acid production. Potato lintner starch had the greatest effect. Insoluble carbohydrates also suppressed production of SCFAs as compared to the control medium. Importantly, potato lintner starch most efficiently suppressed the ratio between the Bacteroidetes and Firmicutes and suppressed growth...... maize starch granules, pectin‐rich potato fiber, and potato lintner starch tested with human fresh fecal microbiota spiked with the probiotic Lactobacillus acidophilus NCFM. Microbial quantification by real‐time polymerase chain‐reaction (qRT‐PCR) revealed that Bacteroidetes was specifically suppressed...... by each insoluble carbohydrate resulting in a clear decrease in the ratio of Bacteroidetes and Firmicutes. Notably, all carbohydrates tested appeared to block the formation of the potentially harmful branched chain fatty acids (BCFA) fermentation products, but supported lactobacilli growth and increase...

  15. Antibiotic resistance genes in the bacteriophage DNA fraction of human fecal samples.

    Science.gov (United States)

    Quirós, Pablo; Colomer-Lluch, Marta; Martínez-Castillo, Alexandre; Miró, Elisenda; Argente, Marc; Jofre, Juan; Navarro, Ferran; Muniesa, Maite

    2014-01-01

    A group of antibiotic resistance genes (ARGs) (blaTEM, blaCTX-M-1, mecA, armA, qnrA, and qnrS) were analyzed by real-time quantitative PCR (qPCR) in bacteriophage DNA isolated from feces from 80 healthy humans. Seventy-seven percent of the samples were positive in phage DNA for one or more ARGs. blaTEM, qnrA, and, blaCTX-M-1 were the most abundant, and armA, qnrS, and mecA were less prevalent. Free bacteriophages carrying ARGs may contribute to the mobilization of ARGs in intra- and extraintestinal environments.

  16. Identification of feces by detection of Bacteroides genes.

    Science.gov (United States)

    Nakanishi, Hiroaki; Shojo, Hideki; Ohmori, Takeshi; Hara, Masaaki; Takada, Aya; Adachi, Noboru; Saito, Kazuyuki

    2013-01-01

    In forensic science, the identification of feces is very important in a variety of crime investigations. However, no sensitive and simple fecal identification method using molecular biological techniques has been reported. Here, we focused on the fecal bacteria, Bacteroides uniformis, Bacteroides vulgatus and Bacteroides thetaiotaomicron, and developed a novel fecal identification method by detection of the gene sequences specific to these bacteria in various body (feces, blood, saliva, semen, urine, vaginal fluids and skin surfaces) and forensic (anal adhesions) specimens. Bacterial gene detection was performed by real-time PCR using a minor groove binding probe to amplify the RNA polymerase β-subunit gene of B. uniformis and B. vulgatus, and the α-1-6 mannanase gene of B. thetaiotaomicron. At least one of these bacteria was detected in the feces of 20 donors; the proportions of B. uniformis, B. vulgatus and B. thetaiotaomicron were 95, 85 and 60%, respectively. Bacteroides vulgatus was also detected in one of six vaginal fluid samples, but B. thetaiotaomicron and B. uniformis were not detected in body samples other than feces. Further, we applied this method to forensic specimens from 18 donors. Eighteen anal adhesions also contained at least one of three bacteria; B. uniformis, B. vulgatus and B. thetaiotaomicron were detected in 89, 78 and 56%, respectively, of the specimens. Thus, these bacteria were present at a high frequency in the fecal and forensic specimens, while either B. uniformis or B. vulgatus was detected in all samples. Therefore, B. uniformis and B. vulgatus represent more appropriate target species than B. thetaiotaomicron for the identification of fecal material. If B. vulgatus and/or B. uniformis are detected, it is likely that the sample contains feces. Taken together, our results suggest that the use of molecular biological techniques will aid the detection of feces in forensic practice, although it is possible that the samples contained

  17. Detection of vancomycin resistance in enterococcus species isolated from clinical samples and feces of colonized patients by phenotypic and genotypic methods

    Directory of Open Access Journals (Sweden)

    Priyanka Paul Biswas

    2016-01-01

    Full Text Available Background: The aim of this study was to find out the clinical correlation between the presence of vancomycin-resistant genes (van A and van B and their expression as detected by phenotypic tests in colonized patients and in clinical isolates. Materials and Methods: Enterococci were isolated from various clinical samples and also from fecal specimens of colonized patients at the time of admission, after 48 h and after 5 days of admission. Identification to species level was done using standard methods. Vancomycin susceptibility in Enterococci was detected by disc diffusion test. Minimum inhibitory concentration was determined by agar dilution method. Multiplex polymerase chain reaction (PCR was used to detect the presence of van genes. Results: Out of all the clinical and fecal samples processed, 12.0% isolates were either vancomycin resistant or vancomycin intermediate. Further, these isolates carried van A or van B genes as confirmed by PCR methods. Expression of van A gene was found to be more in Enterococcus faecalis (28.3% as compared to Enterococcus faecium (25.0% in both clinical and fecal isolates. 16.6% strains of E. faecium and 15.0% strains each of E. faecalis and Enterococcus gallinarum were found to carry van B genes. The overall prevalence of vancomycin resistant Enterococci (VRE in colonized patients was about 9.6%. Prior administration of antibiotics had significant effect (P = 0.001 on VRE carriage. Urinary tract infection was the most common infection caused by vancomycin susceptible Enterococci (VSE, 105/214 (49.0% and VRE, 13/36 (36.1%. There was no significant difference (P = 0.112 in the distribution of VRE and VSE in different infection types. Both clinical and fecal VRE showed maximum resistance to penicillin, ampicillin, and piperacillin. Resistance to linezolid was 2.8% in clinically isolated VRE. Conclusion: VRE in our study were found to be resistant to a number of commonly used antibiotics. The frequency of isolation

  18. 电感耦合等离子体质谱法测定成人全血、膳食、尿样和粪便中轻稀土元素及相关性分析%Determination and Correlation Analysis of Light Rare Earth Elements in Human's Blood, Food, Urine and Feces by ICP-MS

    Institute of Scientific and Technical Information of China (English)

    解清; 欧阳荔; 王京宇

    2012-01-01

    The light rare earth element(REEs) contents in the blood, food, urine and feces of human bodies were determined by ICP-MS. All the samples were obtained from healthy adult men who lived in the four areas (Chengdu, Taiyuan, Tianjin and Zhenjiang) with different dietary patterns in China. All the individual samples were digested by mixed acid and determined by ICP-MS. Descending order of the light REEs concentrations were in feces, food, blood and urine. There was no significant difference on concentrations of the samples among the four cities. And the distribution pattern of the light REEs in blood, food and feces were similar to that in the nature, while the Sm in the urine was abnormal(REEs).%为了全面分析人体内稀土元素通过膳食摄取、血液蓄积、尿粪排出之间的关系,应用等离子体质谱法测定成人全血、膳食、尿样及粪便中轻稀土元素含量,比较4个不同膳食类型的城市人体摄取轻稀土元素的差异,并对相同个体的膳食、全血、尿样及粪便中的轻稀土元素进行相关性分析.分别在天津、成都、镇江、太原4个城市采集30名健康成年男性的全血、72 h膳食、72 h尿样和72 h粪便,膳食、尿样和粪便做冷冻干燥处理后经湿法消解,并采用电感耦合等离子体质谱法测定轻稀土元素含量.4城市采样人体中的稀土含量由高至低依次为粪便、膳食、血液,尿液,但上述样品在4城市人群之间没有显著性差异;同体血、膳食和粪便中轻稀土元素的分布特征符合自然分布特征规律,但尿样中的Sm存在正异常.

  19. Detection of parvoviruses in wolf feces by electron microscopy

    Science.gov (United States)

    Muneer, M.A.; Farah, I.O.; Pomeroy, K.A.; Goyal, S.M.; Mech, L.D.

    1988-01-01

    One hundred fifteen wolf (Canis lupus) feces were collected between 1980 and 1984 from northeastern Minnesota and were examined for canine parvovirus by negative contrast electron microscopy. Of these, seven (6%) samples revealed the presence of parvovirus. Some of these viruses were able to grow in cell cultures forming intranuclear inclusion bodies and giant cells.

  20. Escherichia coli O104 in Feedlot Cattle Feces: Prevalence, Isolation and Characterization.

    Science.gov (United States)

    Shridhar, Pragathi B; Noll, Lance W; Shi, Xiaorong; Cernicchiaro, Natalia; Renter, David G; Bai, J; Nagaraja, T G

    2016-01-01

    Escherichia coli O104:H4, an hybrid pathotype of Shiga toxigenic and enteroaggregative E. coli, involved in a major foodborne outbreak in Germany in 2011, has not been detected in cattle feces. Serogroup O104 with H type other than H4 has been reported to cause human illnesses, but their prevalence and characteristics in cattle have not been reported. Our objectives were to determine the prevalence of E. coli O104 in feces of feedlot cattle, by culture and PCR detection methods, and characterize the isolated strains. Rectal fecal samples from a total of 757 cattle originating from 29 feedlots were collected at a Midwest commercial slaughter plant. Fecal samples, enriched in E. coli broth, were subjected to culture and PCR methods of detection. The culture method involved immunomagnetic separation with O104-specific beads and plating on a selective chromogenic medium, followed by serogroup confirmation of pooled colonies by PCR. If pooled colonies were positive for the wzxO104 gene, then colonies were tested individually to identify wzxO104-positive serogroup and associated genes of the hybrid strains. Extracted DNA from feces were also tested by a multiplex PCR to detect wzxO104-positive serogroup and associated major genes of the O104 hybrid pathotype. Because wzxO104 has been shown to be present in E. coli O8/O9/O9a, wzxO104-positive isolates and extracted DNA from fecal samples were also tested by a PCR targeting wbdDO8/O9/O9a, a gene specific for E. coli O8/O9/O9a serogroups. Model-adjusted prevalence estimates of E. coli O104 (positive for wzxO104 and negative for wbdDO8/O9/O9a) at the feedlot level were 5.7% and 21.2%, and at the sample level were 0.5% and 25.9% by culture and PCR, respectively. The McNemar's test indicated that there was a significant difference (P German outbreak strain and 67.9% to 77.5% homology to human diarrheagenic O104:H7 strains. The 13 isolates obtained from the same feedlot were of the same PFGE subtype with 100% Dice similarity

  1. Prevalence and characterization of Salmonella enterica from the feces of cattle, poultry, swine and hedgehogs in Burkina Faso and their comparison to human Salmonella isolates

    OpenAIRE

    Kagambèga, Assèta; Lienemann, Taru; Aulu, Laura; Traoré, Alfred S; Barro, Nicolas; Siitonen, Anja; Haukka, Kaisa

    2013-01-01

    Background Production and wild animals are major sources of human salmonellosis and animals raised for food also play an important role in transmission of antimicrobial resistant Salmonella strains to humans. Furthermore, in sub-Saharan Africa non-typhoidal Salmonella serotypes are common bloodstream isolates in febrile patients. Yet, little is known about the environmental reservoirs and predominant modes of transmission of these pathogens. The purpose of this study was to discover potential...

  2. An improved qPCR protocol for rapid detection and quantification of Clostridium difficile in cattle feces.

    Science.gov (United States)

    Bandelj, Petra; Logar, Katarina; Usenik, Alenka M; Vengust, Modest; Ocepek, Matjaz

    2013-04-01

    Clostridium difficile (CD) can cause a significant and transmissible disease in animals and humans, with poorly understood epidemiology. Animals have been suggested as a possible source of infection and environment contamination. It is necessary that a precise and rapid diagnostic tool is available for the detection of CD from clinical and/or environmental samples. A quantitative real-time PCR (qPCR) protocol for CD detection defined by Penders et al. (FEMS Microbiol Lett, 243, 2005, 141-147) was modified. The modified protocol, supported by a novel extraction method, was tested on CD-spiked cattle feces and clinical fecal samples from calves. Quantification was performed targeting CD 16S rRNA gene. Three different commonly used TaqMan universal PCR master mixes were also compared. Results indicate that the modified protocol is very sensitive with an LOD of 7.72 CD cells per g CD-spiked feces. The protocol is capable of precise quantification with an LOQ of 77.2 CD cells per g CD-spiked feces, R(2) between 0.9957 and 0.9968, isolation efficiency from 87.89% to 90.96%, and an interassay CV ranging from 3.71% to 9.57%. The qPCR protocol for the detection and quantification of CD from animal feces investigated and described in this article using MIQE guidelines has the lowest detection and quantification limits published to date. Therefore, it can be implemented for precise epidemiological investigations of CD infections in animals and humans. © 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  3. Lack of specificity for PCR assays targeting human Bacteroides 16S rRNA gene: cross-amplification with fish feces

    Science.gov (United States)

    Methods focused on members of the genus Bacteroides have been increasingly utilized in microbial source tracking studies for identifying and quantifying sources of non-point fecal contamination. We present results using real-time PCR to show significant cross-amplification of a human-specific Bacter...

  4. Chimpanzee diet: phytolithic analysis of feces.

    Science.gov (United States)

    Phillips, Caroline; Lancelotti, Carla

    2014-08-01

    Most primate populations remain unobservable; therefore, researchers depend on the analyses of indirect evidence encountered at a study-site in order to understand their behavioral ecology. Diet can be determined through the analyses of scats or feeding remains encountered on-site. This allows aspects of their dietary repertoire to be established, which has implications both for conservation efforts (by locating food resources), and for understanding the evolution of hominin diet (if used as referential models). Macroscopic inspection of fecal samples is a common method applied to ascertain a primate population's diet. However, new approaches are required to identify food-items unrecognizable at this level. We applied a dry ash extraction method to fecal samples (N=50) collected from 10 adult chimpanzees in Kanyawara, Kibale National Park, Uganda and also to plant parts (N=66) from 34 species known to be included in the diet of this community of apes. We recovered phytoliths in 26 of the 34 plant species. Fifteen phytolith morphotypes were only detected in 14 plant species (termed "distinct" phytoliths). We used these distinct phytoliths to identify plant foods (i.e., that they were associated with) in fecal samples. We then validated findings by checking if the 10 chimpanzees had eaten parts of these plants ∼24 hr prior to fecal sample collection; six plant species associated with five distinct phytoliths had been eaten. Finally, we compared plant foods identified in fecal samples from phytolith analyses with plants that had been identified from macroscopic inspection of the same fecal samples. Findings from phytolith analyses corroborate with those from macroscopic inspection by expanding the total number of plant species identified per fecal sample (i.e., we identified certain plant parts that remained unrecognizable at macroscopic level). This study highlights the potential of phytolith analyses of feces to increase our knowledgebase of the dietary

  5. Complete Genome Sequence of Turicibacter sp. Strain H121, Isolated from the Feces of a Contaminated Germ-Free Mouse

    Science.gov (United States)

    Auchtung, T. A.; Holder, M. E.; Gesell, J. R.; Ajami, N. J.; Duarte, R. T. D.; Itoh, K.; Caspi, R. R.; Petrosino, J. F.; Horai, R.

    2016-01-01

    Turicibacter bacteria are commonly detected in the gastrointestinal tracts and feces of humans and animals, but their phylogeny, ecological role, and pathogenic potential remain unclear. We present here the first complete genome sequence of Turicibacter sp. strain H121, which was isolated from the feces of a mouse line contaminated following germ-free derivation. PMID:27013036

  6. The Effects of GH Transgenic Goats on the Microflora of the Intestine, Feces and Surrounding Soil.

    Directory of Open Access Journals (Sweden)

    Zekun Bao

    Full Text Available The development of genetically engineered animals has brought with it increasing concerns about biosafety issues. We therefore evaluated the risks of growth hormone from transgenic goats, including the probability of horizontal gene transfer and the impact on the microbial community of the goats' gastrointestinal tracts, feces and the surrounding soil. The results showed that neither the GH nor the neoR gene could be detected in the samples. Moreover, there was no significant change in the microbial community of the gastrointestinal tracts, feces and soil, as tested with PCR-denaturing gradient gel electrophoresis and 16S rDNA sequencing. Finally, phylogenetic analysis showed that the intestinal content, feces and soil samples all contained the same dominant group of bacteria. These results demonstrated that expression of goat growth hormone in the mammary of GH transgenic goat does not influence the microflora of the intestine, feces and surrounding soil.

  7. The Effects of GH Transgenic Goats on the Microflora of the Intestine, Feces and Surrounding Soil.

    Science.gov (United States)

    Bao, Zekun; Gao, Xue; Zhang, Qiang; Lin, Jian; Hu, Weiwei; Yu, Huiqing; Chen, Jianquan; Yang, Qian; Yu, Qinghua

    2015-01-01

    The development of genetically engineered animals has brought with it increasing concerns about biosafety issues. We therefore evaluated the risks of growth hormone from transgenic goats, including the probability of horizontal gene transfer and the impact on the microbial community of the goats' gastrointestinal tracts, feces and the surrounding soil. The results showed that neither the GH nor the neoR gene could be detected in the samples. Moreover, there was no significant change in the microbial community of the gastrointestinal tracts, feces and soil, as tested with PCR-denaturing gradient gel electrophoresis and 16S rDNA sequencing. Finally, phylogenetic analysis showed that the intestinal content, feces and soil samples all contained the same dominant group of bacteria. These results demonstrated that expression of goat growth hormone in the mammary of GH transgenic goat does not influence the microflora of the intestine, feces and surrounding soil.

  8. Study on organics biodegradation during aerobic thermophilic composting of human feces%粪便中温好氧堆肥过程有机物的降解研究

    Institute of Scientific and Technical Information of China (English)

    白帆; 王晓昌

    2011-01-01

    Batch experiments were conducted using a closed aerobic composting reactor with sawdust as the bulky matrix to simulate the condition of bio-toilet for sanitary disposal of human feces. Under a controlled condition of temperature at 35 ℃ .moisture content at 60% and continuous air supply,the aerobic composting experiments were conducted for 14 days,and the biodegradation of organics in human feces during the composting process was investigated. Results showed that more than 63% fecal organic was removed under thermophilic condition. Compost maturity period lasted for 10-12 days at 35 ℃. Biomass was analyzed by phospholipid fatty acids (PLFA) ,the process of organics biodegradation could be well described by the first class bio-chemical dynamics reaction. Generally speaking, satisfied compost effect was reached under the methepholic condition while composting maturity period is long,it was important for designing and further spread of bio-toilet.%为了生态厕所的推广使用、便于操作和节约能耗,多采用无加热的设施,即在自然条件下(对于小型生态厕所,接近中温条件)的好氧堆肥处理.了解中温好氧堆肥过程有机物的降解特性,对于生态厕所的推广使用和简化设计、操作等具有重要意义.采用密闭式好氧堆肥反应器,模拟中温(35℃)的堆肥温度,以新鲜锯末为空白载体,在含水率为60%以及连续强制供气条件下,进行了为期14d的实验,评估中温好氧堆肥过程中粪便中有机物的降解特性.结果表明,中温条件下粪便中有机物去除率达到63%以上;且堆肥腐熟期长达10~12 d.微生物总量的磷脂分析表明,中温条件下堆肥的微生物总量的变化充分说明了有机物的生物降解特性.中温好氧堆肥过程中具有较高的有机物去除率,只是堆肥腐熟期较长,这对于生态厕所在实际中的推广应用及设计操作等意义重大.

  9. Dielectric characterisation of human tissue samples

    NARCIS (Netherlands)

    Rossum, W.L. van; Nennie, F.; Deiana, D.; Veen, A.J. van der; Monni, S.

    2014-01-01

    The electrical properties of tissues samples are required for investigation and simulation purposes in biomedical applications of EM sensors. While available open literature mostly deals with ex-vivo characterization of isolated tissues, knowledge on dielectric properties of these tissues in their o

  10. Pattern designation of PCBs in human samples

    Energy Technology Data Exchange (ETDEWEB)

    Wolff, M.S.; Fischbein, A.; Rosenman, K.D.; Levin, S.M.

    1986-03-01

    In order to asses the nature of PCB exposures in humans, statistical measures of PCB patterns in blood serum (as Aroclor 1254 or 1260) were made in 348 cases, representing several exposed and non-exposed groups. Although the cases were not representative of any population, most (252/348) had an Arcolor 1260 pattern, with evidence that PCB congeners in blood serum were usually derived from both Aroclor 1254 and 1260. The method is readily applied to routine packed column gc analysis.

  11. Contribution of additives Cu to its accumulation in pig feces: study in Beijing and Fuxin of China

    Institute of Scientific and Technical Information of China (English)

    LI Yan-xia; LI Wei; WU Juan; XU Li-chao; SU Qiu-hong; XIONG Xiong

    2007-01-01

    Massive amounts of pig manure are produced by intensive pig farm in China, and the composition of pig manure has changed much due to the use of feed additives. However, little is known about the exact Cu (copper) feed as additives or present as contaminants in pig feed and the residues in feces. One hundred and thirty-seven feeds and one hundred and forty-two fecal samples from 48 pig farms were collected in Beijing and Fuxin cities in 1999 and 2005, respectively. The concentrations of Cu were in the range of 6.86-395.19 mg/kg in the feed samples, and the mean values were in the order of weaner> grower-finisher> sow's feeds. The high concentrations over EU recommendations implied that excessive levels of Cu are fed on many pig farms in Beijing and Fuxin. Cu was also present in high concentrations in feces, and concentrations were highly variable. Cu concentrations in the feces from grower-finisher and weaner pigs were significantly greater than feces of sows. The super-intensive and small-scale farms had higher levels of Cu in feces than the middle farms. Cu concentrations in pig feces were approximately 5-times greater than in pig feeds. Feed management in grower-finisher pigs on super-intensive and small-scale pig farms is needed to reduce high Cu concentrations in feces and risks to soil contamination while feces are land-applied.

  12. Isolation of Campylobacter from human stool samples

    Directory of Open Access Journals (Sweden)

    S M Salim

    2014-01-01

    Full Text Available Context: Campylobacter is an undetected cause of diarrhoea especially under 5 years of age in most of the countries. Isolation of this organism is difficult, expensive and cumbersome. Aims: Our objective of this study was to isolate this pathogen from the stool specimens on routinely available blood containing laboratory media using the candle jar for creating the microaerophilic atmosphere in our setup. Settings and Designs: A descriptive study. Materials and Methods: A total of 50 stool samples were inoculated onto selective and non-selective media with and without filtration using a 0.45 μm membrane. The inoculated media were simultaneously incubated in microaerophilic conditions using the Anoxomat as well as in candle jars at temperatures 37°C and 42°C. The culture isolates were confirmed by standard phenotypic tests. A simplex polymerase chain reaction (PCR targeting the 16S ribosomal deoxyribonucleic acid of Campylobacter was performed on the deoxyribonucleic acid (DNA of the culture isolates as well as on the DNA extracted from the stool filtrates. Statistical Analysis: Data was expressed as a proportion. Results: Campylobacter could be isolated in 5 out of 50 stool samples using both the Anoxomat as well as the candle jar. Furthermore, we did not find any difference between the isolation using the selective and blood containing media as well as the different incubation temperatures. All the five were confirmed phenotypically and genotypically to be Campylobacter jejuni. The PCR results corroborated with that of the culture. Conclusions: Isolation by culture was as sensitive as that of the PCR.

  13. Rabbit Feces as Feed for Ruminants and as an Energy Source

    Directory of Open Access Journals (Sweden)

    Pier Giorgio Peiretti

    2014-12-01

    Full Text Available There are prospects for using novel feeds from various sources to provide ruminants with alternative sources of protein and energy such as by-products, and animal wastes. Rabbit feces are a concentrated source of fiber and could have commercial potential both as input biomass in anaerobic processes for biogas production, as well as a fibrous source for ruminal degradation. The aims of this work were to assess the potential as ruminant feeding and as biogas production of rabbit feces, in comparison with 12 crops. The chemical composition and the potential and experimental in vitro true digestibility (IVTD and neutral detergent fiber digestibility (NDFD of 148 feces samples were determined by using chemical methods, Daisy system digestibility and/or NIRS predictions. The average biomethane potential (BMP was 286 ± 10 lCH4/kg SV with −4% vs. the crops average. Milk forage unit (milk FU, IVTD and NDFD of feces were 0.54 ± 0.06 milk FU/kg DM, 74% ± 3% and 50% ± 5%, respectively, with comparisons of −19%, −11% and −24% vs. the crops average. Reconstruction of the potential values based on the chemical constituents but using the crop partial least square model well agreed with the NIRS calibrations and cross-validation. In a global NIRS calibration of the feces and crops the relative predicted deviation for IVTD, NDFD and milk FU were 3.1, 2.9 and 2.6, respectively, and only 1.5 for BMP. Running the Daisy system for rabbit feces in rumen fluid gave some inconsistencies, weakened the functional relationships, and appeared not to be correlated with the potential values of IVTD and NDFD. Nevertheless, the energetic potential of feces appears to be similar to some conventional crops at different degrees of maturity. Thus we conclude that rabbit feces has potential value as a ruminant feed and for biogas production.

  14. Attractiveness of native mammal's feces of different trophic guilds to dung beetles (Coleoptera: Scarabaeinae).

    Science.gov (United States)

    Bogoni, Juliano A; Hernández, Malva I M

    2014-01-01

    Mammal feces are the primary food and nesting resource for the majority of dung beetle species, and larval development depends on the quantity and quality of that resource. Physiological necessities, competitive interactions, and resource sharing are common and suggest that dung beetles may show preferences for feces of greater nutritional quality, which may in turn impact beetle assemblages and community structure. This study investigated whether attractiveness of dung beetles to different resource (feces) types varies depending on mammal trophic guild and associated nutritional content. This study was conducted in Atlantic Forest fragments in the Parque Estadual da Serra do Tabuleiro, Santa Catarina, Brazil. To evaluate attractiveness, the feces of the carnivore Puma concolor, the omnivores Cerdocyon thous and Sapajus nigritus, and the herbivore Tapirus terrestris were utilized as bait. Dung was collected from zoo animals fed a standard diet. Sampling was performed in triplicate in five areas in the summer of 2013. Four pitfall traps were established in each area, and each trap was baited with one type of mammal feces. Food preference of the species was analyzed by calculating Rodgers' index for cafeteria-type experiments. In total, 426 individuals from 17 species were collected. Rodgers' index showed that omnivorous mammal feces (C. thous) were most attractive to all dung beetle species, although it is known that dung beetles are commonly opportunistic with respect to search for and allocation of food resources. These results suggest that mammal loss could alter competitive interactions between dung beetles.

  15. Escherichia coli O104 in Feedlot Cattle Feces: Prevalence, Isolation and Characterization.

    Directory of Open Access Journals (Sweden)

    Pragathi B Shridhar

    Full Text Available Escherichia coli O104:H4, an hybrid pathotype of Shiga toxigenic and enteroaggregative E. coli, involved in a major foodborne outbreak in Germany in 2011, has not been detected in cattle feces. Serogroup O104 with H type other than H4 has been reported to cause human illnesses, but their prevalence and characteristics in cattle have not been reported. Our objectives were to determine the prevalence of E. coli O104 in feces of feedlot cattle, by culture and PCR detection methods, and characterize the isolated strains. Rectal fecal samples from a total of 757 cattle originating from 29 feedlots were collected at a Midwest commercial slaughter plant. Fecal samples, enriched in E. coli broth, were subjected to culture and PCR methods of detection. The culture method involved immunomagnetic separation with O104-specific beads and plating on a selective chromogenic medium, followed by serogroup confirmation of pooled colonies by PCR. If pooled colonies were positive for the wzxO104 gene, then colonies were tested individually to identify wzxO104-positive serogroup and associated genes of the hybrid strains. Extracted DNA from feces were also tested by a multiplex PCR to detect wzxO104-positive serogroup and associated major genes of the O104 hybrid pathotype. Because wzxO104 has been shown to be present in E. coli O8/O9/O9a, wzxO104-positive isolates and extracted DNA from fecal samples were also tested by a PCR targeting wbdDO8/O9/O9a, a gene specific for E. coli O8/O9/O9a serogroups. Model-adjusted prevalence estimates of E. coli O104 (positive for wzxO104 and negative for wbdDO8/O9/O9a at the feedlot level were 5.7% and 21.2%, and at the sample level were 0.5% and 25.9% by culture and PCR, respectively. The McNemar's test indicated that there was a significant difference (P < 0.01 between the proportions of samples that tested positive for wzxO104 and samples that were positive for wzxO104, but negative for wbdDO8/O9/O9a by PCR and culture methods

  16. Molecular evidence for the presence of Rickettsia Felis in the feces of wild-living African apes.

    Directory of Open Access Journals (Sweden)

    Alpha Kabinet Keita

    Full Text Available BACKGROUND: Rickettsia felis is a common emerging pathogen detected in mosquitoes in sub-Saharan Africa. We hypothesized that, as with malaria, great apes may be exposed to the infectious bite of infected mosquitoes and release R. felis DNA in their feces. METHODS: We conducted a study of 17 forest sites in Central Africa, testing 1,028 fecal samples from 313 chimpanzees, 430 gorillas and 285 bonobos. The presence of rickettsial DNA was investigated by specific quantitative real-time PCR. Positive results were confirmed by a second PCR using primers and a probe targeting a specific gene for R. felis. All positive samples were sequenced. RESULTS: Overall, 113 samples (11% were positive for the Rickettsia-specific gltA gene, including 25 (22% that were positive for R. felis. The citrate synthase (gltA sequence and outer membrane protein A (ompA sequence analysis indicated 99% identity at the nucleotide level to R. felis. The 88 other samples (78% were negative using R. felis-specific qPCR and were compatible with R. felis-like organisms. CONCLUSION: For the first time, we detected R. felis in wild-living ape feces. This non invasive detection of human pathogens in endangered species opens up new possibilities in the molecular epidemiology and evolutionary analysis of infectious diseases, beside HIV and malaria.

  17. Shigatoxigenic Escherichia coli serogroups O157, O111 and O113 in feces, water and milk samples from dairy farms Escherichia coli Shigatoxigênicas sorogrupos O157, O111 e O113 detectadas em fezes de bovinos, água e leite de propriedades leiteiras

    Directory of Open Access Journals (Sweden)

    Hinig Isa Godoy Vicente

    2005-09-01

    Full Text Available The objective of this study was to determine the prevalence of Shigatoxigenic Escherichia coli (STEC and STEC serogroups O157, O111 and O113 in feces, water and milk sampled in dairy farms in Jaboticabal, SP, Brazil. Feces (n=454, water (n=54 and milk samples (n=30 were collected from 10 herds and assessed for the presence of the virulence genes stx1, stx2 and eae by polymerase chain reaction (PCR. All stx and eae positive samples were submitted to a second PCR reaction targeting the sequences rfb O157, rfb O111 and rfb O113. High prevalence of stx was detected (59.9% in fecal samples, whereas the prevalence of sequences rfb O157, rfb O111 and rfb O113 was 18.9%, 3.3% and 30.4%, respectively. All sequences were detected more frequently in calves and heifers. Sequences stx2 and eae were prevalent in the fecal samples. stx sequences were detected in 1.9% and 3.3% of water and milk samples, respectively. Low prevalence of E. coli O113 was observed in water samples, whereas no E. coli O157 or E. coli O111 was detected. Furthermore, none of the serogroups were identified in milk samples. STEC was identified in all herds (100%, and serogroups O157, O111 and O113 were observed in 40%, 50% and 90% of the herds, respectively. In conclusion, the high STEC prevalence detected in dairy herds evidences that bovine feces might play an important role as a contamination source in the region of Jaboticabal, Brazil.Este trabalho teve como objetivo determinar a prevalência de Escherichia coli Shigatoxigênicas dos sorogrupos O157, O111 e O113 em bovinos leiteiros, água e leite de propriedades rurais do Município de Jaboticabal/SP. Para isso, foram colhidas 454 amostras de fezes, 54 amostras de água e 30 amostras de leite e pesquisada a presença de seqüências stx1, stx2 e eae pela reação em cadeia da polimerase (PCR. Todas as amostras stx e/ eae positivas foram submetidas a uma nova reação de PCR para detecção das seqüências rfb O157, rfb O111 e rfb

  18. Isolation and maintenance of Balantidium coli (Malmsteim, 1857) cultured from fecal samples of pigs and non-human primates.

    Science.gov (United States)

    Barbosa, Alynne da Silva; Bastos, Otilio Machado Pereira; Uchôa, Claudia M Antunes; Pissinatti, Alcides; Ferreira Filho, Paulo Ricardo; Dib, Lais Verdan; Azevedo, Eduarda Peixoto; de Siqueira, Mayara Perlingeiro; Cardozo, Matheus Lessa; Amendoeira, Maria Regina Reis

    2015-06-15

    Balantidium coli is a protozoa that can determine dysentery in humans, pigs and non-human primates having zoonotic potential. The lack of standardization in isolation and maintenance hinders the development of research on its biology and epidemiology. This study is aimed to standardize the isolation and maintenance of this parasite from animal feces, in culture medium, Pavlova modified. From 2012 to 2014, 1905 fecal samples were collected from captive animals of Rio de Janeiro. Were selected for isolation samples with a minimum of 10 trophozoites and/or 30 cysts of B. coli, totaling 88 pigs, 26 Cynomolgus and 90 rhesus macaques. In the presence of cysts, the sample was homogenized in saline solution, 500 μL was removed and inoculated into culture medium. The material that contained trophozoites the inoculum was made from 240 μL of fecal solution. All inoculate tubes with the subcultures were kept at 36°C, and sterile rice starch was always added to the medium. The parasites isolate from pigs, 34%, and from Cynomolgus 38.4% were maintained in vitro for a period of more than 24 months. These procedures proved to be adequate for isolation and maintenance of B. coli from different animals, they were found to be inexpensive and easy to perform.

  19. Specific IgA to lactic acid bacteria in feces of children consuming milk fermented by yoghurt symbiosis and Lactobacillus casei (Danone strain DN 114 001).

    Science.gov (United States)

    Faure, G C; Morisset, M; Gobert, B; Guérin, C; Pedone, C; Bouley, C; Béné, M C

    2001-01-01

    An immunoreactive role of lactic acid bacteria established in animals has seldom been investigated in humans. In a large-scale clinical study, children from day-care centers received either yoghurt (Y), milk fermented by yoghurt symbiosis and Lactobacillus casei (DN 114 001) (YC), or gelified milk (GM) as diet supplements during two 30-day supplementation periods separated by one 30-day period without supplementation. Feces samples were collected before, during, and after the 2nd supplementation period. Proteins were extracted in a buffer containing enzymatic inhibitors. IgA levels were assessed and adjusted to the weight of feces samples. Specific IgA to lactic acid bacteria strains (Streptococcus thermophilus 8901A, 8902A; Lactobacillus bulgaricus; Lactobacillus casei) present in Y and YC were assayed in ELISA and adjusted to individual IgA levels. Mean levels of fecal IgA were within reported ranges for pediatric populations of similar age. IgA levels decreased significantly but transiently in children receiving Y, and increased significantly in children receiving GM, but did not vary in the group of children who were given YC. Specific IgA to the 4 strains tested increased significantly during the supplementation period only in the group of children receiving GM, while it was transient and not significant in children receiving YC. No variation was noted in children given Y Specific IgA to lactic acid bacteria can be assayed in feces. Supplementation with fermented milks might induce a mucosal tolerance to environmental flora.

  20. Manure production and mineral excretion in feces of gilts fed ractopamine - doi: 10.4025/actascianimsci.v35i3.18662

    Directory of Open Access Journals (Sweden)

    Everton Daniel

    2013-07-01

    Full Text Available A study was conducted to evaluate the feces+urine produced per animal (FUPA, dry matter, mineral matter, organic matter, nitrogen, phosphorus, potassium and sulfur in feces of gilts fed diets with increasing levels of ractopamine (0, 5, 10 and 15 mg kg-1 of diet. A total of 468 finishing gilts were allotted into 36 pens. In two days of each week, feces and urine were daily sampled in four pens per treatment, quantifying the feces+urine. To determine the characterization of feces, two samples per week were taken daily, in nine pens per treatment. It was used a split plot design, considering the ractopamine level as the plot and the weeks as the subplots. There was no reduction in nitrogen amount in feces. An interaction was detected between ractopamine concentrations and weeks for FUPA and phosphorus, potassium and sulfur in feces. Ractopamine addition in diets for gilts has reduced the feces+urine production and nitrogen and phosphorus excretion. Higher values estimated for potassium content in feces of animals fed diets with 10 and 15 mg of ractopamine kg-1 were found between the second and third week. Increasing levels of ractopamine from 5 to 15 mg kg-1 promoted higher excretion of sulfur over the weeks of supply.   

  1. Approach-Induced Biases in Human Information Sampling

    Science.gov (United States)

    Hunt, Laurence T.; Rutledge, Robb B.; Malalasekera, W. M. Nishantha; Kennerley, Steven W.; Dolan, Raymond J.

    2016-01-01

    Information sampling is often biased towards seeking evidence that confirms one’s prior beliefs. Despite such biases being a pervasive feature of human behavior, their underlying causes remain unclear. Many accounts of these biases appeal to limitations of human hypothesis testing and cognition, de facto evoking notions of bounded rationality, but neglect more basic aspects of behavioral control. Here, we investigated a potential role for Pavlovian approach in biasing which information humans will choose to sample. We collected a large novel dataset from 32,445 human subjects, making over 3 million decisions, who played a gambling task designed to measure the latent causes and extent of information-sampling biases. We identified three novel approach-related biases, formalized by comparing subject behavior to a dynamic programming model of optimal information gathering. These biases reflected the amount of information sampled (“positive evidence approach”), the selection of which information to sample (“sampling the favorite”), and the interaction between information sampling and subsequent choices (“rejecting unsampled options”). The prevalence of all three biases was related to a Pavlovian approach-avoid parameter quantified within an entirely independent economic decision task. Our large dataset also revealed that individual differences in the amount of information gathered are a stable trait across multiple gameplays and can be related to demographic measures, including age and educational attainment. As well as revealing limitations in cognitive processing, our findings suggest information sampling biases reflect the expression of primitive, yet potentially ecologically adaptive, behavioral repertoires. One such behavior is sampling from options that will eventually be chosen, even when other sources of information are more pertinent for guiding future action. PMID:27832071

  2. Survival of Campylobacter jejuni in naturally and artificially contaminated laying hen feces.

    Science.gov (United States)

    Ahmed, M F M; Schulz, J; Hartung, J

    2013-02-01

    Infected laying hens regularly excrete large amounts of Campylobacter jejuni with their feces, which represent a reservoir of infection within the flock and for animals in the region. However, the knowledge about survival times of C. jejuni in these feces is still scarce. Therefore, orienting laboratory experiments were carried out under controlled conditions to estimate the survival times of C. jejuni both in artificially and naturally contaminated laying hen feces. In 6 different laying hen flocks (3 Campylobacter-free and 3 Campylobacter-positive flocks), fresh excreta were randomly collected and pooled in 20-g samples per flock. In the laboratory, each of the 3 pooled samples from the Campylobacter-free barns were homogenized and mixed with 10 mL of a freshly prepared C. jejuni suspension (3 × 10(8) cfu/mL). The other 3 samples were homogenized only. The 6 samples were stored at 20 ± 1°C and 40 to 60% RH in 2 different incubators. Specimens of 2 g were taken from all 6 samples 1 h after storage and daily at the same time during the next 10 consecutive days and investigated on culturable C. jejuni. The survival times of culturable C. jejuni ranged from 72 to 96 h in artificially inoculated feces and varied from 120 to 144 h in naturally colonized flocks. The flaA typing by RFLP confirmed that the isolates from the artificially contaminated feces were identical with the added strain. A total of 5 different flaA types were identified from the naturally contaminated feces, and survival of these isolates was dependent on flaA type. The demonstrated survival times indicate that contaminated fresh feces are an important reservoir of C. jejuni, representing a permanent source of infection over at least 6 d after excretion. It shows the considerable potential of fresh feces in transmitting the agent within and between flocks during that period. This 6-d span should be considered when poultry manure is applied to land as organic fertilizer.

  3. Droplet digital PCR quantifies host inflammatory transcripts in feces reliably and reproducibly

    Science.gov (United States)

    The gut is the most extensive, interactive, and complex interface between the human host and the environment and therefore a critical site of immunological activity. Non-invasive methods to assess the host response in this organ are currently lacking. Feces are the available analyte which have been ...

  4. Phylogenetic diversity of cultivalble butyrate-producing bacteria from pig gut content and feces

    DEFF Research Database (Denmark)

    Li, Xiaoqiong; Højberg, Ole; Canibe, Nuria

    2016-01-01

    Butyrate is a preferred energy source for colonocytes and is considered crucial for maintaining colonic health in humans and animals. To investigate the diversity of cultivable butyrate-producing bacteria in pig gut, bacteria were isolated from intestinal digesta (Exp. 1) and feces (Exp. 2...

  5. Human-Robot Site Survey and Sampling for Space Exploration

    Science.gov (United States)

    Fong, Terrence; Bualat, Maria; Edwards, Laurence; Flueckiger, Lorenzo; Kunz, Clayton; Lee, Susan Y.; Park, Eric; To, Vinh; Utz, Hans; Ackner, Nir

    2006-01-01

    NASA is planning to send humans and robots back to the Moon before 2020. In order for extended missions to be productive, high quality maps of lunar terrain and resources are required. Although orbital images can provide much information, many features (local topography, resources, etc) will have to be characterized directly on the surface. To address this need, we are developing a system to perform site survey and sampling. The system includes multiple robots and humans operating in a variety of team configurations, coordinated via peer-to-peer human-robot interaction. In this paper, we present our system design and describe planned field tests.

  6. In silico analyses of metagenomes from human atherosclerotic plaque samples

    DEFF Research Database (Denmark)

    Mitra, Suparna; Drautz-Moses, Daniela I; Alhede, Morten

    2015-01-01

    a challenge. RESULTS: To investigate microbiome diversity within human atherosclerotic tissue samples, we employed high-throughput metagenomic analysis on: (1) atherosclerotic plaques obtained from a group of patients who underwent endarterectomy due to recent transient cerebral ischemia or stroke. (2...

  7. A DNA methylation fingerprint of 1628 human samples

    Science.gov (United States)

    Fernandez, Agustin F.; Assenov, Yassen; Martin-Subero, Jose Ignacio; Balint, Balazs; Siebert, Reiner; Taniguchi, Hiroaki; Yamamoto, Hiroyuki; Hidalgo, Manuel; Tan, Aik-Choon; Galm, Oliver; Ferrer, Isidre; Sanchez-Cespedes, Montse; Villanueva, Alberto; Carmona, Javier; Sanchez-Mut, Jose V.; Berdasco, Maria; Moreno, Victor; Capella, Gabriel; Monk, David; Ballestar, Esteban; Ropero, Santiago; Martinez, Ramon; Sanchez-Carbayo, Marta; Prosper, Felipe; Agirre, Xabier; Fraga, Mario F.; Graña, Osvaldo; Perez-Jurado, Luis; Mora, Jaume; Puig, Susana; Prat, Jaime; Badimon, Lina; Puca, Annibale A.; Meltzer, Stephen J.; Lengauer, Thomas; Bridgewater, John; Bock, Christoph; Esteller, Manel

    2012-01-01

    Most of the studies characterizing DNA methylation patterns have been restricted to particular genomic loci in a limited number of human samples and pathological conditions. Herein, we present a compromise between an extremely comprehensive study of a human sample population with an intermediate level of resolution of CpGs at the genomic level. We obtained a DNA methylation fingerprint of 1628 human samples in which we interrogated 1505 CpG sites. The DNA methylation patterns revealed show this epigenetic mark to be critical in tissue-type definition and stemness, particularly around transcription start sites that are not within a CpG island. For disease, the generated DNA methylation fingerprints show that, during tumorigenesis, human cancer cells underwent a progressive gain of promoter CpG-island hypermethylation and a loss of CpG methylation in non-CpG-island promoters. Although transformed cells are those in which DNA methylation disruption is more obvious, we observed that other common human diseases, such as neurological and autoimmune disorders, had their own distinct DNA methylation profiles. Most importantly, we provide proof of principle that the DNA methylation fingerprints obtained might be useful for translational purposes by showing that we are able to identify the tumor type origin of cancers of unknown primary origin (CUPs). Thus, the DNA methylation patterns identified across the largest spectrum of samples, tissues, and diseases reported to date constitute a baseline for developing higher-resolution DNA methylation maps and provide important clues concerning the contribution of CpG methylation to tissue identity and its changes in the most prevalent human diseases. PMID:21613409

  8. Isolation and identification of tyramine-producing enterococci from human fecal samples.

    Science.gov (United States)

    Ladero, Victor; Fernández, María; Alvarez, Miguel A

    2009-02-01

    Lactic acid bacteria (LAB) are recognized as a group of important microorganisms because of their crucial role in food fermentation and their contribution to the maintenance of health homeostasis, as natural inhabitants of the human mucosa. However, the metabolic activities of some strains, such as the ability to synthesize biogenic amines (BAs), can be detrimental to human health. BAs are low molecular weight compounds synthesized by the enzymatic decarboxylation of amino acids. Tyramine, one of the most biologically active BAs, is produced by certain strains of LAB related to food fermentations. Since no data are available as to whether tyramine originates exclusively from food intake, or, like polyamines, could be formed by gut bacteria, this study was focused on the isolation of tyramine-producing LAB from human feces. Different strains of Enterococcus faecium and Enterococcus faecalis able to produce tyramine in culture conditions were isolated.

  9. Acetone and isopropanol in ruminal fluid and feces of lactating dairy cows.

    Science.gov (United States)

    Sato, Hiroshi; Shiogama, Yumiko

    2010-03-01

    Acetone and its metabolite isopropanol are produced by gut microbes as well as by the host's metabolism. To evaluate the production of acetone and isopropanol in alimentary tracts, a total of 80 pair-samples of feces and ruminal fluid were taken in lactating dairy cows that had been fed silage-containing diets. Acetone and isopropanol were analyzed, together with ethanol and volatile fatty acids (VFAs). Isopropanol was detected in 57 fecal and all the ruminal samples; however, the ruminal isopropanol and ethanol concentrations were distinctly lower than those in the feces. Acetone was detected in 13 fecal and 53 ruminal samples; however, there was no significant difference in acetone concentrations between the feces and the ruminal fluid. The group with higher fecal isopropanol concentration showed higher fecal proportions of acetate accompanied by low proportion of minor VFA, which consisted of isobutyrate and iso- and n-valerate. In the group with higher ruminal isopropanol concentration, ethanol concentration was higher; the ruminal VFA profiles showed only a negligible difference. Fecal and ruminal ethanol concentrations were not affected by feed ethanol. Thus, the colon showed an accelerated alcoholic fermentation compared with the rumen of dairy cows; however, acetone was present at higher frequency in the rumen than in the feces.

  10. Child feces disposal practices in rural Orissa: a cross sectional study.

    Directory of Open Access Journals (Sweden)

    Fiona Majorin

    Full Text Available BACKGROUND: An estimated 2.5 billion people worldwide lack access to improved sanitation facilities. While large-scale programs in some countries have increased latrine coverage, they sometimes fail to ensure optimal latrine use, including the safe disposal of child feces, a significant source of exposure to fecal pathogens. We undertook a cross-sectional study to explore fecal disposal practices among children in rural Orissa, India in villages where the Government of India's Total Sanitation Campaign had been implemented at least three years prior to the study. METHODS AND FINDINGS: We conducted surveys with heads of 136 households with 145 children under 5 years of age in 20 villages. We describe defecation and feces disposal practices and explore associations between safe disposal and risk factors. Respondents reported that children commonly defecated on the ground, either inside the household (57.5% for pre-ambulatory children or around the compound (55.2% for ambulatory children. Twenty percent of pre-ambulatory children used potties and nappies; the same percentage of ambulatory children defecated in a latrine. While 78.6% of study children came from 106 households with a latrine, less than a quarter (22.8% reported using them for disposal of child feces. Most child feces were deposited with other household waste, both for pre-ambulatory (67.5% and ambulatory (58.1% children. After restricting the analysis to households owning a latrine, the use of a nappy or potty was associated with safe disposal of feces (OR 6.72, 95%CI 1.02-44.38 though due to small sample size the regression could not adjust for confounders. CONCLUSIONS: In the area surveyed, the Total Sanitation Campaign has not led to high levels of safe disposal of child feces. Further research is needed to identify the actual scope of this potential gap in programming, the health risk presented and interventions to minimize any adverse effect.

  11. Groundbreaking Mars Sample Return for Science and Human Exploration

    Science.gov (United States)

    Cohen, Barbara; Draper, David; Eppler, Dean; Treiman, Allan

    2012-01-01

    Partnerships between science and human exploration have recent heritage for the Moon (Lunar Precursor Robotics Program, LPRP) and nearearth objects (Exploration Precursor Robotics Program, xPRP). Both programs spent appreciable time and effort determining measurements needed or desired before human missions to these destinations. These measurements may be crucial to human health or spacecraft design, or may be desired to better optimize systems designs such as spacesuits or operations. Both LPRP and xPRP recommended measurements from orbit, by landed missions and by sample return. LPRP conducted the Lunar Reconnaissance Orbiter (LRO) and Lunar Crater Observation and Sensing Satellite (LCROSS) missions, providing high-resolution visible imagery, surface and subsurface temperatures, global topography, mapping of possible water ice deposits, and the biological effects of radiation [1]. LPRP also initiated a landed mission to provide dust and regolith properties, local lighting conditions, assessment of resources, and demonstration of precision landing [2]. This mission was canceled in 2006 due to funding shortfalls. For the Moon, adequate samples of rocks and regolith were returned by the Apollo and Luna programs to conduct needed investigations. Many near-earth asteroids (NEAs) have been observed from the Earth and several have been more extensively characterized by close-flying missions and landings (NEAR, Hayabusa, Rosetta). The current Joint Robotic Precursor Activity program is considering activities such as partnering with the New Frontiers mission OSIRIS-Rex to visit a NEA and return a sample to the Earth. However, a strong consensus of the NEO User Team within xPRP was that a dedicated mission to the asteroid targeted by humans is required [3], ideally including regolith sample return for more extensive characterization and testing on the Earth.

  12. Sampling strategy for estimating human exposure pathways to consumer chemicals

    Directory of Open Access Journals (Sweden)

    Eleni Papadopoulou

    2016-03-01

    Full Text Available Human exposure to consumer chemicals has become a worldwide concern. In this work, a comprehensive sampling strategy is presented, to our knowledge being the first to study all relevant exposure pathways in a single cohort using multiple methods for assessment of exposure from each exposure pathway. The selected groups of chemicals to be studied are consumer chemicals whose production and use are currently in a state of transition and are; per- and polyfluorinated alkyl substances (PFASs, traditional and “emerging” brominated flame retardants (BFRs and EBFRs, organophosphate esters (OPEs and phthalate esters (PEs. Information about human exposure to these contaminants is needed due to existing data gaps on human exposure intakes from multiple exposure pathways and relationships between internal and external exposure. Indoor environment, food and biological samples were collected from 61 participants and their households in the Oslo area (Norway on two consecutive days, during winter 2013-14. Air, dust, hand wipes, and duplicate diet (food and drink samples were collected as indicators of external exposure, and blood, urine, blood spots, hair, nails and saliva as indicators of internal exposure. A food diary, food frequency questionnaire (FFQ and indoor environment questionnaire were also implemented. Approximately 2000 samples were collected in total and participant views on their experiences of this campaign were collected via questionnaire. While 91% of our participants were positive about future participation in a similar project, some tasks were viewed as problematic. Completing the food diary and collection of duplicate food/drink portions were the tasks most frequent reported as “hard”/”very hard”. Nevertheless, a strong positive correlation between the reported total mass of food/drinks in the food record and the total weight of the food/drinks in the collection bottles was observed, being an indication of accurate performance

  13. Microstructure and oxygen evolution of Fe-Ce mixed oxides by redox treatment

    Science.gov (United States)

    Li, Kongzhai; Haneda, Masaaki; Ning, Peihong; Wang, Hua; Ozawa, Masakuni

    2014-01-01

    The relationship between structure and reduction/redox properties of Fe-Ce mixed oxides with a Fe content of 5, 10, 20 or 30 mol%, prepared by a coprecipitation method, were investigated by XRD, Raman, TEM, TPR and TPO techniques. It is found that all the iron ions can be incorporated into the ceria lattice to form a solid solution for the FeCe 5 (Fe 5%) sample, but amorphous or crystal Fe2O3 particles were found to be present on the Fe-Ce oxide samples with higher the iron content. The reducibility of single solid solution was much better than the pure CeO2, and the appearance of dispersed Fe2O3 particles improved the surface reducibility of materials. The iron ions incorporated into the CeO2 lattice accelerated the oxygen release from bulk to surface, and surface Fe2O3 particles in close contact to CeO2 acted as a catalyst for the reaction between solid solution and hydrogen. The microstructure of exposed Fe2O3 with Ce-Fe-O solid solution allows the Fe-Ce mixed oxides to own good reducibility and high OSC, which also counteracts the deactivation of the reducibility resulting from the sintering of materials in the redox cycling.

  14. Identifying and analyzing bacteriophages in human fecal samples: what could we discover?

    Science.gov (United States)

    Muniesa, Maite; Jofre, Juan

    2014-01-01

    The human gut is a complex ecosystem, densely populated with microbes including enormous amounts of phages. Metagenomic studies indicate a great diversity of bacteriophages, and because of the variety of gut bacterial species, the human or animal gut is probably a perfect ecological niche for phages that can infect and propagate in their bacterial communities. In addition, some phages have the capacity to mobilize genes, as demonstrated by the enormous fraction of phage particles in feces that contain bacterial DNA. All these facts indicate that, through predation and horizontal gene transfer, bacteriophages play a key role in shaping the size, structure and function of intestinal microbiomes, although our understanding of their effects on gut bacterial populations is only just beginning.

  15. [Fusarium graminearum presence in wheat samples for human consumption].

    Science.gov (United States)

    Martinez, Mauro; Castañares, Eliana; Dinolfo, María I; Pacheco, Walter G; Moreno, María V; Stenglein, Sebastián A

    2014-01-01

    One of the most important diseases in cereal crops is Fusarium head blight, being Fusarium graminearum the main etiological agent. This fungus has the ability to produce a wide spectrum and quantity of toxins, especially deoxynivalenol (DON). During the last crop season (2012-2013) the climatic conditions favored Fusarium colonization. The objective of this work was to determine the presence of this fungus as well as the DON content in 50 wheat grain samples. Our results showed that 80% of the samples were contaminated with Fusarium graminearum. Twenty four percent (24%) of the samples contained ≥ 1μg/g DON, 26% ranged from 0,5 and 0,99μg/g, and the remaining 50% had values lower than 0,5μg/g. Correlation was found between the presence of Fusarium graminearum and DON. It is necessary to establish DON limit values in wheat grains for human consumption.

  16. Identification of dominant bacteria in feces and colonic mucosa from healthy Spanish adults by culturing and by 16S rDNA sequence analysis.

    Science.gov (United States)

    Delgado, Susana; Suárez, Adolfo; Mayo, Baltasar

    2006-04-01

    The aim of this work was to examine by culturing the changes in the total and indicator populations of the feces of two individuals over 1 year and to identify the dominant microbial components of a single sample of feces from each donor. Populations and dominant bacteria from a sample of colonic mucosa from a further individual were also assessed. The culture results were then compared to those obtained with the same samples by 16S rDNA cloning and sequencing. High interindividual variation in representative microbial populations of the gastrointestinal tract (GIT) was revealed by both the culture and the culture-independent techniques. Species belonging to Clostridium clusters (XIVa, IV, and XVIII) predominated in both the fecal and the mucosal samples (except in the mucose cultured isolates), members of Clostridium coccoides cluster XIVa being the most numerous microorganisms. Species of gamma-proteobacteria (Escherichia coli and Shigella spp.), bifidobacteria, and actinobacteria appeared in lower numbers than those of clostridia. From the mucosal cultured sample, only facultative anaerobes and bifidobacteria were recovered, suggesting destruction of the anaerobe population during processing. In accordance with this, the microbial diversity revealed by 16S rDNA sequence analysis was greater than that revealed by culturing. Despite large interindividual differences, distinct human communities may have group-associated GIT microbiota characteristics, such as the low number of Bacteroides seen in the subjects in this study.

  17. Detection and Characterization of Shiga Toxin Producing Escherichia coli, Salmonella spp., and Yersinia Strains from Human, Animal, and Food Samples in San Luis, Argentina.

    Science.gov (United States)

    Favier, Gabriela Isabel; Lucero Estrada, Cecilia; Cortiñas, Teresa Inés; Escudero, María Esther

    2014-01-01

    Shiga toxin producing Escherichia coli (STEC), Salmonella spp., and Yersinia species was investigated in humans, animals, and foods in San Luis, Argentina. A total of 453 samples were analyzed by culture and PCR. The antimicrobial susceptibility of all the strains was studied, the genomic relationships among isolates of the same species were determined by PFGE, and the potencial virulence of Y. enterocolitica strains was analyzed. Yersinia species showed higher prevalence (9/453, 2.0%, 95% CI, 0.7-3.3%) than STEC (4/453, 0.9%, 95% CI, 0-1.8%) and Salmonella spp. (3/453, 0.7%, 95% CI, 0-1.5%). Y. enterocolitica and Y. intermedia were isolated from chicken carcasses (6/80, 7.5%, 95% CI, 1.5-13.5%) and porcine skin and bones (3/10, 30%, 95% CI, 0-65%). One STEC strain was recovered from human feces (1/70, 1.4%, 95% CI, 0-4.2%) and STEC stx1/stx2 genes were detected in bovine stools (3/129, 2.3%, 95% CI, 0-5.0%). S. Typhimurium was isolated from human feces (1/70, 1.4%, 95% CI, 0-4.2%) while one S. Newport and two S. Gaminara strains were recovered from one wild boar (1/3, 33%, 95% CI, 0-99%). The knowledge of prevalence and characteristics of these enteropathogens in our region would allow public health services to take adequate preventive measures.

  18. Virulence factors in Escherichia coli strains isolated from urinary tract infection and pyometra cases and from feces of healthy dogs.

    Science.gov (United States)

    Siqueira, Amanda K; Ribeiro, Marcio G; Leite, Domingos da S; Tiba, Monique R; Moura, Claudia de; Lopes, Maria Denise; Prestes, Nereu Carlos; Salerno, Tatiana; Silva, Aristeu V da

    2009-04-01

    The aim of this study was to compare the prevalence of virulence genes in 158 Escherichia coli strains isolated from 51 clinical cases of UTIs, 52 of pyometra and from 55 fecal samples from healthy dogs by PCR. papC was found in 12 (23.5%) strains isolated from UTIs, 19 (36.5%) from pyometra and 10 (18.2%) from feces. papGII was observed in 3 (5.8%) strains from pyometra, and papGIII in 10 (19.6%) from UTIs, 15 (28.8%) from pyometra and 9 (16.4%) from feces. sfaS was detected in 22 (43.1%) strains from UTIs, 24 (46.1%) from pyometra and 19 (34.5%) from feces. hlyA was observed in 17 (33.3%) strains from UTIs, 18 (34.6%) from pyometra and 7 (12.7%) from feces, while cnf-1 was detected in 11 (21.6%) from UTIs, 21 (40.4%) from pyometra and 9 (16.4%) from feces. iucD was observed in 12 (23.5%) strains from UTIs, 9 (17.3%) from pyometra and 1 (1.8%) from feces. usp was found 17 (33.3%) isolates from UTIs and 36 (69.9%) from pyometra.

  19. Changes in Microbiota in Rumen Digesta and Feces Due to a Grain-Based Subacute Ruminal Acidosis (SARA) Challenge.

    Science.gov (United States)

    Plaizier, Jan C; Li, Shucong; Danscher, Anne Mette; Derakshani, Hooman; Andersen, Pia H; Khafipour, Ehsan

    2017-02-08

    The effects of a grain-based subacute ruminal acidosis (SARA) challenge on bacteria in the rumen and feces of lactating dairy cows were determined. Six lactating, rumen-cannulated Danish Holstein cows were used in a cross-over study with two periods. Periods included two cows on a control diet and two cows on a SARA challenge. The control diet was a total mixed ration containing 45.5% dry matter (DM), 43.8% DM neutral detergent fiber, and 19.6% DM starch. The SARA challenge was conducted by gradually substituting the control diet with pellets containing 50% wheat and 50% barley over 3 days to reach a diet containing 55.6% DM, 31.3% DM neutral detergent fiber, and 31.8% DM starch, which was fed for four more days. Rumen fluid samples were collected at day 7 and 10 of experimental periods. Feces samples were collected on days 8 and 10 of these periods. Extracted DNA from the rumen and feces samples was analyzed to assess their bacterial communities using MiSeq Illumina sequencing of the V4 region of the 16S rRNA gene. The induction of SARA reduced the richness, diversity, and stability of bacterial communities and resulted in distinctly different microbiota in the rumen and feces. Bacteroidetes and Firmicutes were the most abundant phyla and, combined, they represented 76.9 and 94.4% of the bacterial community in the rumen fluid and the feces, respectively. Only the relative abundance of Firmicutes in the rumen was increased by the SARA challenge. In rumen fluid and feces, the abundances of nine out of the 90 and 25 out of the 89 taxa, respectively, were affected by the challenge. Hence, SARA challenge altered the composition of the bacterial community at the lower taxonomical level in the feces and therefore also likely in the hindgut, as well as in the rumen. However, only reductions in the bacterial richness and diversity in the rumen fluid and feces were in agreement with those of other studies and had a biological basis. Although the composition of the

  20. Estimating Sampling Selection Bias in Human Genetics: A Phenomenological Approach

    Science.gov (United States)

    Risso, Davide; Taglioli, Luca; De Iasio, Sergio; Gueresi, Paola; Alfani, Guido; Nelli, Sergio; Rossi, Paolo; Paoli, Giorgio; Tofanelli, Sergio

    2015-01-01

    This research is the first empirical attempt to calculate the various components of the hidden bias associated with the sampling strategies routinely-used in human genetics, with special reference to surname-based strategies. We reconstructed surname distributions of 26 Italian communities with different demographic features across the last six centuries (years 1447–2001). The degree of overlapping between "reference founding core" distributions and the distributions obtained from sampling the present day communities by probabilistic and selective methods was quantified under different conditions and models. When taking into account only one individual per surname (low kinship model), the average discrepancy was 59.5%, with a peak of 84% by random sampling. When multiple individuals per surname were considered (high kinship model), the discrepancy decreased by 8–30% at the cost of a larger variance. Criteria aimed at maximizing locally-spread patrilineages and long-term residency appeared to be affected by recent gene flows much more than expected. Selection of the more frequent family names following low kinship criteria proved to be a suitable approach only for historically stable communities. In any other case true random sampling, despite its high variance, did not return more biased estimates than other selective methods. Our results indicate that the sampling of individuals bearing historically documented surnames (founders' method) should be applied, especially when studying the male-specific genome, to prevent an over-stratification of ancient and recent genetic components that heavily biases inferences and statistics. PMID:26452043

  1. Recognition of human face based on improved multi-sample

    Institute of Scientific and Technical Information of China (English)

    LIU Xia; LI Lei-lei; LI Ting-jun; LIU Lu; ZHANG Ying

    2009-01-01

    In order to solve the problem caused by variation illumination in human face recognition, we bring forward a face recognition algorithm based on the improved muhi-sample. In this algorithm, the face image is processed with Retinex theory, meanwhile, the Gabor filter is adopted to perform the feature extraction. The experimental results show that the application of Retinex theory improves the recognition accuracy, and makes the algorithm more robust to the variation illumination. The Gabor filter is more effective and accurate for extracting more useable facial local features. It is proved that the proposed algorithm has good recognition accuracy and it is stable under variation illumination.

  2. Cr and Yb markers determination in animal feces by energy dispersive X-ray fluorescence

    Energy Technology Data Exchange (ETDEWEB)

    Almeida, Eduardo de; Senicato, Luis A; Nascimento Filho, Virgilio F. [Centro de Energia Nuclear na Agricultura (CENA), Piracicaba, SP (Brazil). Lab. de Instrumentacao Nuclear (LIN)]. E-mail: edualm@usp.br; Gomide, Catarina A. [Universidade de Sao Paulo (USP), Pirassununga, SP (Brazil). Faculdade de Zootecnia e Engenharia de Alimentos (FZEA). Dept. de Zootecnia]. E-mail: cbgomide@usp.br

    2007-07-01

    Chromium and Ytterbium elements are utilized in animal nutritional studies as markers. This paper describes an analytical method for Cr and Yb determination in solid buffalo feces sample using standard addition method and energy dispersive X-ray spectrometry (EDXRF) technique. One gram dried sample was pressed manually in an XRF sample cup with Mylar film (6.3 {mu}m thickness) in the bottom. The experimental conditions were: Mo target X-ray tube with Zr filter, operated at 25 kV/10 mA, and 500 s of acquisition time. The limits of detection for Cr and Yb were 16.6 and 11.4 mg/kg, respectively. This methodology has showed appropriated for simultaneous Cr and Yb determination as marker in animal feces. (author)

  3. Human papillomavirus detection from human immunodeficiency virus-infected Colombian women's paired urine and cervical samples.

    Science.gov (United States)

    Munoz, Marina; Camargo, Milena; Soto-De Leon, Sara C; Sanchez, Ricardo; Parra, Diana; Pineda, Andrea C; Sussmann, Otto; Perez-Prados, Antonio; Patarroyo, Manuel E; Patarroyo, Manuel A

    2013-01-01

    Infection, coinfection and type-specific human papillomavirus (HPV) distribution was evaluated in human immunodeficiency virus (HIV)-positive women from paired cervical and urine samples. Paired cervical and urine samples (n = 204) were taken from HIV-positive women for identifying HPV-DNA presence by using polymerase chain reaction (PCR) with three generic primer sets (GP5+/6+, MY09/11 and pU1M/2R). HPV-positive samples were typed for six high-risk HPV (HR-HPV) (HPV-16, -18, -31, -33, -45 and -58) and two low-risk (LR-HPV) (HPV-6/11) types. Agreement between paired sample results and diagnostic performance was evaluated. HPV infection prevalence was 70.6% in cervical and 63.2% in urine samples. HPV-16 was the most prevalent HPV type in both types of sample (66.7% in cervical samples and 62.0% in urine) followed by HPV-31(47.2%) in cervical samples and HPV-58 (35.7%) in urine samples. There was 55.4% coinfection (infection by more than one type of HPV) in cervical samples and 40.2% in urine samples. Abnormal Papanicolau smears were observed in 25.3% of the women, presenting significant association with HPV-DNA being identified in urine samples. There was poor agreement of cervical and urine sample results in generic and type-specific detection of HPV. Urine samples provided the best diagnosis when taking cytological findings as reference. In conclusion including urine samples could be a good strategy for ensuring adherence to screening programs aimed at reducing the impact of cervical cancer, since this sample is easy to obtain and showed good diagnostic performance.

  4. Human papillomavirus detection from human immunodeficiency virus-infected Colombian women's paired urine and cervical samples.

    Directory of Open Access Journals (Sweden)

    Marina Munoz

    Full Text Available Infection, coinfection and type-specific human papillomavirus (HPV distribution was evaluated in human immunodeficiency virus (HIV-positive women from paired cervical and urine samples. Paired cervical and urine samples (n = 204 were taken from HIV-positive women for identifying HPV-DNA presence by using polymerase chain reaction (PCR with three generic primer sets (GP5+/6+, MY09/11 and pU1M/2R. HPV-positive samples were typed for six high-risk HPV (HR-HPV (HPV-16, -18, -31, -33, -45 and -58 and two low-risk (LR-HPV (HPV-6/11 types. Agreement between paired sample results and diagnostic performance was evaluated. HPV infection prevalence was 70.6% in cervical and 63.2% in urine samples. HPV-16 was the most prevalent HPV type in both types of sample (66.7% in cervical samples and 62.0% in urine followed by HPV-31(47.2% in cervical samples and HPV-58 (35.7% in urine samples. There was 55.4% coinfection (infection by more than one type of HPV in cervical samples and 40.2% in urine samples. Abnormal Papanicolau smears were observed in 25.3% of the women, presenting significant association with HPV-DNA being identified in urine samples. There was poor agreement of cervical and urine sample results in generic and type-specific detection of HPV. Urine samples provided the best diagnosis when taking cytological findings as reference. In conclusion including urine samples could be a good strategy for ensuring adherence to screening programs aimed at reducing the impact of cervical cancer, since this sample is easy to obtain and showed good diagnostic performance.

  5. Spectrophotometric assay of creatinine in human serum sample

    Directory of Open Access Journals (Sweden)

    Avinash Krishnegowda

    2017-05-01

    Full Text Available A new spectrophotometric method for the analysis of creatinine concentration in human serum samples is developed. The method explores the oxidation of p-methylamino phenol sulfate (Metol in the presence of copper sulfate and creatinine which yields an intense violet colored species with maximum absorbance at 530 nm. The calibration graph of creatinine by fixed time assay ranged from 4.4 to 620 μM. Recovery of creatinine in human serum samples varied from 101% to 106%. Limit of detection and limit of quantification were 0.145 μM and 0.487 μM respectively. Sandell’s sensitivity was 0.112 μg cm−2 and molar absorptivity was 0.101 × 104 L mol−1 cm−1. Within day precision was 2.5–4.8% and day-to-day precision range was 3.2–7.8%. The robustness and ruggedness of the method expressed in RSD values ranged from 0.78% to 2.12% and 1.32% to 3.46% respectively, suggesting that the developed method was rugged. This method provides good sensitivity and is comparable to standard Jaffe’s method with comparatively less interference from foreign substances.

  6. Rapid extraction and preservation of genomic DNA from human samples.

    Science.gov (United States)

    Kalyanasundaram, D; Kim, J-H; Yeo, W-H; Oh, K; Lee, K-H; Kim, M-H; Ryew, S-M; Ahn, S-G; Gao, D; Cangelosi, G A; Chung, J-H

    2013-02-01

    Simple and rapid extraction of human genomic DNA remains a bottleneck for genome analysis and disease diagnosis. Current methods using microfilters require cumbersome, multiple handling steps in part because salt conditions must be controlled for attraction and elution of DNA in porous silica. We report a novel extraction method of human genomic DNA from buccal swab and saliva samples. DNA is attracted onto a gold-coated microchip by an electric field and capillary action while the captured DNA is eluted by thermal heating at 70 °C. A prototype device was designed to handle four microchips, and a compatible protocol was developed. The extracted DNA using microchips was characterized by qPCR for different sample volumes, using different lengths of PCR amplicon, and nuclear and mitochondrial genes. In comparison with a commercial kit, an equivalent yield of DNA extraction was achieved with fewer steps. Room-temperature preservation for 1 month was demonstrated for captured DNA, facilitating straightforward collection, delivery, and handling of genomic DNA in an environment-friendly protocol.

  7. FANTOM5 CAGE profiles of human and mouse samples

    KAUST Repository

    Noguchi, Shuhei

    2017-08-29

    In the FANTOM5 project, transcription initiation events across the human and mouse genomes were mapped at a single base-pair resolution and their frequencies were monitored by CAGE (Cap Analysis of Gene Expression) coupled with single-molecule sequencing. Approximately three thousands of samples, consisting of a variety of primary cells, tissues, cell lines, and time series samples during cell activation and development, were subjected to a uniform pipeline of CAGE data production. The analysis pipeline started by measuring RNA extracts to assess their quality, and continued to CAGE library production by using a robotic or a manual workflow, single molecule sequencing, and computational processing to generate frequencies of transcription initiation. Resulting data represents the consequence of transcriptional regulation in each analyzed state of mammalian cells. Non-overlapping peaks over the CAGE profiles, approximately 200,000 and 150,000 peaks for the human and mouse genomes, were identified and annotated to provide precise location of known promoters as well as novel ones, and to quantify their activities.

  8. Corn stalk as matrix in decomposting toilet for treating urine and feces

    Science.gov (United States)

    Sintawardani, N.; Nilawati, D.; Astuti, J. T.

    2017-03-01

    Bio-Toilet technology (BT) which is appropriate for the habits of Indonesian people has been studied and developed. BT is a dry toilet technology commonly uses ligno-cellulosic waste materials as matrix to facilitate the growth of natural microbes. In aerobic condition, microbes degrade feces and urine. Mineral as the leftover of feces and urine, such as nitrogen (N), phosphorus (P), and potassium (K) remain in the rest of matrix waste. After certain period. matrix can be harvested and used as soil conditioner. BT uses much less water, mobile, and very useful to be applied in areas where water availability is limited. BT type with different capacities, user amounts and mixing systems has been developed using sawdust for matrix. Since corn stalk is categorized as useless and priceless waste, its application in BT is challenging. Performance of BT with corn stalk as matrix to degrade feces and urine of carnivore imitating the human waste was observed. BT M-15 manual mixing type with paddle was filled with chopped corn stalk as much as 45% of total volume. This BT was designed for 15 person as users per day if 80% reactor volume was filled with ligno-cellulosic matrix. It is assumed that 150 g of feces are discharged once per person/day and 1000 mL of urine 6-8 times per day. Start up process was made in the beginning to initialize the needed microbes in the reactor (matrix). The discharge of feces and urine were increased slowly and gradually the users were increased from 1 to 4 users per day. Performance of BT was indicated by the change in the pile that showed by moisture content, temperature and pH. C/N ratio in matrix decreased significantly from 43 to 17. This result showed that the corn stalk could be used as matrix in BT.

  9. Isolation of Cancer Stem Cells From Human Prostate Cancer Samples

    Science.gov (United States)

    Vidal, Samuel J.; Quinn, S. Aidan; de la Iglesia-Vicente, Janis; Bonal, Dennis M.; Rodriguez-Bravo, Veronica; Firpo-Betancourt, Adolfo; Cordon-Cardo, Carlos; Domingo-Domenech, Josep

    2014-01-01

    The cancer stem cell (CSC) model has been considerably revisited over the last two decades. During this time CSCs have been identified and directly isolated from human tissues and serially propagated in immunodeficient mice, typically through antibody labeling of subpopulations of cells and fractionation by flow cytometry. However, the unique clinical features of prostate cancer have considerably limited the study of prostate CSCs from fresh human tumor samples. We recently reported the isolation of prostate CSCs directly from human tissues by virtue of their HLA class I (HLAI)-negative phenotype. Prostate cancer cells are harvested from surgical specimens and mechanically dissociated. A cell suspension is generated and labeled with fluorescently conjugated HLAI and stromal antibodies. Subpopulations of HLAI-negative cells are finally isolated using a flow cytometer. The principal limitation of this protocol is the frequently microscopic and multifocal nature of primary cancer in prostatectomy specimens. Nonetheless, isolated live prostate CSCs are suitable for molecular characterization and functional validation by transplantation in immunodeficient mice. PMID:24686446

  10. Phylogenetic analysis of Escherichia coli strains isolated from human samples

    Directory of Open Access Journals (Sweden)

    Abdollah Derakhshandeh

    2013-12-01

    Full Text Available Escherichia coli (E. coli is a normal inhabitant of the gastrointestinal tract of vertebrates, including humans. Phylogenetic analysis has shown that E. coli is composed of four main phylogenetic groups (A, B1, B2 and D. Group A and B1 are generally associated with commensals, whereas group B2 is associated with extra-intestinal pathotypes. Most enteropathogenic isolates, however, are assigned to group D. In the present study, a total of 102 E. coli strains, isolated from human samples, were used. Phylogenetic grouping was done based on the Clermont triplex PCR method using primers targeted at three genetic markers, chuA, yjaA and TspE4.C2. Group A contained the majority of the collected isolates (69 isolates, 67.64%, followed by group B2 (18 isolates, 17.64% and D (15 isolates, 14.7% and no strains were found to belong to group B1. The distribution of phylogenetic groups in our study suggests that although the majority of strains were commensals, the prevalence of enteropathogenic and extra-intestinal pathotypes was noteworthy. Therefore, the role of E. coli in human infections including diarrhea, urinary tract infections and meningitis should be considered.

  11. Longitudinal characterization of antimicrobial resistance genes in feces shed from cattle fed different subtherapeutic antibiotics

    Directory of Open Access Journals (Sweden)

    Read Ronald R

    2011-01-01

    Full Text Available Abstract Background Environmental transmission of antimicrobial-resistant bacteria and resistance gene determinants originating from livestock is affected by their persistence in agricultural-related matrices. This study investigated the effects of administering subtherapeutic concentrations of antimicrobials to beef cattle on the abundance and persistence of resistance genes within the microbial community of fecal deposits. Cattle (three pens per treatment, 10 steers per pen were administered chlortetracycline, chlortetracycline plus sulfamethazine, tylosin, or no antimicrobials (control. Model fecal deposits (n = 3 were prepared by mixing fresh feces from each pen into a single composite sample. Real-time PCR was used to measure concentrations of tet, sul and erm resistance genes in DNA extracted from composites over 175 days of environmental exposure in the field. The microbial communities were analyzed by quantification and denaturing gradient gel electrophoresis (DGGE of PCR-amplified 16S-rRNA. Results The concentrations of 16S-rRNA in feces were similar across treatments and increased by day 56, declining thereafter. DGGE profiles of 16S-rRNA differed amongst treatments and with time, illustrating temporal shifts in microbial communities. All measured resistance gene determinants were quantifiable in feces after 175 days. Antimicrobial treatment differentially affected the abundance of certain resistance genes but generally not their persistence. In the first 56 days, concentrations of tet(B, tet(C, sul1, sul2, erm(A tended to increase, and decline thereafter, whereas tet(M and tet(W gradually declined over 175 days. At day 7, the concentration of erm(X was greatest in feces from cattle fed tylosin, compared to all other treatments. Conclusion The abundance of genes coding for antimicrobial resistance in bovine feces can be affected by inclusion of antibiotics in the feed. Resistance genes can persist in feces from cattle beyond 175 days

  12. Nest sanitation through defecation: antifungal properties of wood cockroach feces

    Science.gov (United States)

    Rosengaus, Rebeca B.; Mead, Kerry; Du Comb, William S.; Benson, Ryan W.; Godoy, Veronica G.

    2013-11-01

    The wood cockroach Cryptocercus punctulatus nests as family units inside decayed wood, a substrate known for its high microbial load. We tested the hypothesis that defecation within their nests, a common occurrence in this species, reduces the probability of fungal development. Conidia of the entomopathogenic fungus, Metarhizium anisopliae, were incubated with crushed feces and subsequently plated on potato dextrose agar. Relative to controls, the viability of fungal conidia was significantly reduced following incubation with feces and was negatively correlated with incubation time. Although the cockroach's hindgut contained abundant β-1,3-glucanase activity, its feces had no detectable enzymatic function. Hence, these enzymes are unlikely the source of the fungistasis. Instead, the antifungal compound(s) of the feces involved heat-sensitive factor(s) of potential microbial origin. When feces were boiled or when they were subjected to ultraviolet radiation and subsequently incubated with conidia, viability was "rescued" and germination rates were similar to those of controls. Filtration experiments indicate that the fungistatic activity of feces results from chemical interference. Because Cryptocercidae cockroaches have been considered appropriate models to make inferences about the factors fostering the evolution of termite sociality, we suggest that nesting in microbe-rich environments likely selected for the coupling of intranest defecation and feces fungistasis in the common ancestor of wood cockroaches and termites. This might in turn have served as a preadaptation that prevented mycosis as these phylogenetically related taxa diverged and evolved respectively into subsocial and eusocial organizations.

  13. Determination of Hg and diet identification in otter (Lontra longicaudis) feces

    Energy Technology Data Exchange (ETDEWEB)

    Ferreira Josef, Carla [Centro de Energia Nuclear na Agricultura - CENA-USP, Universidade de Sao Paulo, Av. Centenario 303, 13400-970 Piracicaba, Sao Paulo (Brazil)], E-mail: carlajosef@hotmail.com; Ramos Adriano, Leonardo; Franca, Elvis Joacir de; Gustinelli Arantes de Carvalho, Gabriel [Centro de Energia Nuclear na Agricultura - CENA-USP, Universidade de Sao Paulo, Av. Centenario 303, 13400-970 Piracicaba, Sao Paulo (Brazil); Ferreira, Jose Roberto [Centro de Energia Nuclear na Agricultura - CENA-USP, Universidade de Sao Paulo, Av. Centenario 303, 13400-970 Piracicaba, Sao Paulo (Brazil); Polo de Pesquisa e Desenvolvimento Regional do Centro Sul - APTA, Agencia Paulista de Tecnologia dos Agronegocios. Rua Alberto Coral 1500, 13400-000 Piracicaba, Sao Paulo (Brazil)

    2008-04-15

    An analytical procedure for the determination of Hg in otter (Lontra longicaudis) feces was developed, to separate fish scales for the identification of the animal diet. Samples were washed with ultra-pure water and the suspension was sampled and transferred for digestion. The solubilization was performed with nitric-perchloric acid mixture, and detection carried out by the atomic fluorescence spectrometry (AFS). The quality of the analytical procedure was assessed by analyzing in-house standard solutions and certified reference materials. Total Hg concentrations were in the range of 7.6-156 ng g{sup -1} (July 2004), 25.6-277 ng g{sup -1} (January 2005) and 14.6-744 ng g{sup -1} (May 2005) that is approximately the same order of magnitude for all samples collected in two reservoirs at the Tiete River, Brazil. Although Hg concentrations varied with sampling periods and diet, high levels were correlated to the percentage of carnivorous fish scales present in the otter feces. - The importance of otter feces preparation for Hg analysis, focusing the food web.

  14. CTX-M producing Escherichia coli isolated from cattle feces in Bogor slaughterhouse,Indonesia

    Institute of Scientific and Technical Information of China (English)

    Mirnawati Bachrum Sudarwanto; Denny Widaya Lukman; Hadri Latif; Herwin Pisestyani; Eddy Sukmawinata; mer Akineden; Ewald Usleber

    2016-01-01

    Objective: To determine the occurrence of CTX-M producing Escherichia coli(E. coli)from cattle feces in Bogor slaughterhouse, Indonesia.Methods: A total of 220 cattle feces samples were collected from Bogor slaughterhouse from March to April 2015. Presence of extended-spectrum beta-lactamase(ESBL) producing E. coli was detected by disc diffusion test based on the recommendation from Clinical and Laboratory Standards Institute(2014). Bacterial strains which were confirmed as producing ESBLs were further analyzed for the presence of bla genes of the ESBL by PCR.Results: The results showed that CTX-M producing E. coli isolates were detected in 19 samples from 220 samples(8.6%). The b-lactamase genes detected were CTX-M-1(n = 10) and CTX-M-9(n = 9). All of the CTX-M producing E. coli isolates showed multidrug resistance phenotypes to at least four antibiotics. The highest incidence of antibiotics resistance was showed to ampicillin(100.0%), cefotaxime(100.0%), and cefpodoxime(100.0%), followed by streptomycin(84.3%), trimethoprim-sulfamethoxazole(73.7%), erythromycin(52.6%), kanamycin(26.3%), doxycycline(10.5%), and ceftazidime(0.0%).Conclusions: Detection of CTX-M-producing E. coli in cattle feces raises important questions as they can represent a potential risk factor to public health.

  15. CTX-M producing Escherichia coli isolated from cattle feces in Bogor slaughterhouse, Indonesia

    Institute of Scientific and Technical Information of China (English)

    Mirnawati Bachrum Sudarwanto; Denny Widaya Lukman; Hadri Latif; Herwin Pisestyani; Eddy Sukmawinata; Omer Akineden; Ewald Usleber

    2016-01-01

    Objective: To determine the occurrence of CTX-M producing Escherichia coli (E. coli) from cattle feces in Bogor slaughterhouse, Indonesia. Methods: A total of 220 cattle feces samples were collected from Bogor slaughterhouse from March to April 2015. Presence of extended-spectrum beta-lactamase (ESBL) producing E. coli was detected by disc diffusion test based on the recommendation from Clinical and Laboratory Standards Institute (2014). Bacterial strains which were confirmed as producing ESBLs were further analyzed for the presence of bla genes of the ESBL by PCR. Results: The results showed that CTX-M producing E. coli isolates were detected in 19 samples from 220 samples (8.6%). The b-lactamase genes detected were CTX-M-1 (n = 10) and CTX-M-9 (n = 9). All of the CTX-M producing E. coli isolates showed multidrug resistance phenotypes to at least four antibiotics. The highest incidence of an-tibiotics resistance was showed to ampicillin (100.0%), cefotaxime (100.0%), and cef-podoxime (100.0%), followed by streptomycin (84.3%), trimethoprim-sulfamethoxazole (73.7%), erythromycin (52.6%), kanamycin (26.3%), doxycycline (10.5%), and ceftazi-dime (0.0%). Conclusions: Detection of CTX-M-producing E. coli in cattle feces raises important questions as they can represent a potential risk factor to public health.

  16. Evaluation of chromium concentration in cattle feces using different acid digestion and spectrophotometric quantification techniques

    Directory of Open Access Journals (Sweden)

    N.K.P. Souza

    2013-10-01

    Full Text Available The objective of this work was to evaluate combinations between acid digestion techniques and spectrophotometric quantification to measure chromium concentration in cattle feces. Digestion techniques were evaluated based on the use of nitric and perchloric acids, sulfuric and perchloric acids, and phosphoric acid. The chromium quantification in the solutions was performed by colorimetry and by atomic absorption spectrophotometry (AAS. When AAS was used, the addition of calcium chloride to the solutions as a releasing agent was also evaluated. Several standard samples containing known chromium contents were produced (0, 2, 4, 6, 8 and 10g of chromium per kg of feces using cattle feces obtained from three different animals to evaluate the accuracy of the different combinations of techniques. The accuracy was evaluated by adjusting a simple linear regression model of the estimated values on the actual values of chromium content in the standard samples. Regardless of the digestion technique, the chromium content estimates in the standard samples obtained by colorimetry were not accurate (P0.05. The use of the digestion technique in phosphoric acid provided incomplete recovery of the fecal chromium (P0.05 fecal chromium contents.

  17. Study on fingerprint and identification of origin of feces%粪便源指纹及其鉴别研究

    Institute of Scientific and Technical Information of China (English)

    徐恒振; 马新东; 王洪艳; 周传光; 姚子伟

    2013-01-01

    利用气相色谱-质谱联用仪检测了人和32种动物粪便中以及5个不同排污口表层沉积物中10种粪固醇,以10种粪固醇含量和9个粪固醇比值为指标,分别应用层次聚类分析法对人和32种动物粪便以及5个沉积物的粪便污染来源进行了鉴别,结果表明:草食动物粪便中主要含C29-甾醇,5β-甾醇比5α-甾醇含量多,草食动物粪便的主要指示物是24-乙基粪醇和谷甾醇;肉食动物粪便中主要含C27-甾醇,胆固醇是肉食动物的主要指示物;杂食粪便中粪固醇组成多变;人粪便中主要含粪醇,占总粪固醇的51.5%;应用10种粪固醇含量和9个粪固醇比值以及层次聚类分析法,可以准确地鉴别人和32种动物粪便以及5个沉积物样品的粪便污染来源.%10 kinds of faecal sterols from feces in humans and 32 kinds of animals and surface sediments from 5 different outfalls of sewages were analyzed by GC/MS. The origin of pollution from the feces was identified by the contents and the ratio values of faecal sterols with the hierarchical cluster analysis, respectively. The results showed that the profiles of C29 -sterols were dominated, and 5|3-stanols were greater abundance than 5α-stanols in the feces from herbivores, and 24-ethycoprostanol and sitosterol were as the principal faecal biomarker for herbivores. The profiles of C27 -sterols in the feces from carnivores were dominated and cholesterol was as the principal faecal biomarker of carnivores. It showed that the lots of sterols in faces from omnivores were varied, and the major of faecal sterol from feces of human was coprostanol constituted 51.5% of the total sterols in human faeces. It also showed that the origins of pollution from feces in humans and 32 kinds of animals and samples of surface sediments from 5 different outfalls of sewages could be accurately identified by 10 kinds of contents and 9 types of ratio values of faecal sterols with the hierarchical cluster

  18. Molecular detection of hepatitis E virus in feces and slurry from swine farms, Rio Grande do Sul, Southern Brazil

    Directory of Open Access Journals (Sweden)

    J. Vasconcelos

    2015-06-01

    Full Text Available Hepatitis E virus (HEV is highly disseminated among swine herds worldwide. HEV is also a threat to public health, since particularly genotypes 3 and 4 may cause acute hepatitis in human beings. No previous studies were done on the occurrence of HEV in environmental samples in Rio Grande do Sul, Brazil. In the present study, reverse transcriptase-polymerase chain reaction (RT-PCR was employed to detect the presence of HEV in swine feces and in effluents from slurry lagoons in farms located in the municipality of Teutônia, inside the area of swine husbandry in the state. Pooled fecal samples from the floor of pig barns from 9 wean-to-finish farms and liquid manure samples were collected from the slurry lagoons from 8 of these farms. From the pooled fecal samples, 8/9 were positive for the HEV ORF1 gene by RT-PCR; all the slurry lagoon samples were positive for HEV RNA (100%. The identity of the HEV ORF1 amplicons was confirmed by sequencing belonging to HEV genotype 3, which was previously shown to be circulating in South America.

  19. Effect on biodegradation and nitrogen holding property from temperature during aerobic composting for sanitary disposal of human feces%粪便好氧堆肥过程中温度对有机物的降解和氮的保持特性影响

    Institute of Scientific and Technical Information of China (English)

    白帆; 王晓昌

    2011-01-01

    It would be favorable to improve biodegradation of feces and hold more nitrogen in compost if the effect on biodegradation and nitrogen holding property from temperature was found out during aerobic composting in bio-toilet for sanitary disposal of human feces.In this study, batch experiments were conducted using a closed aerobic composting reactor with sawdust as the bulky matrix to simulate the condition of a bio-toilet for sanitary disposal of human feces.Attention was paid to the effect on biodegradation and nitrogen holding property from temperature.Under a controlled condition of temperature at 60℃ and 35℃, moisture content at 60%, and continuous air supply, more than 70% fecal organic removal was obtained with merely 17% fecal nitrogen loss observed at 60℃ in a two-week composting period, while more than 63% fecal organic removal with 31.4% fecal nitrogen loss at 35℃.Compost maturity period decreased from 10 ~ 12 days at 35℃ to 6 ~8 days at 60℃.The nitrogen loss ( 17% ) was found to occur mainly in the first day with quick depletion of inorganic nitrogen, but almost unchanged organic nitrogen content at 60℃.While the nitrogen loss ( 31.4% ) was mainly in the first 4 days from both inorganic nitrogen and organic nitrogen content at 35℃.The result showed that temperature has obviously effect on composting, controlling temperature could improve compost effect, shorten eomposting maturity period, decreased fecal nitrogen loss, keep high organic nitrogen content in the composts for better fertilizer utilization.%研究堆肥过程温度对有机物的降解,尤其是氮的迁移转化的影响,对于提高堆肥效率和保持更多的氮在产物中具有重要意义.本研究采用密闭式好氧堆肥反应器,模拟高温(60℃)和中温(35℃)两种典型的堆肥温度,以新鲜锯末为空白载体,在含水率60%以及连续强制供气的条件下,进行了为期两周的试验,评估温度对于粪便中有机物的降

  20. Semi-automated extraction of microbial DNA from feces for qPCR and phylogenetic microarray analysis

    NARCIS (Netherlands)

    Nylund, L.; Heilig, G.H.J.; Salminen, S.; Vos, de W.M.; Satokari, R.M.

    2010-01-01

    The human gastrointestinal tract (GI-tract) harbors a complex microbial ecosystem, largely composed of so far uncultured species, which can be detected only by using techniques such as PCR and by different hybridization techniques including phylogenetic microarrays. Manual DNA extraction from feces

  1. Semi-automated extraction of microbial DNA from feces for qPCR and phylogenetic microarray analysis

    NARCIS (Netherlands)

    Nylund, L.; Heilig, G.H.J.; Salminen, S.; Vos, de W.M.; Satokari, R.M.

    2010-01-01

    The human gastrointestinal tract (GI-tract) harbors a complex microbial ecosystem, largely composed of so far uncultured species, which can be detected only by using techniques such as PCR and by different hybridization techniques including phylogenetic microarrays. Manual DNA extraction from feces

  2. Probiotic properties of lactobacilli species isolated from children's feces.

    Science.gov (United States)

    Tulumoglu, Sener; Yuksekdag, Zehra Nur; Beyatli, Yavuz; Simsek, Omer; Cinar, Berat; Yaşar, Esra

    2013-12-01

    In the present research, the 20 lactobacilli isolated from children feces aged 4-15 years old were investigated for their capabilities to survive at pH 2.0, 2.5, 3.0 and in the presence of 0.25, 0.50 and 0.75% bile salts, their effect on the growth of pathogens, in addition to their sensitivity against 13 selected antibiotics. All the lactobacilli strains were able to survive in low pH and bile salt conditions at pH 2.0 and 0.25% bile salt for 2 h. Moreover, all lactobacilli strains exhibited inhibitory activity against Escherichia coli ATCC 11229, Pseudomonas aeruginosa ATCC 27853 and Staphylococcus aureus ATCC 29213. In addition, all lactobacilli strains indicated resistance to teicoplanin, vancomycin, and bacitracin. The amount of exopolysaccharide (EPS) produced by the strains was 70 and 290 mg/L. The capabilities to autoaggregation and coaggregate with E. coli ATCC 11229 of the strains were also evaluated. High EPS-producing strains indicated significant autoaggregation and coaggregation capability with test bacteria (p lactobacilli could be utilized for preliminary screening in order to identify potentially probiotic bacteria suitable for human.

  3. Strong-LAMP: A LAMP Assay for Strongyloides spp. Detection in Stool and Urine Samples. Towards the Diagnosis of Human Strongyloidiasis Starting from a Rodent Model

    Science.gov (United States)

    Gandasegui, Javier; Bajo Santos, Cristina; López-Abán, Julio; Saugar, José María; Rodríguez, Esperanza; Vicente, Belén; Muro, Antonio

    2016-01-01

    Background Strongyloides stercoralis, the chief causative agent of human strongyloidiasis, is a nematode globally distributed but mainly endemic in tropical and subtropical regions. Chronic infection is often clinically asymptomatic but it can result in severe hyperinfection syndrome or disseminated strongyloidiasis in immunocompromised patients. There is a great diversity of techniques used in diagnosing the disease, but definitive diagnosis is accomplished by parasitological examination of stool samples for morphological identification of parasite. Until now, no molecular method has been tested in urine samples as an alternative to stool samples for diagnosing strongyloidiasis. This study aimed to evaluate the use of a new molecular LAMP assay in a well-established Wistar rat experimental infection model using both stool and, for the first time, urine samples. The LAMP assay was also clinically evaluated in patients´ stool samples. Methodology/Principal Findings Stool and urine samples were obtained daily during a 28-day period from rats infected subcutaneously with different infective third-stage larvae doses of S. venezuelensis. The dynamics of parasite infection was determined by daily counting the number of eggs per gram of feces from day 1 to 28 post-infection. A set of primers for LAMP assay based on a DNA partial sequence in the 18S rRNA gene from S. venezuelensis was designed. The set up LAMP assay (namely, Strong-LAMP) allowed the sensitive detection of S. venezuelensis DNA in both stool and urine samples obtained from each infection group of rats and was also effective in S. stercoralis DNA amplification in patients´ stool samples with previously confirmed strongyloidiasis by parasitological and real-time PCR tests. Conclusions/Significance Our Strong-LAMP assay is an useful molecular tool in research of a strongyloidiasis experimental infection model in both stool and urine samples. After further validation, the Strong-LAMP could also be potentially

  4. Antibiotic effect of amoxicillin on the feces composting process and reactivation of bacteria by intermittent feeding of feces.

    Science.gov (United States)

    Kakimoto, Takashi; Osawa, Teruma; Funamizu, Naoyuki

    2007-12-01

    The aim of this work was to investigate the single exposure effect of amoxicillin on the feces composting process and the possibility of its bacterial reactivation by intermittent feeding of feces. The respiratory activity of the bacteria during the process after receiving exposure to several dosages of amoxicillin indicated a decrement of treatment performance, which was caused by the reduction of the initial viable bacterial count and activity brought about by the amoxicillin dosage. The amount of remaining feces was proportional to the initial concentration of amoxicillin, and even though no amoxicillin was detected, no automatic reactivation was observed, either. An intermittent feces feeding test was conducted to reactivate the bacteria. For the 10 and 100microg-amoxicillin/g dry systems, reactivation was observed, but for the 1000microg/g dry, no reactivation was seen. Finally, an intermittent feces feeding test was conducted with sterilized sawdust and the result indicated that the feces acted as a substrate rather than as a bacterial carrier.

  5. DNA extraction from protozoan oocysts/cysts in feces for diagnostic PCR.

    Science.gov (United States)

    Hawash, Yousry

    2014-06-01

    PCR detection of intestinal protozoa is often restrained by a poor DNA recovery or by inhibitors present in feces. The need for an extraction protocol that can overcome these obstacles is therefore clear. QIAamp® DNA Stool Mini Kit (Qiagen) was evaluated for its ability to recover DNA from oocysts/cysts directly from feces. Twenty-five Giardia-positive, 15 Cryptosporidium-positive, 15 Entamoeba histolytica-positive, and 45 protozoa-free samples were processed as control by microscopy and immunoassay tests. DNA extracts were amplified using 3 sets of published primers. Following the manufacturer's protocol, the kit showed sensitivity and specificity of 100% towards Giardia and Entamoeba. However, for Cryptosporidium, the sensitivity and specificity were 60% (9/15) and 100%, respectively. A series of optimization experiments involving various steps of the kit's protocol were conducted using Cryptosporidium-positive samples. The best DNA recoveries were gained by raising the lysis temperature to the boiling point for 10 min and the incubation time of the InhibitEX tablet to 5 min. Also, using a pre-cooled ethanol for nucleic acid precipitation and small elution volume (50-100 µl) were valuable. The sensitivity of the amended protocol to Cryptosporidium was raised to 100%. Cryptosporidium DNA was successfully amplified by either the first or the second primer set. When applied on parasite-free feces spiked with variable oocysts/cysts counts, ≈ 2 oocysts/cysts were theoretically enough for detection by PCR. To conclude, the Qiagen kit with the amended protocol was proved to be suitable for protozoan DNA extraction directly from feces and support PCR diagnosis.

  6. Evaluation of immunomagnetic separation and PCR for the detection of Escherichia coli O157 in animal feces and meats

    NARCIS (Netherlands)

    Islam, M.A.; Heuvelink, A.E.; Talukder, K.A.; Zwietering, M.H.; Boer, de E.

    2006-01-01

    Series of animal feces and meat samples artificially contaminated with strains of Escherichia coli O157 isolated from different sources were tested by both an immunomagnetic separation (IMS)-based method and a PCR method using primers specific for a portion of the rfbE gene of E. coli O157. IMS is l

  7. Evaluation of immunomagnetic separation and PCR for the detection of Escherichia coli O157 in animal feces and meats

    NARCIS (Netherlands)

    Islam, M.A.; Heuvelink, A.E.; Talukder, K.A.; Zwietering, M.H.; Boer, de E.

    2006-01-01

    Series of animal feces and meat samples artificially contaminated with strains of Escherichia coli O157 isolated from different sources were tested by both an immunomagnetic separation (IMS)-based method and a PCR method using primers specific for a portion of the rfbE gene of E. coli O157. IMS is l

  8. A molecular epidemiologic investigation of Salmonella from a meat source to the feces of captive cheetah (Acinonyx jubatus).

    Science.gov (United States)

    Venter, Estelle H; van Vuuren, Moritz; Carstens, Johann; van der Walt, Martha L; Nieuwoudt, Badenhorst; Steyn, Helena; Kriek, Nick P J

    2003-03-01

    Low cheetah (Acinonyx jubatus) birth rates were observed for a long time in a captive breeding facility in which Salmonella, which was possibly present in contaminated beef, was isolated from still-born lion (Panthera leo) cubs. Salmonella, including 14 isolates of Salmonella serovar typhimurium and 19 isolates of Salmonella serovar muenchen, was subsequently isolated 47 times from 378 meat samples at the facility during a 13-mo period. Salmonella, including 26 isolates of S. serovar typhimurium, 10 of S. serovar muenchen, and 11 other serovars, also was isolated 54 times from 119 fecal samples. Only three plasmid profiles were identified in 59 S. typhimurium isolates from both meat and fecal samples. Although random-amplified polymorphic DNA fingerprinting using different primers in the polymerase chain reaction was able to distinguish between S. typhimurium and S. muenchen and to demonstrate similar chromosomal DNA fingerprints in some of the isolates from meat and feces, the results were not consistent enough to prove that the Salmonella in the feces originated from contaminated meat. However, the predominance of only two serovars in the meat fed to carnivores and in the feces of these animals suggests that the meat was the source of the Salmonella organisms in the feces.

  9. Effect of fenbendazole and ivermectin on development of strongylate nematode eggs and larvae in calf feces.

    Science.gov (United States)

    Miller, J E; Morrison, D G

    1992-07-01

    Thirty-nine weaned steer calves (mean weight 284 kg) were maintained under dry-lot conditions and assigned (based on fecal nematode egg count) to one of three treatment groups of 13 animals each as follows: control (no treatment), fenbendazole (5 mg kg-1), and ivermectin (0.2 mg kg-1). Fecal samples were collected 12 h before treatment, at treatment, and 12, 24, 48 and 72 h after treatment for determination of nematode eggs per gram, and (after culture) infective larvae per gram and population distribution. The effect of treatment on egg development was observed in feces collected 12 and 24 h after treatment. There was essentially no difference in efficacy, based on egg counts, of fenbendazole and ivermectin. Egg count was reduced 100% by both anthelmintics at 72 h after treatment. Viability, based on percent of eggs reaching the infective larval stage, of developing stages at 12, 24, and 48 h after fenbendazole treatment was 0.1%, 1.1%, and 0%; after ivermectin treatment the corresponding values were 23.7%, 30.1%, and 28.6%, respectively. Fenbendazole treatment resulted in little or no development of eggs and/or larvae in feces deposited 12 and 24 h after treatment, whereas development proceeded normally (compared with the control group) in ivermectin treated feces. Population distribution of infective larvae was predominantly Haemonchus and Cooperia with some Ostertagia and Oesophagostomum.

  10. Salmonellae in fish feces analyzed by in situ hybridization and quantitative polymerase chain reaction.

    Science.gov (United States)

    Sha, Qiong; Forstner, Michael R J; Bonner, Timothy H; Hahn, Dittmar

    2013-09-01

    The potential of fish to transfer salmonellae from heterogeneous aquatic biofilms into feces was assessed in controlled aquarium studies with Suckermouth Catfish Hypostomus plecostomus and with biofilms inoculated with salmonellae. Neither the presence of catfish nor inoculation with salmonellae had detectable effects on the abundance of the microbial community. Densities of the microbial community were about 10(5) cells/mL in the water during a 1-week period, whereas densities of the microbial community increased 10-fold (10(6) to 10(7) cells/mg) in catfish feces during the same period. Salmonellae were detected by both quantitative polymerase chain reaction (qPCR) and situ hybridization in water samples immediately after inoculation, in numbers of about 10(4) cells/mL, representing up to 20% of the cells of the microbial community. Numbers decreased by three orders of magnitude within the first 3 d of the study, which represented only 0.01% of the community, and became undetectable after day 5. In catfish feces, numbers of Salmonella initially increased to up to 6% of the cells of the community but then declined. These results suggest that Salmonella are not biomagnified during gut passage, and thus, fish only provide a means for the translocation of this pathogen.

  11. Within-day repeatability for absolute quantification of Lawsonia intracellularis bacteria in feces from growing pigs

    DEFF Research Database (Denmark)

    Pedersen, Ken Steen; Pedersen, Klaus H.; Hjulsager, Charlotte Kristiane;

    2012-01-01

    standard deviation for individual pigs was 0.27 log10 bacteria/g feces. The average coefficient of variation within pigs was 0.04, ranging from 0.006 to 0.08. The results imply that absolute quantification of L. intracellularis by qPCR has acceptable repeatability within 1 day. However, a population...... of the current study was to investigate overall variation within a day for fecal numbers of L. intracellularis bacteria determined by real-time PCR in growing pigs. From each of 30 pigs with an infection of L. intracellularis, 5 fecal samples were collected within 1 day. A total of 150 fecal samples were...

  12. OCCURRENCE OF INTRINSIC VANCOMYCIN RESISTANT ENTEROCOCCI IN ANIMAL FECES

    Science.gov (United States)

    A survey was conducted to determine the occurrence of vancomycin resistant enterococci (VRE) in animal and human fecal samples. Fecal samples from 14 animal species and humans were analyzed by quantitative culture for enterococci and VRE. Over 800 VRE isolates were characterize...

  13. Clinical and epidemiologic characteristics of human bocavirus in Danish infants: results from a prospective birth cohort study

    DEFF Research Database (Denmark)

    von Linstow, Marie-Louise; Høgh, Mette; Høgh, Birthe

    2008-01-01

    BACKGROUND: Human bocavirus (HBoV) is a recently discovered parvovirus that has been detected in respiratory samples from children with acute respiratory tract infection (ARTI) and in feces from children with gastroenteritis. However, its role as a causative agent of respiratory disease is not de......BACKGROUND: Human bocavirus (HBoV) is a recently discovered parvovirus that has been detected in respiratory samples from children with acute respiratory tract infection (ARTI) and in feces from children with gastroenteritis. However, its role as a causative agent of respiratory disease...

  14. A polyclonal antibody based immunoassay detects seven subtypes of Shiga toxin 2 produced by Escherichia coli in human and environmental samples.

    Directory of Open Access Journals (Sweden)

    Xiaohua He

    Full Text Available BACKGROUND: Shiga toxin-producing Escherichia coli (STEC are frequent causes of severe human diseases ranging from diarrhea to hemolytic uremic syndrome. The existing strategy for detection of STEC relies on the unique sorbitol-negative fermentation property of the O157 strains, the most commonly identified serotype has been E. coli O157. It is becoming increasingly evident, however, that numerous non-O157 STEC serotypes also cause outbreaks and severe illnesses. It is necessary to have new methods that are capable of detecting all STEC strains. METHODS AND FINDINGS: Here we describe the development of a sandwich ELISA assay for detecting both O157 and non-O157 STECs by incorporating a novel polyclonal antibody (pAb against Stx2. The newly established immunoassay was capable of detecting Stx2a spiked in environmental samples with a limit of detection between 10 and 100 pg/mL in soil and between 100 and 500 pg/mL in feces. When applied to 36 bacterial strains isolated from human and environmental samples, this assay detected Stx2 in all strains that were confirmed to be stx2-positive by real-time PCR, demonstrating a 100% sensitivity and specificity. CONCLUSIONS: The sandwich ELISA developed in this study will enable any competent laboratory to identify and characterize Stx2-producing O157 and non-O157 strains in human and environmental samples, resulting in rapid diagnosis and patient care. The results of epitope mapping from this study will be useful for further development of a peptide-based antibody and vaccine.

  15. [Glycoprotein hexoses in feces of infants with lactose intolerance].

    Science.gov (United States)

    Filippvskiĭ, G K; Klimov, L Ia

    1995-01-01

    A modified method for estimation of total glycoprotein hexoses in feces, based on their measurements in the blood serum, is presented. Sixty-six nursing children with lactose intolerance, breastfed or formula fed, were examined; formula fed babies were kept on mixtures with high and low lactose content. Glycoprotein hexose parameters were as follows (X +/- m): 13.51 +/- 1.93, 12.05 +/- 2.20, and 3.69 +/- 0.47 g/l feces. In control children without lactose intolerance (n = 33) this value was 3.6 +/- 0.79 g/l. Increased glycoprotein excretion is connected with glycocalix and small intestinal enterocyte alteration.

  16. Rapid microbiome changes in freshly deposited cow feces under field conditions

    Directory of Open Access Journals (Sweden)

    Kelvin eWong

    2016-04-01

    Full Text Available Although development of next generation sequencing (NGS has substantially improved our understanding of the microbial ecology of animal feces, previous studies have mostly focused on freshly excreted feces. There is still limited understanding of the aging process dynamics of fecal microbiomes in intact cowpats exposed to natural environments. Fresh cowpats were sampled at multiple time points for 57 days under field conditions; half the samples were exposed to sunlight (unshaded while the other half was protected from sunlight (shaded. The 16SRNA hypervariable region 4 was amplified from each sample and sequenced on an Illumina MiSeq Platform. While Clostridia, Bacteroidia and Sphingobacteria were dominant classes of bacteria in fresh cowpats, Alphaproteobacteria, Betaproteobacteria, Actinobacteria, and Bacilli were the dominant classes by the end of the study, indicating a general shift from anaerobic to aerobic bacterial populations. This change was most likely influenced by the shift from cattle gut (anaerobic to pasture ground (aerobic. Reduced moisture in cowpats may also contribute to the community shift since air can penetrate the dryer cowpat more easily. Twelve genera consisting pathogenic bacteria were detected, with Mycobacterium, Bacillus, and Clostridium being the most abundant; their combined abundance accounts for 90% of the total pathogenic genera. Taxonomic richness and diversity increased throughout the study for most samples, which could be due to bacteria regrowth and colonization of bacteria from the environment. In contrast to the high taxonomic diversity, the changes of PICRUSt inferred function profile were minimal for all cowpats throughout the study, which suggest that core functions predicted by PICRUSt may be too conserved to distinguish differences between aerobe and anaerobe. To the best of our knowledge, this is the first study demonstrating that cowpat exposure to air and sunlight can cause drastic microbiome

  17. Hanging drop cultures of human testis and testis cancer samples

    DEFF Research Database (Denmark)

    Jørgensen, Anne; Young, J; Nielsen, J E

    2014-01-01

    limited by the lack of experimental models. The aim of this study was to establish an experimental tissue culture model to maintain normal and malignant germ cells within their niche and allow investigation of treatment effects. METHODS: Human testis and testis cancer specimens from orchidectomies were...

  18. A unique circovirus-like genome detected in pig feces

    Science.gov (United States)

    Using a metagenomic approach and molecular cloning methods, we identified, cloned, and sequenced the complete genome of a novel circular DNA virus, porcine stool-associated virus (PoSCV4), from pig feces. Phylogenetic analysis of the deduced replication initiator protein showed that PoSCV4 is most r...

  19. Characterizing healthy samples for studies of human cognitive aging

    OpenAIRE

    Geldmacher, David S.; Levin, Bonnie E.; Wright, Clinton B.

    2012-01-01

    Characterizing the cognitive declines associated with aging, and differentiating them from the effects of disease in older adults, are important goals for human neuroscience researchers. This is also an issue of public health urgency in countries with rapidly aging populations. Progress toward understanding cognitive aging is complicated by numerous factors. Researchers interested in cognitive changes in healthy older adults need to consider these complexities when they design and interpre...

  20. Identification of lactic acid bacteria in the rumen and feces of dairy cows fed total mixed ration silage to assess the survival of silage bacteria in the gut.

    Science.gov (United States)

    Han, H; Ogata, Y; Yamamoto, Y; Nagao, S; Nishino, N

    2014-09-01

    The survival of silage lactic acid bacteria (LAB) in the gut of dairy cows was evaluated by examining the LAB communities of silage and gut contents. Samples were collected at 2 different research institutes (Mie and Okayama) that offered total mixed ration (TMR) silage throughout the year. Silage and feces were sampled in August, October, and November at the Mie institute, whereas silage, rumen fluid, and feces were sampled in June and August at the Okayama institute. Denaturing gradient gel electrophoresis using Lactobacillus-specific primers was performed to detect LAB species in the samples. The selected bands were purified for species identification and the band patterns were used for principal component analysis. Lactic acid was the predominant fermentation product in all the TMR silages analyzed, and the lactic acid level tended to be constant regardless of the sampling time and region. A total of 14 LAB species were detected in the TMR silage samples, of which 5 (Lactobacillus acetotolerans, Lactobacillus pontis, Lactobacillus casei, Lactobacillus suebicus, and Lactobacillus plantarum) were detected in the dairy cow feces. Most of the denaturing gradient gel electrophoresis bands for the feces samples were also detected in the rumen fluid, suggesting that any elimination of silage LAB occurred in the rumen and not in the postruminal gut segments. The principal component analysis indicated that the LAB communities in the silage, rumen fluid, and feces were separately grouped; hence, the survival of silage LAB in the cow rumen and lower gut was deemed difficult. It was concluded that, although the gut LAB community is robust and not easily affected by the silage conditions, several LAB species can inhabit both silage and feces, which suggests the potential of using silage as a vehicle for conveying probiotics.

  1. Neospora caninum-like oocysts observed in feces of free-ranging red foxes (Vulpes vulpes) and coyotes (Canis latrans).

    Science.gov (United States)

    Wapenaar, Wendela; Jenkins, Mark C; O'Handley, Ryan M; Barkema, Herman W

    2006-12-01

    The aim of this study was to examine the feces of free-ranging foxes and coyotes for the presence of Neospora caninum oocysts. Feces were collected from 271 foxes and 185 coyotes in the Canadian province of Prince Edward Island, processed by sucrose flotation, and examined by light microscopy for the presence of coccidian oocysts. In 2 fox and 2 coyote samples, oocysts morphologically and morphometrically similar to oocysts of N. caninum were observed. DNA was extracted from these samples and subjected to nested polymerase chain reaction (PCR) using primers to the N. caninum-specific Nc5 genomic sequence. Through DNA sequencing, alignment of the sequences of at least 3 clones from each isolate to sequences deposited in GenBank revealed 95-99% similarity to the Nc5 sequence of N. caninum. PCR using primers specific for Hammondia heydorni failed to yield an amplification product from these DNA samples.

  2. Inhibition of adhesion of S-fimbriated Escherichia coli to epithelial cells by meconium feces of breast fed and formula fed newborns - mucins are the major inhibitor component

    OpenAIRE

    Schroten, H; Lethen, A.; Hanisch, F.; Plogmann, R; Hacker, Jörg; Nobis-Bosch, R; Wahn, V

    2011-01-01

    We investigated the ability of meconium, feces from human milk-fed (HMF) newborns, and feces from formula-fed (FF) newborns to inhibit adhesion of S-fimbriated E. coli to human buccal epithelial cells. S-fimbriae are a common property of E.·coli strains causing sepsis and meningitis in neonates. Meconium had the highest content of neuraminic acid and the strongest inhibitory effect on bacterial adhesion. HMF also exerted high inhibitory activity while FF was markedly less active: To achieve i...

  3. Sampling Based Trajectory Planning for Robots in Dynamic Human Environments

    DEFF Research Database (Denmark)

    Svenstrup, Mikael

    2010-01-01

    Open-ended human environments, such as pedestrian streets, hospital corridors, train stations etc., are places where robots start to emerge. Hence, being able to plan safe and natural trajectories in these dynamic environments is an important skill for future generations of robots. In this work...... method for selecting the best trajectory in the RRT, according to the cost of traversing a potential field. Furthermore the RRT expansion is enhanced to direct the search and account for the kinodynamic robot constraints. A model predictive control (MPC) approach is taken to accommodate...

  4. Sampling

    CERN Document Server

    Thompson, Steven K

    2012-01-01

    Praise for the Second Edition "This book has never had a competitor. It is the only book that takes a broad approach to sampling . . . any good personal statistics library should include a copy of this book." —Technometrics "Well-written . . . an excellent book on an important subject. Highly recommended." —Choice "An ideal reference for scientific researchers and other professionals who use sampling." —Zentralblatt Math Features new developments in the field combined with all aspects of obtaining, interpreting, and using sample data Sampling provides an up-to-date treat

  5. [Analysis of human tissue samples for volatile fire accelerants].

    Science.gov (United States)

    Treibs, Rudolf

    2014-01-01

    In police investigations of fires, the cause of a fire and the fire debris analysis regarding traces of fire accelerants are important aspects for forensic scientists. Established analytical procedures were recently applied to the remains of fire victims. When examining lung tissue samples, vapors inhaled from volatile ignitable liquids could be identified and differentiated from products of pyrolysis caused by the fire. In addition to the medico-legal results this evidence allowed to draw conclusions as to whether the fire victim was still alive when the fire started.

  6. Dansylation isotope labeling liquid chromatography mass spectrometry for parallel profiling of human urinary and fecal submetabolomes

    Energy Technology Data Exchange (ETDEWEB)

    Su, Xiaoling [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Wang, Nan [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Chen, Deying [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Li, Yunong [Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Lu, Yingfeng [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Huan, Tao [Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Xu, Wei [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Li, Liang, E-mail: Liang.Li@ualberta.ca [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2 (Canada); Li, Lanjuan, E-mail: ljli@zju.edu.cn [State Key Laboratory and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China)

    2016-01-15

    Human urine and feces can be non-invasively collected for metabolomics-based disease biomarker discovery research. Because urinary and fecal metabolomes are thought to be different, analysis of both biospecimens may generate a more comprehensive metabolomic profile that can be better related to the health state of an individual. Herein we describe a method of using differential chemical isotope labeling (CIL) liquid chromatography mass spectrometry (LC-MS) for parallel metabolomic profiling of urine and feces. Dansylation labeling was used to quantify the amine/phenol submetabolome changes among different samples based on {sup 12}C-labeling of individual samples and {sup 13}C-labeling of a pooled urine or pooled feces and subsequent analysis of the {sup 13}C-/{sup 12}C-labeled mixture by LC-MS. The pooled urine and pooled feces are further differentially labeled, mixed and then analyzed by LC-MS in order to relate the metabolite concentrations of the common metabolites found in both biospecimens. This method offers a means of direct comparison of urinary and fecal submetabolomes. We evaluated the analytical performance and demonstrated the utility of this method in the analysis of urine and feces collected daily from three healthy individuals for 7 days. On average, 2534 ± 113 (n = 126) peak pairs or metabolites could be detected from a urine sample, while 2507 ± 77 (n = 63) peak pairs were detected from a fecal sample. In total, 5372 unique peak pairs were detected from all the samples combined; 3089 and 3012 pairs were found in urine and feces, respectively. These results reveal that the urine and fecal metabolomes are very different, thereby justifying the consideration of using both biospecimens to increase the probability of finding specific biomarkers of diseases. Furthermore, the CIL LC-MS method described can be used to perform parallel quantitative analysis of urine and feces, resulting in more complete coverage of the human metabolome

  7. Human papillomavirus self-sampling for screening nonattenders

    DEFF Research Database (Denmark)

    Lam, Janni Uyen Hoa; Rebolj, Matejka; Ejegod, Ditte Møller

    2017-01-01

    In organized cervical screening programs, typically 25% of the invited women do not attend. The Copenhagen Self-sampling Initiative (CSi) aimed to gain experiences on participation among screening nonattenders in the Capital Region of Denmark. Here, we report on the effectiveness of different...... region of Denmark were identified via the organized national invitation module. Screening history was obtained via the nationwide pathology registry. Twenty-four thousand women were invited, and as an alternative to the regular communication platforms (letter and phone), women could request a home test...... via a mobile-friendly webpage. Instruction material and video-animation in several languages were made available online. Chi-square test was used to test differences. Out of all invited, 31.7% requested a home test, and 20% returned it to the laboratory. In addition, 10% were screened at the physician...

  8. Improved culture methods for isolation of Salmonella organisms from swine feces

    DEFF Research Database (Denmark)

    Hoorfar, Jeffrey; Mortensen, Alicja

    2000-01-01

    . Procedure-4 experiments were performed to evaluate the following: 1) diagnostic sensitivity of the selective preenrichment and rapid isolation novel technology (SPRINT) protocol, compared with that of the modified ISO protocol; 2) detection limit of the SPRINT protocol for Salmonella organisms; 3) use...... of preenrichment of samples between the use of UPE broth or BPW. Conclusions and Clinical Relevance-The SPRINT protocol may provide a faster alternative for isolation of Salmonella organisms from swine fecal samples. Furthermore, the use of TTN broth instead of SC broth may increase the sensitivity of the modified......Objective-To compare 3 alternative culture techniques for the detection of Salmonella organisms in swine feces with a modification of the international Standard Organization (ISO) 6579 standard protocol. Sample Population-Fecal samples from swine herds suspected of having Salmonella infections...

  9. Reducing sample complexity of polyclonal human autoantibodies by chromatofocusing.

    Science.gov (United States)

    Hagemann, Sascha; Faude, Alexander; Rabenstein, Monika; Balzer-Geldsetzer, Monika; Nölker, Carmen; Bacher, Michael; Dodel, Richard

    2010-08-15

    Chromatofocusing was performed in order to separate a polyclonal antigen-specific mixture of human immunoglobulins (IgGs) that would then allow for further analyses of as few different IgGs as possible. Because polyclonal IgGs only differ by amino acid sequence and possible post-translational modifications but not by molecular weight, we chose chromatofocusing for protein separation by different isoelectric points. We isolated antigen-specific IgGs from commercially available intravenous immunoglobulins (IVIG) using a combination of affinity- and size exclusion-chromatography and in order to reduce the complexity of the starting material IVIG was then replaced by single-donor plasmapheresis material. Using two-dimensional gel electrophoresis (2-DE), we observed a clear decrease in the number of different light and heavy chains in the chromatofocusing peak as compared to the starting material. In parallel, we monitored slight problems with the selected peak in isoelectric focusing as the first dimension of 2-DE, displayed in by the less proper focusing of the spots. When we tested whether IgGs were binding to their specific antigen after chromatofocusing, we were able to show that they were still in native conformation. In conclusion, we showed that chromatofocusing can be used as a first step in the analysis of mixtures of very similar proteins, e.g. polyclonal IgG preparations, in order to minimize the amount of different proteins in separated fractions in a reproducible way. Copyright 2010 Elsevier B.V. All rights reserved.

  10. Assessment of human exposure to airborne fungi in agricultural confinements: personal inhalable sampling versus stationary sampling.

    Science.gov (United States)

    Adhikari, Atin; Reponen, Tiina; Lee, Shu-An; Grinshpun, Sergey A

    2004-01-01

    Accurate exposure assessment to airborne fungi in agricultural environments is essential for estimating the associated occupational health hazards of workers. The objective of this pilot study was to compare personal and stationary sampling for assessing farmers' exposure to airborne fungi in 3 different agricultural confinements located in Ohio, USA (hog farm, dairy farm, and grain farm), using Button Personal Inhalable Samplers. Personal exposures were measured with samplers worn by 3 subjects (each carrying 2 samplers) during 3 types of activities, including animal feeding in the hog farm, cleaning and animal handling in the dairy farm, and soybean unloading and handling in the grain farm. Simultaneously, the stationary measurements were performed using 5 static Button Samplers and 1 revolving Button Sampler. The study showed that the total concentration of airborne fungi ranged from 1.4 x 10(4)-1.2 x 10(5) spores m(-3) in 3 confinements. Grain unloading and handling activity generated highest concentrations of airborne fungi compared to the other 2 activities. Prevalent airborne fungi belonged to Cladosporium, Aspergillus/Penicillium, Ascospores, smut spores, Epicoccum, Alternaria, and Basidiospores. Lower coefficients of variations were observed for the fungal concentrations measured by personal samplers (7-12%) compared to the concentrations measured by stationary samplers (27-37%). No statistically significant difference was observed between the stationary and personal measurement data for the total concentrations of airborne fungi (p > 0.05). Revolving stationary and static stationary Button Samplers demonstrated similar performance characteristics for the collection of airborne fungi. This reflects the low sensitivity of the sampler's efficiency to the wind speed and direction. The results indicate that personal exposure of agricultural workers in confinements may be adequately assessed by placing several Button Samplers simultaneously operating in a

  11. Detection of novel polyomaviruses, TSPyV, HPyV6, HPyV7, HPyV9 and MWPyV in feces, urine, blood, respiratory swabs and cerebrospinal fluid.

    Directory of Open Access Journals (Sweden)

    Rebecca J Rockett

    Full Text Available Eight novel human polyomaviruses have been discovered since 2007. Prevalence rates and tissue tropism for the most recent members HPyV 6, 7, 9, TSPyV and MWPyV are largely unknown. We used real-time PCR to determine the presence of HPyV 6, 7, 9, TSPyV and MWPyV in feces (n = 263, urine (n = 189, blood (n = 161, respiratory swabs (n = 1385 and cerebrospinal fluid (n = 171 from both healthy control children and children and adults undergoing diagnostic testing. Whole genome sequencing was able to be performed on 9 MWPyV positive specimens. Novel polyomaviruses were only detected in respiratory swabs and feces, with no detections of HPyV 9 in any sample type. MWPyV was found to be the most prevalent novel polyomavirus, being detected in 18 (1.5% respiratory specimens from symptomatic patients, 16 (9.8% respiratory sample from healthy control children, 11 (5.9% fecal specimens from patient suffering gastrointestinal illness, and in 13 (15.3% of feces from healthy control children. MWPyV was found only in respiratory and fecal specimens from children, the oldest being 9 years old. HPyV 6, 7, 9 and TSPyV were also detected in respiratory specimens and fecal specimens at low prevalence (<1.3%. The majority of these detections were found in immunocompromised patients. Our findings suggest that MWPyV can result in a subclinical infection, persistent or intermittent shedding, particularly in young children. The other novel polyomaviruses were also found in respiratory and fecal specimens, but at lower prevalence and most commonly in immunocompromised individuals.

  12. Terrestrial mammal feces: a morphometric summary and description

    Directory of Open Access Journals (Sweden)

    Marcia Chame

    2003-01-01

    Full Text Available The study of feces of terrestrial mammals brings out biological and ecological data such as the species presence, diet, behaviour, territory, parasitic fauna, and home-range use, which can be applied for conservation projects and support paleoecological research that use coprolites as the main source of study. Although the new biotechnological techniques allow more accurate data, the diagnosis based on morphometric analyses permits the primary identification of the taxonomic group origin to support the best choice of subsequent analyses. We present the compilation list of fecal shape and measurements available in the literature published in North America, Eastern and Southern Africa, Europe, and new data from Brazil. Shape and diameters are the best characteristics for taxonomic identification. Feces were assembled in 9 groups that reflect the Order, sometimes the Family, and even their common origin.

  13. Microbial and Oligosaccharides Treatments of Feces and Slurry in Reducing Ammonia of the Poultry Farm

    Directory of Open Access Journals (Sweden)

    Y. Yusrizal

    2012-12-01

    Full Text Available This study was conducted to investigate the effectiveness of Lactobacillus sp and fructooligosaccaride (FOS to reduce the volatile ammonia formation from chicken excreta and layer slurry. For each treatment-replication, 150 g of fecal material were collected from the poultry farm and placed in 500 ml beaker glass. The fecal sample was then treated with 2% Lactobacillus sp (2.6x106 cfu/g and 2% FOS and covered with plastic wraps. The volatile ammonia contents and pH were measured after one hour of standing (0 d and repeated at 48 h intervals for 6 d. For the dropping slurry study, 300 g of each layer dropping slurry sample were used. Results indicated that 2% Lactobacillus sp or FOS supplementations in the feces and dropping slurry after 1 h up to 6 d reduced the ammonia odor formation, fecal pH, and moisture content. The Lactobacillus sp and FOS treated manure resulted in increasing Lactobacillus sp count and reducing in E. coli, Salmonella, and Campylobacter in 6 days for both feces and layer dropping slurry. In addition, reducing moisture content was observed in treated manure. It is concluded that Lactobacillus sp and FOS reduced the volatile ammonia formation and pathogenic bacteria from chicken excreta and layer slurry.

  14. Tannic acid degradation by Klebsiella strains isolated from goat feces

    Directory of Open Access Journals (Sweden)

    Arezoo Tahmourespour

    2016-03-01

    Full Text Available Background and Objectives: Tannins are toxic polyphenols that either bind and precipitate or condense proteins. The high tannin content of some plants is the preliminary limitation of using them as a ruminant feed. So, the aim of this study was the isolation and characterization of tannic acid degrading bacterial strains from goat feces before and after feeding on Pis- tachio-Soft Hulls as tannin rich diet (TRD.Materials and Methods: Bacterial strains capable of utilizing tannic acid as sole carbon and energy source were isolated and characterized from goat feces before and after feeding on TRD. Tannase activity, maximum tolerable concentration and biodegradation potential were assessed.Results: Four tannase positive isolates were identified as Klebsiella pneumoniae. Isolated strains showed the maximum tolerable concentration of 64g/L of tannin. The tannic acid degradation percentage at a concentration of 15.0 g/L reached a maximum of 68% after 24 h incubation, and more than 98% after 72 h incubation. The pH of the medium also decreased along with tannic acid utilization.Conclusions: It is obvious that TRD induced adaptive responses. Thus, while the bacteria were able to degrade and detoxify the tannic acids, they had to adapt in the presence of high concentrations of tannic acid. So, these isolates have an amazing potential for application in bioremediation, waste water treatment, also reduction of tannins antinutritional effects in animal feeds.Keywords: Biodegradation; Goat feces; Klebsiella strains; Tannic acid

  15. Optimization of techniques for multiple platform testing in small, precious samples such as human chorionic villus sampling.

    Science.gov (United States)

    Pisarska, Margareta D; Akhlaghpour, Marzieh; Lee, Bora; Barlow, Gillian M; Xu, Ning; Wang, Erica T; Mackey, Aaron J; Farber, Charles R; Rich, Stephen S; Rotter, Jerome I; Chen, Yii-der I; Goodarzi, Mark O; Guller, Seth; Williams, John

    2016-11-01

    Multiple testing to understand global changes in gene expression based on genetic and epigenetic modifications is evolving. Chorionic villi, obtained for prenatal testing, is limited, but can be used to understand ongoing human pregnancies. However, optimal storage, processing and utilization of CVS for multiple platform testing have not been established. Leftover CVS samples were flash-frozen or preserved in RNAlater. Modifications to standard isolation kits were performed to isolate quality DNA and RNA from samples as small as 2-5 mg. RNAlater samples had significantly higher RNA yields and quality and were successfully used in microarray and RNA-sequencing (RNA-seq). RNA-seq libraries generated using 200 versus 800-ng RNA showed similar biological coefficients of variation. RNAlater samples had lower DNA yields and quality, which improved by heating the elution buffer to 70 °C. Purification of DNA was not necessary for bisulfite-conversion and genome-wide methylation profiling. CVS cells were propagated and continue to express genes found in freshly isolated chorionic villi. CVS samples preserved in RNAlater are superior. Our optimized techniques provide specimens for genetic, epigenetic and gene expression studies from a single small sample which can be used to develop diagnostics and treatments using a systems biology approach in the prenatal period. © 2016 John Wiley & Sons, Ltd. © 2016 John Wiley & Sons, Ltd.

  16. Sex determination of sika deer (Cervus nippon yesoensis) using nested PCR from feces collected in the field.

    Science.gov (United States)

    Yamazaki, Shoki; Motoi, Yuta; Nagai, Kazuya; Ishinazaka, Tsuyoshi; Asano, Makoto; Suzuki, Masatsugu

    2011-12-01

    We describe a method for determining the sex of sika deer (Cervus nippon yesoensis) from feces collected in the field. Using a nested polymerase chain reaction (nested PCR), partial sequences of the sex determination region of the Y chromosome (SRY) gene and X zinc finger protein (ZFX) gene were amplified. In 19 individuals with sex information, the correct sex was successfully detected and sequences of target amplicons were completely matched between muscle and feces from the rectum. Among 75 fecal samples collected noninvasively in the field, 68-71 samples (90.7-94.7%) yielded successful sex determinations. Using this technique, feces collected in the field would enhance the utility of genetic analysis. As a result, instead of biomaterials, these samples can serve as new or alternative materials. Finally, it can be expected that this technique will contribute to reveal in advanced detail of the population dynamics and genetic diversity that needed to carry out effective population control and to reduce the extinction risk of sika deer.

  17. Application of a real-time fluorescence resonance energy transfer polymerase chain reaction assay with melting curve analysis for the detection of Paragonimus heterotremus eggs in the feces of experimentally infected cats.

    Science.gov (United States)

    Tantrawatpan, Chairat; Intapan, Pewpan M; Thanchomnang, Tongjit; Sanpool, Oranuch; Janwan, Penchom; Lulitanond, Viraphong; Anamnart, Witthaya; Maleewong, Wanchai

    2013-09-01

    Paragonimus heterotremus is a medically important lung fluke that causes human and animal paragonimiasis in Southeast Asia, including Thailand. In the current study, a real-time fluorescence resonance energy transfer polymerase chain reaction (real-time FRET PCR) with melting curve analysis was developed and evaluated to detect P. heterotremus eggs in the feces of experimentally infected cats. The detection limit of this method for the P. heterotremus DNA sequence was 3 × 10(2) copies of the positive control plasmid and 10(-3) ng of P. heterotremus genomic DNA. The assay system could detect 10 eggs of P. heterotremus per gram of cat feces. No fluorescence signal was observed when DNA purified from 16 other organisms or genomic DNA from cats and human beings were tested. Real-time FRET PCR yielded positive results for all fecal samples from 17 P. heterotremus-infected cats and showed a negative relationship (r = -0.852, P analysis. This assay can be useful for the detection of, and epidemiological studies on, P. heterotremus infection in endemic areas.

  18. Human DNA quantification and sample quality assessment: Developmental validation of the PowerQuant(®) system.

    Science.gov (United States)

    Ewing, Margaret M; Thompson, Jonelle M; McLaren, Robert S; Purpero, Vincent M; Thomas, Kelli J; Dobrowski, Patricia A; DeGroot, Gretchen A; Romsos, Erica L; Storts, Douglas R

    2016-07-01

    Quantification of the total amount of human DNA isolated from a forensic evidence item is crucial for DNA normalization prior to short tandem repeat (STR) DNA analysis and a federal quality assurance standard requirement. Previous commercial quantification methods determine the total human DNA and total human male DNA concentrations, but provide limited information about the condition of the DNA sample. The PowerQuant(®) System includes targets for quantification of total human and total human male DNA as well as targets for evaluating whether the human DNA is degraded and/or PCR inhibitors are present in the sample. A developmental validation of the PowerQuant(®) System was completed, following SWGDAM Validation Guidelines, to evaluate the assay's specificity, sensitivity, precision and accuracy, as well as the ability to detect degraded DNA or PCR inhibitors. In addition to the total human DNA and total human male DNA concentrations in a sample, data from the degradation target and internal PCR control (IPC) provide a forensic DNA analyst meaningful information about the quality of the isolated human DNA and the presence of PCR inhibitors in the sample that can be used to determine the most effective workflow and assist downstream interpretation.

  19. Towards diagnostic metagenomics of Campylobacter in fecal samples

    DEFF Research Database (Denmark)

    Andersen, Sandra Christine; Kiil, Kristoffer; Harder, Christoffer Bugge

    2017-01-01

    of the challenges in diagnostic metagenomics are, that it requires a great next-generation sequencing depth and unautomated data analysis. DNA from human fecal samples spiked with 7.75 × 101-7.75 × 107 colony forming unit (CFU)/ml Campylobacter jejuni and chicken fecal samples spiked with 1 × 102-1 × 106 CFU....../g Campylobacter jejuni was sequenced and data analysis was done by the metagenomic tools Kraken and CLARK. More hits were obtained at higher spiking levels, however with no significant linear correlations (human samples p = 0.12, chicken samples p = 0.10). Therefore, no definite detection limit could...... be determined, but the lowest spiking levels found positive were 7.75 × 104 CFU/ml in human feces and 103 CFU/g in chicken feces. Eight human clinical fecal samples with estimated Campylobacter infection loads from 9.2 × 104-1.0 × 109 CFU/ml were analyzed using the same methods. It was possible to detect...

  20. Urea in Weaver Ant Feces: Quantification and Investigation of the Uptake and Translocation of Urea in Coffea Arabica

    DEFF Research Database (Denmark)

    Vidkjær, Nanna Hjort; Wollenweber, Bernd; Jensen, Karl-Martin Vagn

    2016-01-01

    investigate the interactions of weaver ants with the host plants with respect to plant nutrition. Here, we report the identification and quantification of urea, a highly effective foliar nutrient present in the fecal depositions of O. smaragdina. Feces samples obtained from six O. smaragdina colonies were......Weaver ants are tropical insects that nest in tree canopies, and for centuries these ants have been used for pest control in tropical orchards. Trees hosting weaver ants might benefit not only from the pest protective properties of these insects but also an additional supply of nutrients from ant...... feces deposited on the leaves. In a recent study, we demonstrated that Coffea arabica plants hosting Oecophylla smaragdina weaver ants under laboratory conditions experienced enhanced nitrogen availability compared with plants grown without ants. Therefore, the aim of the present study was to further...

  1. Adenoviruses of canine and human origins in stool samples from free-living pampas foxes (Lycalopex gymnocercus) and crab-eating foxes (Cerdocyon thous) in São Francisco de Paula, Rio dos Sinos basin.

    Science.gov (United States)

    Monteiro, G S; Fleck, J D; Kluge, M; Rech, N K; Soliman, M C; Staggemeier, R; Rodrigues, M T; Barros, M P; Heinzelmann, L S; Spilki, F R

    2015-05-01

    The spread of enteric viruses of domestic animals and human beings to wild species can be facilitated by the resistance of these viruses on the environment and their ability to be transmitted by water and contaminated food. The health status of the populations of pampas foxes Lycalopex gymnocercus) and crab-eating foxes (Cerdocyon thous) is largely unknown and the landscapes occupied by these animals in southern Brazil have been threatened by human occupation and expansion of agriculture. In this work, the search of genomes of human and canine adenoviruses in feces from these wild carnivores was used to track the dissemination of domestic animals and human pathogens to the free-living populations in a wildlife reserve located in southern Brazil. This was performed by virus-specific differential real-time polymerase chain reactions (qPCR) on stool specimens, avoiding capture and additional stress to the animals. Genus-specific conventional reverse-transcriptase PCR (RT-PCR) was complementarily performed aiming the detection of enteroviruses (EV) and rotaviruses (RV) on these same samples. HAdV genomes were found on 14 out of the 17 (82.35%) stool samples analysed, whereas CAV was found co-infecting 5 of these samples. RV genomes were detected on 7 of the 17 samples (41.18%) and all samples were negative for EV. The results point to the dispersion of HAdV and RV at a high rate to these species of South American wild carnivores, which can be an effect of growing anthropisation of the habitat of these animals.

  2. The Characterization of Feces and Urine: A Review of the Literature to Inform Advanced Treatment Technology.

    Science.gov (United States)

    Rose, C; Parker, A; Jefferson, B; Cartmell, E

    2015-09-02

    The safe disposal of human excreta is of paramount importance for the health and welfare of populations living in low income countries as well as the prevention of pollution to the surrounding environment. On-site sanitation (OSS) systems are the most numerous means of treating excreta in low income countries, these facilities aim at treating human waste at source and can provide a hygienic and affordable method of waste disposal. However, current OSS systems need improvement and require further research and development. Development of OSS facilities that treat excreta at, or close to, its source require knowledge of the waste stream entering the system. Data regarding the generation rate and the chemical and physical composition of fresh feces and urine was collected from the medical literature as well as the treatability sector. The data were summarized and statistical analysis was used to quantify the major factors that were a significant cause of variability. The impact of this data on biological processes, thermal processes, physical separators, and chemical processes was then assessed. Results showed that the median fecal wet mass production was 128 g/cap/day, with a median dry mass of 29 g/cap/day. Fecal output in healthy individuals was 1.20 defecations per 24 hr period and the main factor affecting fecal mass was the fiber intake of the population. Fecal wet mass values were increased by a factor of 2 in low income countries (high fiber intakes) in comparison to values found in high income countries (low fiber intakes). Feces had a median pH of 6.64 and were composed of 74.6% water. Bacterial biomass is the major component (25-54% of dry solids) of the organic fraction of the feces. Undigested carbohydrate, fiber, protein, and fat comprise the remainder and the amounts depend on diet and diarrhea prevalence in the population. The inorganic component of the feces is primarily undigested dietary elements that also depend on dietary supply. Median urine

  3. Bacteria-human somatic cell lateral gene transfer is enriched in cancer samples.

    Directory of Open Access Journals (Sweden)

    David R Riley

    Full Text Available There are 10× more bacterial cells in our bodies from the microbiome than human cells. Viral DNA is known to integrate in the human genome, but the integration of bacterial DNA has not been described. Using publicly available sequence data from the human genome project, the 1000 Genomes Project, and The Cancer Genome Atlas (TCGA, we examined bacterial DNA integration into the human somatic genome. Here we present evidence that bacterial DNA integrates into the human somatic genome through an RNA intermediate, and that such integrations are detected more frequently in (a tumors than normal samples, (b RNA than DNA samples, and (c the mitochondrial genome than the nuclear genome. Hundreds of thousands of paired reads support random integration of Acinetobacter-like DNA in the human mitochondrial genome in acute myeloid leukemia samples. Numerous read pairs across multiple stomach adenocarcinoma samples support specific integration of Pseudomonas-like DNA in the 5'-UTR and 3'-UTR of four proto-oncogenes that are up-regulated in their transcription, consistent with conversion to an oncogene. These data support our hypothesis that bacterial integrations occur in the human somatic genome and may play a role in carcinogenesis. We anticipate that the application of our approach to additional cancer genome projects will lead to the more frequent detection of bacterial DNA integrations in tumors that are in close proximity to the human microbiome.

  4. OCCURRENCE OF VANCOMYCIN RESISTANT ENTEROCOCCI IN ANIMAL FECES

    Science.gov (United States)

    A survey was conducted to determine the occurrence of vancomycin resistant Enterococci (VRE) in animal and human fecal samples. A selective agar mEI, and mEI supplemented with 4 micrograms/ml vancomycin was used in a membrane filtration procedure to determine quantitative levels ...

  5. Mitochondrial Respiration Chain Enzymatic Activities in the Human Brain: Methodological Implications for Tissue Sampling and Storage.

    Science.gov (United States)

    Ronsoni, Marcelo Fernando; Remor, Aline Pertile; Lopes, Mark William; Hohl, Alexandre; Troncoso, Iris H Z; Leal, Rodrigo Bainy; Boos, Gustavo Luchi; Kondageski, Charles; Nunes, Jean Costa; Linhares, Marcelo Neves; Lin, Kátia; Latini, Alexandra Susana; Walz, Roger

    2016-04-01

    Mitochondrial respiratory chain complexes enzymatic (MRCCE) activities were successfully evaluated in frozen brain samples. Epilepsy surgery offers an ethical opportunity to study human brain tissue surgically removed to treat drug resistant epilepsies. Epilepsy surgeries are done with hemodynamic and laboratory parameters to maintain physiology, but there are no studies analyzing the association among these parameters and MRCCE activities in the human brain tissue. We determined the intra-operative parameters independently associated with MRCCE activities in middle temporal neocortex (Cx), amygdala (AMY) and head of hippocampus (HIP) samples of patients (n = 23) who underwent temporal lobectomy using multiple linear regressions. MRCCE activities in Cx, AMY and HIP are differentially associated to trans-operative mean arterial blood pressure, O2 saturation, hemoglobin, and anesthesia duration to time of tissue sampling. The time-course between the last seizure occurrence and tissue sampling as well as the sample storage to biochemical assessments were also associated with enzyme activities. Linear regression models including these variables explain 13-17 % of MRCCE activities and show a moderate to strong effect (r = 0.37-0.82). Intraoperative hemodynamic and laboratory parameters as well as the time from last seizure to tissue sampling and storage time are associated with MRCCE activities in human samples from the Cx, AMYG and HIP. Careful control of these parameters is required to minimize confounding biases in studies using human brain samples collected from elective neurosurgery.

  6. A duplex PCR for rapid and simultaneous detection of Brucella spp. in human blood samples.

    Science.gov (United States)

    Mirnejad, Reza; Mohamadi, Mozafar; Piranfar, Vahbeh; Mortazavi, Seied Mojtaba; Kachuei, Reza

    2013-06-01

    To design a duplex PCR for rapid and simultaneous detection of Brucella species. in human blood samples. Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis. Following DNA extraction, PCR assay were performed, using three primers that could simultaneously identify and differentiate three major species of pathogenic Brucella in humans and animals. Of the 52 peripheral bloods samples tested, 25 sample (48%) showed positive reactions in PCR. Twelve samples were positive for Brucella abortus 39 (B. abortus 39) (23%), 13 for Brucella melitensis 39 (B. melitensis 39) (25%) and 0 for Brucella ovis 39 (B. ovis 39) (0%). This work demonstrates that in case where specific primers were utilized, duplex PCR has proved to be a simple, fast, and relatively inexpensive method for simultaneous detection of important species of Brucella in clinical samples. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  7. A duplex PCR for the rapid and simultaneous detection of Brucella spp. in human blood samples

    Institute of Scientific and Technical Information of China (English)

    Reza Mirnejad; Mozafar mohamadi; Vahbeh Piranfar; Seied Mojtaba Mortazavi; Reza Kachuei

    2013-01-01

    Objective: To design a duplex PCR for rapid and simultaneous detection of Brucella species. in human blood samples. Methods: Fifty-two peripheral bloods samples were collected from suspicious patients with brucellosis. Following DNA extraction, PCR assay were performed, using three primers that could simultaneously identify and differentiate three major species of pathogenic Brucella in humans and animals. Results: Of the 52 peripheral bloods samples tested, 25 sample (48%) showed positive reactions in PCR. Twelve samples were positive for Brucella abortus (B. abortus) (23%), 13 for Brucella melitensis (B. melitensis) (25%) and 0 for Brucella ovis (B. ovis) (0%). Conclusions: This work de=monstrates that in case where specific primers were utilized, duplex PCR has proved to be a simple, fast, and relatively inexpensive method for simultaneous detection of important species of Brucella in clinical samples.

  8. Mini-FLOTAC for counting Toxoplasma gondii oocysts from cat feces--comparison with cell counting plates.

    Science.gov (United States)

    Djokic, Vitomir; Blaga, Radu; Rinaldi, Laura; Le Roux, Delphine; Ducry, Tamara; Maurelli, Maria Paola; Perret, Catherine; Djurkovic Djakovic, Olgica; Cringoli, Giuseppe; Boireau, Pascal

    2014-12-01

    Oocysts of Toxoplasma gondii represent one of the most common environmental contaminants causing the zoonotic infection toxoplasmosis. The aim of the present study was to compare the Mini-FLOTAC device with traditional cell counting plates (Kova Slide) for the detection of T. gondii oocysts from feline feces. Two types of experiments were performed: (i) purified oocysts were counted in different dilutions and (ii) specific pathogen free T. gondii-negative cat feces was inoculated with numbers of purified oocysts and counting was performed directly from feces. Our analysis showed a thousand times higher sensitivity of Mini-FLOTAC (5 × 10(2) oocysts) compared to Kova Slide (5 × 10(5) oocysts). Also, when compared by McNemar's test, counting of the purified oocysts showed a higher sensitivity of Mini-FLOTAC compared to Kova Slide, for a dilution of 10(3) oocysts/ml (chi(2) = 6.1; P microscopes in any laboratory or field conditions. We therefore recommend its use for regular screening. Further studies are needed to validate Mini-FLOTAC for the detection of oocysts in soil and water samples in field conditions.

  9. Animal Feces Contribute to Domestic Fecal Contamination: Evidence from E. coli Measured in Water, Hands, Food, Flies, and Soil in Bangladesh

    Science.gov (United States)

    2017-01-01

    Fecal-oral pathogens are transmitted through complex, environmentally mediated pathways. Sanitation interventions that isolate human feces from the environment may reduce transmission but have shown limited impact on environmental contamination. We conducted a study in rural Bangladesh to (1) quantify domestic fecal contamination in settings with high on-site sanitation coverage; (2) determine how domestic animals affect fecal contamination; and (3) assess how each environmental pathway affects others. We collected water, hand rinse, food, soil, and fly samples from 608 households. We analyzed samples with IDEXX Quantitray for the most probable number (MPN) of E. coli. We detected E. coli in source water (25%), stored water (77%), child hands (43%), food (58%), flies (50%), ponds (97%), and soil (95%). Soil had >120 000 mean MPN E. coli per gram. In compounds with vs without animals, E. coli was higher by 0.54 log10 in soil, 0.40 log10 in stored water and 0.61 log10 in food (p contamination, and fecal indicator bacteria do not strictly indicate human fecal contamination when animals are present. PMID:28686435

  10. Variation in the extraction efficiency of estradiol and progesterone in moist and lyiphilized feces of the black howler monkey (Alouatta pigra: alternative methods

    Directory of Open Access Journals (Sweden)

    Vianey Del Rocio Torres

    2011-12-01

    Full Text Available several fecal steroid extraction techniques have been developed to measure the ovary function in different species of mammals. However, regardless of the method of extraction and the sample type chosen, is has been observed that they can yield results with different percentages of recuperation. The objective of this study was to determine whether the type of substratum, solvent and extraction method used have any influence on the extraction efficiency in the feces of Alouatta pigra (black howler monkey. For this purpose we used two methods: agitation and ebullition. With each method, we utilized moist and lyophilized feces. The validation of radioimmunoassay method was accurate and precise for quantify estradiol and progesterone in lyophilized feces of Alouatta pigra. To both of which ethanol and methanol, absolute and at 80%, were added, besides the hormones 125I-Estradiol and 125I-Progesterone. The extraction efficiency for 125I-Estradiol was from 87.72 ± 3.97% to 41.24 ± 2.67%, and for 125I-Progesterone from 71.15 ± 4.24% to 42.30 ± 1.19% when we used the agitation method. Whereas with the ebullition method, the extraction efficiency for 125I-Estradiol ranged from 86.89 ± 2.66% to 71.68 ± 3.02% and for 125I-Progesterone from 98.31 ± 1.26% to 85.40 ± 1.98%. Due to the differences found in these assays, which depend on the method used, the type of feces employed and the type of solvent added to them, we recommend the ebullition method and the lyophilized feces of Alouatta pigra for extracting the hormones, since in moist feces there may exist variables which might interfere in the quantification of 125I-Estradiol and 125I-Progesterone.

  11. Genotyping of cystic echinococcosis isolates from clinical samples of human and domestic animals

    Directory of Open Access Journals (Sweden)

    S.A. Fadhil

    2016-12-01

    Full Text Available Cystic hydatid disease is a cosmopolitan important disease in both human and animals. Many strains were investigated in this parasite. The aim of study was to characterize genotype variations of Echinococcus granulosus isolates collected from human and domestic animals in Al-Qadisiyah province/ Iraq based on sequencing of nad1 mitochondrial gene. Eighty hydatid cysts of human (12, sheep (15, cattle (36, and camels (17 were collected from hospital and slaughter house of the province, during October 2014 to June 2015; microscopic examination was made for cysts fluid to determine the fertility. DNAs extraction was done for each sample in addition to purify and concentrate of extracted DNA samples was performed to determine nad1 (400bp gene used conventional PCR method. Phylogenetic analysis was performed using NCBI-Blast Alignment identification and Unweighted Pair Group Method with Arithmetic Mean. Twenty five (10 from human and 5 from each studied animals samples were chosen due to their fertility and high DNA purity, in which three strains (genotypes were investigated including sheep strain (G1 40%, buffalo strain (G3 48% and camel strain (G6 12%, where human samples related to G1(20% and G3(80%; sheep samples related to G1(80% and G3(20%; cattle samples related to G1(60%, G3 (20% and G6 (20%; camels samples related to G1(20%, G3(40% and G6(40%. The dominant strain is a buffalo strain (G3; both of buffalo strain (G3 and sheep strain (G1 represented the actual source of human infection. There is no host specificity of detected genotypes.

  12. Prevalence of Campylobacter, Arcobacter, Helicobacter, and Sutterella spp. in human fecal samples as estimated by a reevaluation of isolation methods for Campylobacters

    DEFF Research Database (Denmark)

    Engberg, J.; On, Stephen L.W.; Harrington, C.S.;

    2000-01-01

    CCDA recovered significantly more thermophilic Campylobacter spp. than Skirrow's medium (P = 0.0034). No significant difference between Skirrow's medium and CAT agar was observed in this study. Another six taxa were identified, namely, Campylobacter concisus, Campylobacter curvus-like bacteria, Arcobacter...... butzleri, Arcobacter cryaerophilus, Helicobacter cinaedi, and Sutterella wadsworthensis. Most of these strains were isolated after 5 to 6 days of incubation by use of the filter technique. This paper pro,ides evidence for the existence of S. wadsworthensis in human feces from clinical cases...

  13. Lawsonia intracellularis in the feces of wild rodents and stray cats captured around equine farms.

    Science.gov (United States)

    Hwang, Jeong-Min; Seo, Myung-Ji; Yeh, Jung-Yong

    2017-08-11

    Proliferative enteropathy is a global enteric disease of particular importance in pigs. The causative bacterium, Lawsonia intracellularis, has a wide range of susceptible host species. Recently, L. intracellularis has been recognized as an etiologic agent of an emerging enteric disease in foals called equine proliferative enteropathy (EPE). The presence of L. intracellularis in nonruminant wildlife has raised questions regarding the role of these species in EPE transmission. This study investigated exposure to L. intracellularis in wild rodents and feral cats from eight farms with confirmed EPE. Serum (42) and fecal (40) samples from resident foals and fecal samples (131), intestinal mucosa tissues (14), and mesenteric lymph nodes (14) from wild and feral animals were collected for the evaluation of the farm status and the molecular detection of L. intracellularis following the diagnosis of EPE in index cases. Fresh feces from wild rodents and feral cats were collected from the ground while walking the premises or after trapping the animals using live traps. A total of 3 brown rats, 7 house mice, 1 striped field mouse, 2 grey red-backed voles, and 3 feral cats showed evidence of prior exposure to L. intracellularis. Our data add to increasing evidence demonstrating the potential for L. intracellularis transmission and infection in wild rodents and feral cats and provide possible evidence of interspecies transmission. The exposure of wild rodents and feral cats provides potential evidence for the spillover of L. intracellularis to wildlife species and raises the question of spillback to horses. Additionally, these animals may represent an indicator of environmental exposure or may be actively involved in the transmission of L. intracellularis to foals by acting as potential reservoir/amplifier hosts. This study is the first to demonstrate the magnitude of L. intracellularis shedding in the feces of wild rodents and feral cats and to indicate the significant

  14. Method development and validation: solid Phase extraction-ultra performance liquid chromatography-tandem mass spectrometry quantification of pirlimycin in bovine feces and urine.

    Science.gov (United States)

    Ray, Partha; Knowlton, Katharine F; Shang, Chao; Xia, Kang

    2014-01-01

    Pirlimycin, a lincosamide antibiotic, is one of the most commonly used antibiotics for the treatment of mastitis in dairy cows. Assessment of pirlimycin loadingto the environment via fecal and urinary excretion is critical to develop efficient management strategies to reduce environmental pollution by the livestock industry. Therefore, the aim of this study was to develop and validate an analytical method to identify and quantify pirlimycin in bovine feces and urine. Samples were extracted with methanol- phosphate buffer and cleaned up by SPE before analysis for pirlimycin using UPLC-MS/MS. This method was sensitive (LOQ 1.47 ng/g wet feces, 0.90 ng/mL urine), accurate (recovery, 80-108%), and precise (repeatability, 2.3-13%; reproducibility, 2.3-14%) for both bovine feces and urine. With the application of this method to samples collected in the first 10 h and then every 24 h for 120 h following intramammary dosing (50 mg/cow; n = 3 cows), pirlimycin was detected at 40.5-287 ng/g and 46.1-254 ng/mL in feces and urine, respectively. This robust, sensitive, and accurate method can be used to assess the fate and environmental impact of antibiotics used on farms.

  15. Determination of cadmium and lead in human biological samples by spectrometric techniques: a review.

    Science.gov (United States)

    Lemos, Valfredo Azevedo; de Carvalho, Anaildes Lago

    2010-12-01

    The analysis of human biological samples, such as blood, urine, nails, and hair, is generally used for the verification of human exposure to toxic metals. In this review, various spectrometric methods for the determination of cadmium and lead in biological samples are discussed and compared. Several spectrometric techniques are presented and discussed with respect to various characteristics such as sensitivity, selectivity, and cost. Special attention is drawn to the procedures for digestion prior to the determination of cadmium and lead in hair, nails, blood, and urine.

  16. Detection of Campylobacter in human and animal field samples in Cambodia.

    Science.gov (United States)

    Osbjer, Kristina; Tano, Eva; Chhayheng, Leang; Mac-Kwashie, Akofa Olivia; Fernström, Lise-Lotte; Ellström, Patrik; Sokerya, Seng; Sokheng, Choup; Mom, Veng; Chheng, Kannarath; San, Sorn; Davun, Holl; Boqvist, Sofia; Rautelin, Hilpi; Magnusson, Ulf

    2016-06-01

    Campylobacter are zoonotic bacteria and a leading cause of human gastroenteritis worldwide with Campylobacter jejuni and C. coli being the most commonly detected species. The aim of this study was to detect Campylobacter in humans and livestock (chickens, ducks, pigs, cattle, water buffalo, quail, pigeons and geese) in rural households by routine culturing and multiplex PCR in faecal samples frozen before analysis. Of 681 human samples, 82 (12%) tested positive by PCR (C. jejuni in 66 samples and C. coli in 16), but none by routine culture. Children were more commonly Campylobacter positive (19%) than adult males (8%) and females (7%). Of 853 livestock samples, 106 (12%) tested positive by routine culture and 352 (41%) by PCR. Campylobacter jejuni was more frequent in chickens and ducks and C. coli in pigs. In conclusion, Campylobacter proved to be highly prevalent by PCR in children (19%), ducks (24%), chickens (56%) and pigs (72%). Routine culturing was insufficiently sensitive in detecting Campylobacter in field samples frozen before analysis. These findings suggest that PCR should be the preferred diagnostic method for detection of Campylobacter in humans and livestock where timely culture is not feasible. © 2016 The Authors. APMIS published by John Wiley & Sons Ltd on behalf of Scandinavian Societies for Medical Microbiology and Pathology.

  17. Proteomic biomarker discovery in 1000 human plasma samples with mass spectrometry

    DEFF Research Database (Denmark)

    Cominetti, Ornella; Núñez Galindo, Antonio; Corthésy, John

    2016-01-01

    the quality of the MS data and provided descriptive statistics. The data set was interrogated for proteins with most stable expression levels in that set of plasma samples. We evaluated standard clinical variables that typically impact forthcoming results and assessed body mass index-associated and gender......-specific proteins at two time points. We demonstrate that analyzing a large number of human plasma samples for biomarker discovery with MS using isobaric tagging is feasible, providing robust and consistent biological results....

  18. Assessment of Salmonella spp. in feces, cloacal swabs, and eggs (eggshell and content separately) from a laying hen farm.

    Science.gov (United States)

    García, C; Soriano, J M; Benítez, V; Catalá-Gregori, P

    2011-07-01

    Microbial pathogens of the genus Salmonella are among the leading causes of foodborne illness in the world. The present study was done on a laying hen farm with a Salmonella enterica serovar Enteritidis-positive result according to the testing specified by European regulation 2160/2003. The aim of this study was to compare the Salmonella contamination on a laying hen farm with the Salmonella presence in the hen eggs. The strains were isolated by ISO method 6579:2002 (standard method for the detection of Salmonella spp. in the European regulation for food and animal feeding stuffs, animal feces, and environmental samples from the primary production stage, including poultry farms) and were confirmed as Salmonella Enteritidis by the Kauffmann-White method. In addition, strains were compared with genomic macrorestriction followed by pulsed-field gel electrophoresis. Four types of samples, namely, feces (n = 50), cloacal swabs (n = 150), eggshells (n = 50), and egg contents (n = 50), were taken from each of 50 randomly selected battery cages. Results demonstrated that feces (92%) were the most positive sample, followed by eggshells (34%) and cloacal swabs (4%). No Salmonella spp. were detected in the egg contents. Our results show that a Salmonella Enteritidis-positive result on a laying hen farm, according to the testing specified by European regulation 2160/2003, did not imply the presence of the pathogen in the egg contents. Additionally, XbaI-digested genomic DNA of Salmonella Enteritidis strains isolated from several samples resulted in the same pattern, so were probably of the same origin.

  19. Rapid and Decentralized Human Waste Treatment by Microwave Radiation.

    Science.gov (United States)

    Nguyen, Tu Anh; Babel, Sandhya; Boonyarattanakalin, Siwarutt; Koottatep, Thammarat

    2016-09-07

    This study evaluates the technical feasibility of using microwave radiation for the rapid treatment of human feces. Human feces of 1000 g were radiated with a commercially available household microwave oven (with rotation) at different exposure time lengths (30, 50, 60, 70, and 75 minutes) and powers (600, 800, and 1000 W). Volume reduction over 90% occurred after 1000 W microwave radiation for 75 minutes. Pathogen eradiation performances of six log units or more at a high range of microwave powers were achieved. Treatments with the same energy input of 1000 Wh, but at lower powers with prolonged exposure times, significantly enhanced moisture removal and volume reduction. Microwave radiation caused carbonization and resulted in a more stable end product. The energy content of the samples after microwave treatment at 1000 W and 75 minutes is 3517 ± 8.85 calories/g of dried sample, and the product can also be used as compost.

  20. Isolation of Viable Toxoplasma gondii from Tissues and Feces of Cats from Addis Ababa, Ethiopia

    Science.gov (United States)

    Cats are important in the epidemiology of Toxoplasma gondii because they are the only hosts that excrete environmentally resistant oocysts in feces. In the present study, hearts, serum, and feces from 36 feral cats from Addis Ababa area, Ethiopia were examined for T. gondii infection. Antibodies to ...

  1. Quantification of bioavailable chlortetracycline in pig feces using a bacterial whole-cell biosensor

    DEFF Research Database (Denmark)

    Hansen, Lars Hestbjerg; Aarestrup, Frank; Sørensen, Søren Johannes

    2002-01-01

    Bacterial whole-cell biosensors were used to measure the concentration of chlortetracycline (CTC) in the feces of pigs. In this study, the Escherichia coli biosensor used has a detection limit of 0.03 mg/kg CTC in pig feces. The tetracycline concentration was correlated with the appearance...

  2. Quantification of bioavailable chlortetracycline in pig feces using a bacterial whole-cell biosensor

    DEFF Research Database (Denmark)

    Hansen, L. H.; Aarestrup, Frank Møller; Sørensen, S. J.

    2002-01-01

    Bacterial whole-cell biosensors were used to measure the concentration of chlortetracycline (CTC) in the feces of pigs. In this study, the Escherichia coli biosensor used has a detection limit of 0.03 mg/kg CTC in pig feces. The tetracycline concentration was correlated with the appearance...

  3. China's human resources for maternal and child health: a national sampling survey.

    Science.gov (United States)

    Ren, Zhenghong; Song, Peige; Theodoratou, Evropi; Guo, Sufang; An, Lin

    2015-12-16

    In order to achieve the Millennium Development Goals (MDG) 4 and 5, the Chinese Government has invested greatly in improving maternal and child health (MCH) with impressive results. However, one of the most important barriers for further improvement is the uneven distribution of MCH human resources. There is little information about the distribution, quantity and capacity of the Chinese MCH human resources and we sought to investigate this. Cities at prefectural level were selected by random cluster sampling. All medical and health institutions providing MCH-related services in the sampled areas were investigated using a structured questionnaire. The data were weighted based on the proportion of the sampled districts/cities. Amount, proportions and numbers per 10,000 population of MCH human resources were estimated in order to reveal the quantity of the Chinese MCH human resources. The capacity of MCH human resources was evaluated by analyzing data on the education level and professional skills of the staff. There were 77,248 MCH workers in China in 2010. In general, 67.6% and 71.9% of the women's and children's health care professionals had an associate degree or higher, whereas around 30% had only high-school or lower degrees. More than 40% of the women's health workers were capable of providing skilled birth attendance, but these proportions varied between different institutions and locations. Evidence from this study highlights that Chinese MCH human resources are not in shortage in the national level. However, the quantity and capacity of MCH human resources are not evenly distributed among different institutions and locations. Finally there is a need in the improvement of the MCH services by improving the quality of MCH human resources.

  4. Detection of feline coronavirus in cheetah (Acinonyx jubatus) feces by reverse transcription-nested polymerase chain reaction in cheetahs with variable frequency of viral shedding.

    Science.gov (United States)

    Gaffney, Patricia M; Kennedy, Melissa; Terio, Karen; Gardner, Ian; Lothamer, Chad; Coleman, Kathleen; Munson, Linda

    2012-12-01

    Cheetahs (Acinonyx jubatus) are a highly threatened species because of habitat loss, human conflict, and high prevalence of disease in captivity. An epidemic of feline infectious peritonitis and concern for spread of infectious disease resulted in decreased movement of cheetahs between U.S. zoological facilities for managed captive breeding. Identifying the true feline coronavirus (FCoV) infection status of cheetahs is challenging because of inconsistent correlation between seropositivity and fecal viral shedding. Because the pattern of fecal shedding of FCoV is unknown in cheetahs, this study aimed to assess the frequency of detectable fecal viral shedding in a 30-day period and to determine the most efficient fecal sampling strategy to identify cheetahs shedding FCoV. Fecal samples were collected from 16 cheetahs housed at seven zoological facilities for 30 to 46 consecutive days; the samples were evaluated for the presence of FCoV by reverse transcription-nested polymerase chain reaction (RT-nPCR). Forty-four percent (7/16) of cheetahs had detectable FCoV in feces, and the proportion of positive samples for individual animals ranged from 13 to 93%. Cheetahs shed virus persistently, intermittently, or rarely over 30-46 days. Fecal RT-nPCR results were used to calculate the probability of correctly identifying a cheetah known to shed virus given multiple hypothetical fecal collection schedules. The most efficient hypothetical fecal sample collection schedule was evaluation of five individual consecutive fecal samples, resulting in a 90% probability of identifying a known shedder. Demographic and management risk factors were not significantly associated (P cheetahs shed virus intermittently to rarely, fecal sampling schedules meant to identify all known shedders would be impractical with current tests and eradication of virus from the population unreasonable. Managing the captive population as endemically infected with FCoV may be a more feasible approach.

  5. Use of culture, PCR analysis, and DNA microarrays for detection of Campylobacter jejuni and Campylobacter coli from chicken feces

    DEFF Research Database (Denmark)

    Keramas, Georgios; Bang, Dang Duong; Lund, Marianne

    2004-01-01

    A DNA microarray for detection of Campylobacter spp. was recently developed and applied to detect Campylobacter spp. directly from chicken feces. Sixty-five pooled chicken cloacal swab samples from 650 individual broiler chickens were included in the study. The results of Campylobacter sp....... detection obtained with DNA microarrays were compared to those obtained by conventional culture and gel electrophoresis. By conventional culture, 60% of the samples were positive for either Campylobacter jejuni or Campylobacter coli. By PCR and capillary electrophoresis, 95% of the samples were positive...... for Campylobacter spp., whereas with DNA microarrays all samples were positive for Campylobacter spp. By application of DNA microarray analysis, the isolates in 4 samples (6%) could not be identified to the species level, whereas by PCR-capillary electrophoresis, the isolates in 12 samples (19%) remained...

  6. Efficient discrimination and removal of phospholipids during electromembrane extraction from human plasma samples

    DEFF Research Database (Denmark)

    Vårdal, Linda; Gjelstad, Astrid; Huang, Chuixiu

    2017-01-01

    AIM: For the first time, extracts obtained from human plasma samples by electromembrane extraction (EME) were investigated comprehensively with particular respect to phospholipids using ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). Thhe purpose was to invest...

  7. Imitation of Tongue Protrusion in Human Neonates: Specificity of the Response in a Large Sample

    Science.gov (United States)

    Nagy, Emese; Pilling, Karen; Orvos, Hajnalka; Molnar, Peter

    2013-01-01

    Although a large body of evidence has accumulated on the young human infant's ability to imitate, the phenomenon has failed to gain unanimous acceptance. Imitation of tongue protrusion, the most tested gesture to date, was examined in a sample of 115 newborns in the first 5 days of life in 3 seating positions. An ethologically based…

  8. Analyses of human colonic mucus obtained by an in vivo sampling technique

    NARCIS (Netherlands)

    Hamer, H.M.; Jonkers, D.M.A.E.; Loof, A.; Houtvin, S.A.L.W. van; Troost, F.J.; Venema, K.; Kodde, A.; Koek, G.H.; Schipper, R.G.; Heerde, W.L. van; Brummer, R.J.

    2009-01-01

    Background: The mucus layer is an important dynamic component of the epithelial barrier. It contains mucin glycoproteins and other compounds secreted by the intestinal epithelium, such as secretory IgA. However, a standardized in vivo sampling technique of mucus in humans is not yet available. Aim:

  9. Groundwater sampling methods using glass wool filtration to trace human enteric viruses in Madison, Wisconsin

    Science.gov (United States)

    Human enteric viruses have been detected in the Madison, Wisconsin deep municipal well system. Earlier projects by the Wisconsin Geological and Natural History Survey (WGNHS) have used glass wool filters to sample groundwater for these viruses directly from the deep municipal wells. Polymerase chain...

  10. Elimination of bioweapons agents from forensic samples during extraction of human DNA.

    Science.gov (United States)

    Timbers, Jason; Wilkinson, Della; Hause, Christine C; Smith, Myron L; Zaidi, Mohsin A; Laframboise, Denis; Wright, Kathryn E

    2014-11-01

    Collection of DNA for genetic profiling is a powerful means for the identification of individuals responsible for crimes and terrorist acts. Biologic hazards, such as bacteria, endospores, toxins, and viruses, could contaminate sites of terrorist activities and thus could be present in samples collected for profiling. The fate of these hazards during DNA isolation has not been thoroughly examined. Our goals were to determine whether the DNA extraction process used by the Royal Canadian Mounted Police eliminates or neutralizes these agents and if not, to establish methods that render samples safe without compromising the human DNA. Our results show that bacteria, viruses, and toxins were reduced to undetectable levels during DNA extraction, but endospores remained viable. Filtration of samples after DNA isolation eliminated viable spores from the samples but left DNA intact. We also demonstrated that contamination of samples with some bacteria, endospores, and toxins for longer than 1 h compromised the ability to complete genetic profiling.

  11. Long-term performance and stability of molecular shotgun lipidomic analysis of human plasma samples.

    Science.gov (United States)

    Heiskanen, Laura A; Suoniemi, Matti; Ta, Hung Xuan; Tarasov, Kirill; Ekroos, Kim

    2013-09-17

    The stability of the lipid concentration levels in shotgun lipidomics analysis was tracked over a period of 3.5 years. Concentration levels in several lipid classes, such as phospholipids, were determined in human plasma lipid extracts. Impact of the following factors on the analysis was investigated: sample amount, internal standard amount, and sample dilution factor. Moreover, the reproducibility of lipid profiles obtained in both polarity modes was evaluated. Total number of samples analyzed was approximately 6800 and 7300 samples in negative and positive ion modes, respectively, out of which 610 and 639 instrument control samples were used in stability calculations. The assessed shotgun lipidomics approach showed to be remarkably robust and reproducible, requiring no batch corrections. Coefficients of variation (CVs) of lipid mean concentration measured with optimized analytical parameters were typically less than 15%. The high reproducibility indicated that no lipid degradation occurred during the monitored time period.

  12. A tool kit for quantifying eukaryotic rRNA gene sequences from human microbiome samples.

    Science.gov (United States)

    Dollive, Serena; Peterfreund, Gregory L; Sherrill-Mix, Scott; Bittinger, Kyle; Sinha, Rohini; Hoffmann, Christian; Nabel, Christopher S; Hill, David A; Artis, David; Bachman, Michael A; Custers-Allen, Rebecca; Grunberg, Stephanie; Wu, Gary D; Lewis, James D; Bushman, Frederic D

    2012-07-03

    Eukaryotic microorganisms are important but understudied components of the human microbiome. Here we present a pipeline for analysis of deep sequencing data on single cell eukaryotes. We designed a new 18S rRNA gene-specific PCR primer set and compared a published rRNA gene internal transcribed spacer (ITS) gene primer set. Amplicons were tested against 24 specimens from defined eukaryotes and eight well-characterized human stool samples. A software pipeline https://sourceforge.net/projects/brocc/ was developed for taxonomic attribution, validated against simulated data, and tested on pyrosequence data. This study provides a well-characterized tool kit for sequence-based enumeration of eukaryotic organisms in human microbiome samples.

  13. A human gut metaproteomic dataset from stool samples pretreated or not by differential centrifugation

    Directory of Open Access Journals (Sweden)

    Alessandro Tanca

    2015-09-01

    Full Text Available We present a human gut metaproteomic dataset deposited in the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD001573. Ten aliquots of a single stool sample collected from a healthy human volunteer were either pretreated by differential centrifugation (DC; N=5 or not centrifuged (NC; N=5. Protein extracts were then processed by filter-aided sample preparation, single-run liquid chromatography and high-resolution mass spectrometry, and peptide identification was carried out using Sequest-HT as search engine within the Proteome Discoverer informatic platform. The dataset described here is also related to the research article entitled “Enrichment or depletion? The impact of stool pretreatment on metaproteomic characterization of the human gut microbiota” published in Proteomics (Tanca et al., 2015, [1].

  14. Easily-handled method to isolate mesenchymal stem cells from coagulated human bone marrow samples

    Institute of Scientific and Technical Information of China (English)

    Heng-Xiang; Wang; Zhi-Yong; Li; Zhi-Kun; Guo; Zi-Kuan; Guo

    2015-01-01

    AIM:To establish an easily-handled method to isolate mesenchymal stem cells(MSCs) from coagulated human bone marrow samples. METHODS: Thrombin was added to aliquots of seven heparinized human bone marrow samples to mimic marrow coagulation. The clots were untreated,treated with urokinase or mechanically cut into pieces before culture for MSCs. The un-coagulated samples and the clots were also stored at 4 ℃ for 8 or 16 h before the treatment. The numbers of colony-forming unit-fibroblast(CFU-F) in the different samples were determined. The adherent cells from different groups were passaged and their surface profile was analyzed with flow cytometry. Their capacities of in vitro osteogenesis and adipogenesis were observed after the cells were exposed to specific inductive agents.RESULTS: The average CFU-F number of urokinasetreated samples(16.85 ± 11.77/106) was comparable to that of un-coagulated control samples(20.22 ± 10.65/106,P = 0.293),which was significantly higher than those of mechanically-cut clots(6.5 ± 5.32/106,P < 0.01) and untreated clots(1.95 ± 1.86/106,P < 0.01). The CFU-F numbers decreased after samples were stored,but those of control and urokinase-treated clots remained higher than the other two groups. Consistently,the numbers of the attached cells at passage 0 were higher in control and urokinase-treated clots than those of mechanically-cut clots and untreated clots.The attached cells were fibroblast-like in morphology and homogenously positive for CD44,CD73 and CD90,and negative for CD31 and CD45. Also,they could be induced to differentiate into osteoblasts and adipocytes in vitro. CONCLUSION: Urokinase pretreatment is an optimal strategy to isolate MSCs from human bone marrow samples that are poorly aspirated and clotted.

  15. Rigorous Training of Dogs Leads to High Accuracy in Human Scent Matching-To-Sample Performance.

    Directory of Open Access Journals (Sweden)

    Sophie Marchal

    Full Text Available Human scent identification is based on a matching-to-sample task in which trained dogs are required to compare a scent sample collected from an object found at a crime scene to that of a suspect. Based on dogs' greater olfactory ability to detect and process odours, this method has been used in forensic investigations to identify the odour of a suspect at a crime scene. The excellent reliability and reproducibility of the method largely depend on rigor in dog training. The present study describes the various steps of training that lead to high sensitivity scores, with dogs matching samples with 90% efficiency when the complexity of the scents presented during the task in the sample is similar to that presented in the in lineups, and specificity reaching a ceiling, with no false alarms in human scent matching-to-sample tasks. This high level of accuracy ensures reliable results in judicial human scent identification tests. Also, our data should convince law enforcement authorities to use these results as official forensic evidence when dogs are trained appropriately.

  16. Sequence space coverage, entropy of genomes and the potential to detect non-human DNA in human samples

    Directory of Open Access Journals (Sweden)

    Maley Carlo C

    2008-10-01

    Full Text Available Abstract Background Genomes store information for building and maintaining organisms. Complete sequencing of many genomes provides the opportunity to study and compare global information properties of those genomes. Results We have analyzed aspects of the information content of Homo sapiens, Mus musculus, Drosophila melanogaster, Caenorhabditis elegans, Arabidopsis thaliana, Saccharomyces cerevisiae, and Escherichia coli (K-12 genomes. Virtually all possible (> 98% 12 bp oligomers appear in vertebrate genomes while 98% to D. melanogaster (12–17 bp, C. elegans (11–17 bp, A. thaliana (11–17 bp, S. cerevisiae (10–16 bp and E. coli (9–15 bp. Frequencies of unique oligomers in the genomes follow similar patterns. We identified a set of 2.6 M 15-mers that are more than 1 nucleotide different from all 15-mers in the human genome and so could be used as probes to detect microbes in human samples. In a human sample, these probes would detect 100% of the 433 currently fully sequenced prokaryotes and 75% of the 3065 fully sequenced viruses. The human genome is significantly more compact in sequence space than a random genome. We identified the most frequent 5- to 20-mers in the human genome, which may prove useful as PCR primers. We also identified a bacterium, Anaeromyxobacter dehalogenans, which has an exceptionally low diversity of oligomers given the size of its genome and its GC content. The entropy of coding regions in the human genome is significantly higher than non-coding regions and chromosomes. However chromosomes 1, 2, 9, 12 and 14 have a relatively high proportion of coding DNA without high entropy, and chromosome 20 is the opposite with a low frequency of coding regions but relatively high entropy. Conclusion Measures of the frequency of oligomers are useful for designing PCR assays and for identifying chromosomes and organisms with hidden structure that had not been previously recognized. This information may be used to detect

  17. Development and evaluation of a sensitive PCR-ELISA system for detection of schistosoma infection in feces.

    Directory of Open Access Journals (Sweden)

    Luciana Inácia Gomes

    Full Text Available BACKGROUND: A PCR-enzyme-linked immunosorbent assay (PCR-ELISA was developed to overcome the need for sensitive techniques for the efficient diagnosis of Schistosoma infection in endemic settings with low parasitic burden. METHODOLOGY/PRINCIPAL FINDINGS: This system amplifies a 121-base pair tandem repeat DNA sequence, immobilizes the resultant 5' biotinylated product on streptavidin-coated strip-well microplates and uses anti-fluorescein antibodies conjugated to horseradish peroxidase to detect the hybridized fluorescein-labeled oligonucleotide probe. The detection limit of the Schistosoma PCR-ELISA system was determined to be 1.3 fg of S. mansoni genomic DNA (less than the amount found in a single cell and estimated to be 0.15 S. mansoni eggs per gram of feces (fractions of an egg. The system showed good precision and genus specificity since the DNA target was found in seven Schistosoma DNA samples: S. mansoni, S. haematobium, S. bovis, S. intercalatum, S. japonicum, S. magrebowiei and S. rhodaini. By evaluating 206 patients living in an endemic area in Brazil, the prevalence of S. mansoni infection was determined to be 18% by examining 12 Kato-Katz slides (41.7 mg/smear, 500 mg total of a single fecal sample from each person, while the Schistosoma PCR-ELISA identified a 30% rate of infection using 500-mg of the same fecal sample. When considering the Kato-Katz method as the reference test, artificial sensitivity and specificity rates of the PCR-ELISA system were 97.4% and 85.1%, respectively. The potential for estimating parasitic load by DNA detection in feces was assessed by comparing absorbance values and eggs per gram of feces, with a Spearman correlation coefficient of 0.700 (P<0.0001. CONCLUSIONS/SIGNIFICANCE: This study reports the development and field evaluation of a sensitive Schistosoma PCR-ELISA, a system that may serve as an alternative for diagnosing Schistosoma infection.

  18. Quantification of 15 bile acids in lake charr feces by ultra-high performance liquid chromatography–tandem mass spectrometry

    Science.gov (United States)

    Li, Ke; Buchinger, Tyler J.; Bussy, Ugo; Fissette, Skye D; Johnson, Nicholas; Li, Weiming

    2015-01-01

    Many fishes are hypothesized to use bile acids (BAs) as chemical cues, yet quantification of BAs in biological samples and the required methods remain limited. Here, we present an UHPLC–MS/MS method for simultaneous, sensitive, and rapid quantification of 15 BAs, including free, taurine, and glycine conjugated BAs, and application of the method to fecal samples from lake charr (Salvelinus namaycush). The analytes were separated on a C18 column with acetonitrile–water (containing 7.5 mM ammonium acetate and 0.1% formic acid) as mobile phase at a flow rate of 0.25 mL/min for 12 min. BAs were monitored with a negative electrospray triple quadrupole mass spectrometer (Xevo TQ-S™). Calibration curves of 15 BAs were linear over the concentration range of 1.00–5,000 ng/mL. Validation revealed that the method was specific, accurate, and precise. The method was applied to quantitative analysis of feces extract of fry lake charr and the food they were eating. The concentrations of analytes CA, TCDCA, TCA, and CDCA were 242.3, 81.2, 60.7, and 36.2 ng/mg, respectively. However, other taurine conjugated BAs, TUDCA, TDCA, and THDCA, were not detected in feces of lake charr. Interestingly, TCA and TCDCA were detected at high concentrations in food pellets, at 71.9 and 38.2 ng/mg, respectively. Application of the method to feces samples from lake charr supported a role of BAs as chemical cues, and will enhance further investigation of BAs as chemical cues in other fish species.

  19. Evaluation of non-invasive biological samples to monitor Staphylococcus aureus colonization in great apes and lemurs.

    Directory of Open Access Journals (Sweden)

    Frieder Schaumburg

    Full Text Available INTRODUCTION: Reintroduction of endangered animals as part of conservational programs bears the risk of importing human pathogens from the sanctuary to the natural habitat. One bacterial pathogen that serves as a model organism to analyze this transmission is Staphylococcus aureus as it can colonize and infect both humans and animals. The aim of this study was to evaluate the utility of various biological samples to monitor S. aureus colonization in great apes and lemurs. METHODS: Mucosal swabs from wild lemurs (n=25, Kirindy, Madagascar, feces, oral and genital swabs from captive chimpanzees (n=58, Ngamba and Entebbe, Uganda and fruit wadges and feces from wild chimpanzees (n=21, Taï National Parc, Côte d'Ivoire were screened for S. aureus. Antimicrobial resistance and selected virulence factors were tested for each isolate. Sequence based genotyping (spa typing, multilocus sequence typing was applied to assess the population structure of S. aureus. RESULTS: Oro-pharyngeal carriage of S. aureus was high in lemurs (72%, n=18 and captive chimpanzees (69.2%, n=27 and 100%, n=6, respectively. Wild chimpanzees shed S. aureus through feces (43.8, n=7 and fruit wadges (54.5, n=12. Analysis of multiple sampling revealed that two samples are sufficient to detect those animals which shed S. aureus through feces or fruit wadges. Genotyping showed that captive animals are more frequently colonized with human-associated S. aureus lineages. CONCLUSION: Oro-pharyngeal swabs are useful to screen for S. aureus colonization in apes and lemurs before reintroduction. Duplicates of stool and fruit wadges reliably detect S. aureus shedding in wild chimpanzees. We propose to apply these sampling strategies in future reintroduction programs to screen for S. aureus colonization. They may also be useful to monitor S. aureus in wild populations.

  20. SAMPLING INTENSITY WITH FIXED PRECISION WHEN ESTIMATING VOLUME OF HUMAN BRAIN COMPARTMENTS

    Directory of Open Access Journals (Sweden)

    Rhiannon Maudsley

    2011-05-01

    Full Text Available Cavalieri sampling and point counting are frequently applied in combination with magnetic resonance (MR imaging to estimate the volume of human brain compartments. Current practice involves arbitrarily choosing the number of sections and sampling intensity within each section, and subsequently applying error prediction formulae to estimate the precision. The aim of this study is to derive a reference table for researchers who are interested in estimating the volume of brain regions, namely grey matter, white matter, and their union, to a given precision. In particular, this table, which is based on subsampling of a large brain data set obtained from coronal MR images, offers a recommendation for the minimum number of sections and mean number of points per section that are required to achieve a pre-defined coefficient of error of the volume estimator. Further analysis onMR brain data from a second human brain shows that the sampling intensity recommended is appropriate.

  1. Determination of cholesterol concentration in human milk samples using attenuated total reflectance Fourier transform infrared spectroscopy

    Science.gov (United States)

    Kamelska, A. M.; Pietrzak-Fiećko, R.; Bryl, K.

    2013-03-01

    Results of an inexpensive and rapid evaluation of the cholesterol concentration in human milk using ATR-FTIR techniques are presented. The FTIR spectrum of pure cholesterol was characterized and quantitatively estimated in the region between 2800 and 3200 cm-1. 125 samples at different stages of lactation were analyzed. There were no differences between the cholesterol concentrations in the samples of early (1-3 months), medium (4-6 months), and late (> 6 months) lactation stages ( p = 0.096968). The cholesterol concentration ranged from 4.30 to 21.77 mg/100 cm3. Such a broad range was due to the differences between the samples from different women ( p = 0.000184). The results indicate that ATR-FTIR has potential for rapid estimation of cholesterol concentration in human milk.

  2. Use of Serial Quantitative PCR of the vapA Gene of Rhodococcus equi in Feces for Early Detection of R. equi Pneumonia in Foals.

    Science.gov (United States)

    Madrigal, R G; Shaw, S D; Witkowski, L A; Sisson, B E; Blodgett, G P; Chaffin, M K; Cohen, N D

    2016-01-01

    Current screening tests for Rhodococcus equi pneumonia in foals lack adequate accuracy for clinical use. Real-time, quantitative PCR (qPCR) for virulent R. equi in feces has not been systematically evaluated as a screening test. The objective of this study was to evaluate the accuracy of qPCR for vapA in serially collected fecal samples as a screening test for R. equi pneumonia in foals. One hundred and twenty-five foals born in 2011 at a ranch in Texas. Fecal samples were collected concurrently with thoracic ultrasonography (TUS) screening examinations at ages 3, 5, and 7 weeks. Affected (pneumonic) foals (n = 25) were matched by age and date-of-birth to unaffected (n = 25) and subclinical (ie, having thoracic TUS lesions but no clinical signs of pneumonia) foals (n = 75). DNA was extracted from feces using commercial kits and concentration of virulent R. equi in feces was determined by qPCR. Subsequently affected foals had significantly greater concentrations of vapA in feces than foals that did not develop pneumonia (unaffected and subclinical foals) at 5 and 7 weeks of age. Accuracy of fecal qPCR, however, was poor as a screening test to differentiate foals that would develop clinical signs of pneumonia from those that would remain free of clinical signs (including foals with subclinical pulmonary lesions attributed to R. equi) using receiver operating characteristic (ROC) methods. In the population studied, serial qPCR on feces lacked adequate accuracy as a screening test for clinical R. equi foal pneumonia. Copyright © 2016 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.

  3. Role of intestinal microbiota in transformation of bismuth and other metals and metalloids into volatile methyl and hydride derivatives in humans and mice.

    Science.gov (United States)

    Michalke, Klaus; Schmidt, Annette; Huber, Britta; Meyer, Jörg; Sulkowski, Margareta; Hirner, Alfred V; Boertz, Jens; Mosel, Frank; Dammann, Philip; Hilken, Gero; Hedrich, Hans J; Dorsch, Martina; Rettenmeier, Albert W; Hensel, Reinhard

    2008-05-01

    The present study shows that feces samples of 14 human volunteers and isolated gut segments of mice (small intestine, cecum, and large intestine) are able to transform metals and metalloids into volatile derivatives ex situ during anaerobic incubation at 37 degrees C and neutral pH. Human feces and the gut of mice exhibit highly productive mechanisms for the formation of the toxic volatile derivative trimethylbismuth [(CH(3))(3)Bi] at rather low concentrations of bismuth (0.2 to 1 mumol kg(-1) [dry weight]). An increase of bismuth up to 2 to 14 mmol kg(-1) (dry weight) upon a single (human volunteers) or continuous (mouse study) administration of colloidal bismuth subcitrate resulted in an average increase of the derivatization rate from approximately 4 pmol h(-1) kg(-1) (dry weight) to 2,100 pmol h(-1) kg(-1) (dry weight) in human feces samples and from approximately 5 pmol h(-1) kg(-1) (dry weight) to 120 pmol h(-1) kg(-1) (dry weight) in mouse gut samples, respectively. The upshift of the bismuth content also led to an increase of derivatives of other elements (such as arsenic, antimony, and lead in human feces or tellurium and lead in the murine large intestine). The assumption that the gut microbiota plays a dominant role for these transformation processes, as indicated by the production of volatile derivatives of various elements in feces samples, is supported by the observation that the gut segments of germfree mice are unable to transform administered bismuth to (CH(3))(3)Bi.

  4. Identification of human Norovirus (HNoV in domestic pig stool samples

    Directory of Open Access Journals (Sweden)

    María F. Gutiérrez

    2011-08-01

    Full Text Available To determine the presence of NoVs as a possible causal zoonotic agent of acute diarrhea in pigs and humans. Materialsand methods. We collected a total of 77 samples from diarrheal children under 5 years and pigs under 2 months from La Chambatown in Tolima, Colombia. These samples were transported to the Laboratory of Virology of the Pontificia Universidad Javerianain Bogotá, and extraction with Trizol-reagent was done following the manufacturer’s instructions. After obtaining the RNA, thenext step was to perform RT-PCR for obtaining the expected amplification product of 213- bp NoVs. Finally, the positive samplesobtained in the RT-PCR were sequenced and analyzed by bioinformatics methods. Results. Six positive diarrheic samples fromchildren and a positive diarrheic sample from pigs were detected by a band of 231 bp. Five of the six positive samples in childrenand the positive pig sample were sequenced and analyzed. Conclusion. Given the close genetic relationship between pig andhuman sequences, this could be an indication of the potential existence of a common animal acting as a reservoir for human orother animal strains.

  5. Total reflection X-ray fluorescence as a fast multielemental technique for human placenta sample analysis

    Science.gov (United States)

    Marguí, E.; Ricketts, P.; Fletcher, H.; Karydas, A. G.; Migliori, A.; Leani, J. J.; Hidalgo, M.; Queralt, I.; Voutchkov, M.

    2017-04-01

    In the present contribution, benchtop total reflection X-ray fluorescence spectrometry (TXRF) has been evaluated as a cost-effective multielemental analytical technique for human placenta analysis. An easy and rapid sample preparation consisting of suspending 50 mg of sample in 1 mL of a Triton 1% solution in deionized water showed to be the most suitable for this kind of samples. However, for comparison purposes, an acidic microwave acidic digestion procedure was also applied. For both sample treatment methodologies, limits of detection for most elements were in the low mg/kg level. Accurate and precise results were obtained using internal standardization as quantification approach and applying a correction factor to compensate for absorption effects. The correction factor was based on the proportional ratio between the slurry preparation results and those obtained for the analysis of a set of human placenta samples analysed by microwave acidic digestion and ICP-AES analysis. As a study case, the developed TXRF methodology was applied for multielemental analysis (K, Ca, Fe, Cu, Zn, As, Se, Br, Rb and Sr) of several healthy women's placenta samples from two regions in Jamaica.

  6. Residues of PCDDs and PCDFs in human milk samples in Ahmedabad, India

    Energy Technology Data Exchange (ETDEWEB)

    Kashyap, R.; Bhatnagar, V.; Sadhu, H.; Jhamb, N.; Karanjkar, R.; Saiyed, H. [National Inst. of Occupational Health, Ahmedabad (India)

    2004-09-15

    Polychlorinated dibenzo-p-dioxins (PCDDs) and Polychlorinated dibenzo furans (PCDFs) represent a class of organic environmental pollutants. They are unwanted byproduct of incineration, uncontrolled burning and certain industrial processes. They are persistent in nature and bioaccumulates through food chain. These are hazardous to human health and environment. The residues of these toxicants have been detected in human adipose tissue, blood and milk. WHO has coordinated two rounds of follow up studies on levels of PCDD/Fs and PCBs in human milk and the data shows a decreasing trend during the last 30 years. However, in India there is no data available on the exposure and residues of these contaminants. This study presents first time the levels of dioxin and furans in human milk samples collected from the Ahmedabad city in India.

  7. Occurrence and characterization of toxigenic Bacillus cereus in food and infant feces

    Institute of Scientific and Technical Information of China (English)

    Sameer; Rushdi; Organji; Hussein; Hasan; Abulreesh; Khaled; Elbanna; Gamal; Ebrahim; Haridy; Osman; Manal; Khider

    2015-01-01

    Objective: To investigate the true incidence of Bacillus cereus(B. cereus) in food and children diarrhea cases. Methods: A total of 110 samples of various dairy products such as raw milk, long life pasteurized milk, yoghurt and infant powdered milk formulas, raw rice, and feces were examined for the presence of B. cereus by selective plating on mannitol-egg-yolk-polymyxin agar. Confirmation of B. cereus was carried out by biochemical tests and PCR. Identification of non-B. cereus isolates was carried out by 16 S r DNA sequencing. Antimicrobial susceptibility was done by disk diffusion method.Results: Overall 35 samples(31.8%, n = 110) yielded Bacillus-like growth. Of which 19 samples(54.28%) were positive for B. cereus. All isolates were positive for enterotoxin production. No psychrotolerant B. cereus strains were detected in all samples. All B. cereus isolates were resistant to penicillin G, but susceptible to vancomycin, erythromycin and clindamycin. Conclusions: The results of this study confirm the importance of including B. cereus in disease control and prevention programs, as well as in routine clinical and food quality control laboratories in both Saudi Arabia and Egypt.

  8. Enumeration and isolation of cpe-positive Clostridium perfringens spores from feces.

    Science.gov (United States)

    Heikinheimo, Annamari; Lindström, Miia; Korkeala, Hannu

    2004-09-01

    A hydrophobic grid membrane filter-colony hybridization (HGMF-CH) method for the enumeration and isolation of cpe gene-carrying (cpe-positive) Clostridium perfringens spores from feces was developed. A 425-bp DNA probe specific for the cpe gene was sensitive and specific when tested with bacterial DNA and pure cultures. The enumeration of cpe-positive C. perfringens by the HGMF-CH method proved to be as sensitive as nested PCR combined with the most-probable number technique when tested with fecal samples from healthy individuals. With the aid of the HGMF-CH method, positive hybridization signals were detected from two out of seven fecal samples obtained from healthy individuals. Furthermore, cpe-positive C. perfringens was successfully isolated from both of these samples. The detection of cpe-positive C. perfringens by the HGMF-CH method is dependent on the ratio of cpe-positive C. perfringens colonies to total C. perfringens colonies growing on the HGMF-tryptose-sulfite-cycloserine plate. cpe-positive C. perfringens could be isolated if the ratio of cpe-positive C. perfringens spores to total C. perfringens spores was 6 x 10(-5) or higher. The HGMF-CH method provides an aid in the investigation of fecal samples of patients suffering from food poisoning or other diseases caused by cpe-positive C. perfringens. The method also offers a new approach in the investigation of the epidemiology of cpe-positive C. perfringens strains.

  9. Occurrence and characterization of toxigenic Bacillus cereus in food and infant feces

    Institute of Scientific and Technical Information of China (English)

    Sameer Rushdi Organji; Hussein Hasan Abulreesh; Khaled Elbanna; Gamal Ebrahim Haridy Osman; Manal Khider

    2015-01-01

    Objective:To investigate the true incidence of Bacillus cereus (B. cereus) in food and children diarrhea cases. Methods:A total of 110 samples of various dairy products such as raw milk, long life pasteurized milk, yoghurt and infant powdered milk formulas, raw rice, and feces were examined for the presence of B. cereus by selective plating on mannitol-egg-yolk-polymyxin agar. Confirmation of B. cereus was carried out by biochemical tests and PCR. Identification of non-B. cereus isolates was carried out by 16S rDNA sequencing. Antimicrobial susceptibility was done by disk diffusion method. Results:Overall 35 samples (31.8%, n=110) yielded Bacillus-like growth. Of which 19 samples (54.28%) were positive for B. cereus. All isolates were positive for enterotoxin production. No psychrotolerant B. cereus strains were detected in all samples. All B. cereus isolates were resistant to penicillin G, but susceptible to vancomycin, erythromycin and clindamycin. Conclusions:The results of this study confirm the importance of including B. cereus in disease control and prevention programs, as well as in routine clinical and food quality control laboratories in both Saudi Arabia and Egypt.

  10. Sensitive Procedure for Rapid Detection of Human Brucellosis, Based on PCR Method in Contaminated Serum Samples

    Directory of Open Access Journals (Sweden)

    Eslam Ghezelsofla

    2013-07-01

    Full Text Available AbstractBackground and objective: Brucellosis is a zoonosis transmittable to humans poses a significant public health problem in many developing countries and requires rapid and accurate diagnostic methods. Here, our aim was to develop a diagnostic polymerase chain reaction (PCR assay in artificially contaminated serum samples as a model for rapid and accurate laboratory confirmation of human brucellosis. Material and methods: In this study, initially the standard Brucella abortus strain (2308 were cultured on Brucella agar medium and then colonies were inactivated by formalin 10 %. Genomic DNA was extracted from inactivated bacterial colonies. Serial dilutions of bacterial-DNA were prepared in fetal bovine serum (FBS and water and subsequently DNA extraction were repeated on these artificially contaminated samples. The two pairs of primers amplified two different fragments included in: a gene encoding an outer membrane protein (omp-2 (primers JPF/JPR and a sequence 16S rRNA of B. abortus (primers F4/R2. Results: The two primers assayed showed a difference in sensitivity for detecting Brucella DNA, ranging between 5 pg and 50 pg for artificially contaminated serum samples and 50Fg and 5 pg for contaminated control samples. Therefore, the sensitivity of PCR using F4/R2 primers was greater than the PCR using JPF/JPR primers.Conclusion: Although the sensitivity of PCR using these primers was affected by serum inhibitors, they are still the most sensitive and they could provide a useful tool for the diagnosis of human brucellosis.

  11. Impact of Sample Type and DNA Isolation Procedure on Genomic Inference of Microbiome Composition

    DEFF Research Database (Denmark)

    Knudsen, Berith Elkær; Bergmark, Lasse; Munk, Patrick

    2016-01-01

    resistance from different reservoirs. Here, we compare in a stepwise approach a total of eight commercially available DNA extraction kits and 16 procedures based on these for three specimentypes (human feces, pig feces, and hospital sewage). We assess DNA extraction using spike-in controls and different...... types of beads for bead beating, facilitating cell lysis. We evaluate DNA concentration, purity, and stability and microbial community composition using 16S rRNA gene sequencing and for selected samples using shotgun metagenomic sequencing. Our results suggest that inferred community composition...... was dependent on inherent specimen properties as well as DNA extraction method. We further show that bead beating or enzymatic treatment can increase the extraction of DNA from Gram-positive bacteria. Final DNA quantities could be increased by isolating DNA from a larger volume of cell lysate than...

  12. A percutaneous needle biopsy technique for sampling the supraclavicular brown adipose tissue depot of humans.

    Science.gov (United States)

    Chondronikola, M; Annamalai, P; Chao, T; Porter, C; Saraf, M K; Cesani, F; Sidossis, L S

    2015-10-01

    Brown adipose tissue (BAT) has been proposed as a potential target tissue against obesity and its related metabolic complications. Although the molecular and functional characteristics of BAT have been intensively studied in rodents, only a few studies have used human BAT specimens due to the difficulty of sampling human BAT deposits. We established a novel positron emission tomography and computed tomography-guided Bergström needle biopsy technique to acquire human BAT specimens from the supraclavicular area in human subjects. Forty-three biopsies were performed on 23 participants. The procedure was tolerated well by the majority of participants. No major complications were noted. Numbness (9.6%) and hematoma (2.3%) were the two minor complications noted, which fully resolved. Thus, the proposed biopsy technique can be considered safe with only minimal risk of adverse events. Adoption of the proposed method is expected to increase the sampling of the supraclavicular BAT depot for research purposes so as to augment the scientific knowledge of the biology of human BAT.

  13. [Detection of human parvovirus B19, human bocavirus and human parvovirus 4 infections in blood samples among 95 patients with liver disease in Nanjing by nested PCR].

    Science.gov (United States)

    Tong, Rui; Zhou, Wei-Min; Liu, Xi-Jun; Wang, Yue; Lou, Yong-Liang; Tan, Wen-Jie

    2013-04-01

    To analyze the infection of human parvovirus B19, human bocavirus (HBoV) and human parvovirus 4 (PARV4) in blood samples among patients with liver disease in Nanjing by molecular detection. Nested PCR assays were designed and validated to detect B19, HBoV and PARV4, respectively. The assays were used to screen three parvoviruses in blood samples from 95 patients with different liver disease in Nanjing. The parvovirus infection was analyzed statistically. The detection limits were 10 copies of genomic DNA equivalents per reaction for each assays and the good specificity were observed. The frequency of B19 and HBoV were 2/95 (2.1%) and 9/95 (9.5%) in blood samples respectively. No PARV4 was detected. HBoV was detected in 3/5 patients with drug-induced hepatitis. Both B19 and HBoV infection were detected in blood from patients with liver disease.

  14. 50 CFR 23.16 - What are the U.S. CITES requirements for urine, feces, and synthetically derived DNA?

    Science.gov (United States)

    2010-10-01

    ... urine, feces, and synthetically derived DNA? 23.16 Section 23.16 Wildlife and Fisheries UNITED STATES... Requirements § 23.16 What are the U.S. CITES requirements for urine, feces, and synthetically derived DNA? (a) CITES documents. We do not require CITES documents to trade in urine, feces, or synthetically...

  15. Antigenic typing of brazilian rabies virus samples isolated from animals and humans, 1989-2000

    Directory of Open Access Journals (Sweden)

    FAVORETTO Silvana Regina

    2002-01-01

    Full Text Available Animal and human rabies samples isolated between 1989 and 2000 were typified by means of a monoclonal antibody panel against the viral nucleoprotein. The panel had been previously established to study the molecular epidemiology of rabies virus in the Americas. Samples were isolated in the Diagnostic Laboratory of the Pasteur Institute and in other rabies diagnostic centers in Brazil. In addition to the fixed virus samples CVS-31/96-IP, preserved in mouse brain, and PV-BHK/97, preserved in cell culture, a total of 330 rabies virus samples were isolated from dogs, cats, cattle, horses, bats, sheep, goat, swine, foxes, marmosets, coati and humans. Six antigenic variants that were compatible with the pre-established monoclonal antibodies panel were defined: numbers 2 (dog, 3 (Desmodus rotundus, 4 (Tadarida brasiliensis, 5 (vampire bat from Venezuela, 6 (Lasiurus cinereus and Lab (reacted to all used antibodies. Six unknown profiles, not compatible with the panel, were also found. Samples isolated from insectivore bats showed the greatest variability and the most commonly isolated variant was variant-3 (Desmodus rotundus. These findings may be related to the existence of multiple independent transmission cycles, involving different bat species.

  16. Comparison of chlorzoxazone one-sample methods to estimate CYP2E1 activity in humans

    DEFF Research Database (Denmark)

    Kramer, Iza; Dalhoff, Kim; Clemmesen, Jens O

    2003-01-01

    OBJECTIVE: Comparison of a one-sample with a multi-sample method (the metabolic fractional clearance) to estimate CYP2E1 activity in humans. METHODS: Healthy, male Caucasians ( n=19) were included. The multi-sample fractional clearance (Cl(fe)) of chlorzoxazone was compared with one...... estimates, Cl(est) at 3 h or 6 h, and MR at 3 h, can serve as reliable markers of CYP2E1 activity. The one-sample clearance method is an accurate, renal function-independent measure of the intrinsic activity; it is simple to use and easily applicable to humans.......-time-point clearance estimation (Cl(est)) at 3, 4, 5 and 6 h. Furthermore, the metabolite/drug ratios (MRs) estimated from one-time-point samples at 1, 2, 3, 4, 5 and 6 h were compared with Cl(fe). RESULTS: The concordance between Cl(est) and Cl(fe) was highest at 6 h. The minimal mean prediction error (MPE) of Cl...

  17. Selection of Candidate Housekeeping Genes for Normalization in Human Postmortem Brain Samples

    Directory of Open Access Journals (Sweden)

    Aldo Pagano

    2011-08-01

    Full Text Available The most frequently used technique to study the expression profile of genes involved in common neurological disorders is quantitative real-time RT-PCR, which allows the indirect detection of very low amounts of selected mRNAs in tissue samples. Expression analysis by RT-qPCR requires an appropriate normalization to the expression level of genes characterized by a stable, constitutive transcription. However, the identification of a gene transcribed at a very stable level is difficult if not impossible, since significant fluctuations of the level of mRNA synthesis often accompanies changes of cell behavior. The aim of this study is to identify the most stable genes in postmortem human brain samples of patients affected by Alzheimer’s disease (AD suitable as reference genes. The experiments analyzed 12 commonly used reference genes in brain samples from eight individuals with AD and seven controls. After a careful analysis of the results calculated by geNorm and NormFinder algorithms, we found that CYC1 and EIF4A2 are the best reference genes. We remark on the importance of the determination of the best reference genes for each sample to be analyzed and suggest a practical combination of reference genes to be used in the analysis of human postmortem samples.

  18. Quantitative chiral and achiral determination of ketamine and its metabolites by LC-MS/MS in human serum, urine and fecal samples.

    Science.gov (United States)

    Hasan, Mahmoud; Hofstetter, Robert; Fassauer, Georg M; Link, Andreas; Siegmund, Werner; Oswald, Stefan

    2017-05-30

    Ketamine (KET) is a widely used anesthetic drug which is metabolized by CYP450 enzymes to norketamine (n-KET), dehydronorketamine (DHNK), hydroxynorketamine (HNK) and hydroxyketamine (HK). Ketamine is a chiral compound and S-ketamine is known to be the more potent enantiomer. Here, we present the development and validation of three LC-MS/MS assays; the first for the quantification of racemic KET, n-KET and DHNK in human serum, urine and feces; the second for the separation and quantification of the S- and R-enantiomers of KET, n-KET and DHNK, and the third for separation and quantification of 2S,6S-hydroxynorketamine (2S,6S-HNK) and 2R,6R-hydroxynorketamine (2R,6R-HNK) in serum and urine with the ability to separate and detect 10 additional hydroxylated norketamine metabolites of racemic ketamine. Sample preparation was done by liquid-liquid extraction using methyl tert-butyl ether. For achiral determination of KET and its metabolites, an isocratic elution with ammonium acetate (pH 3.8; 5mM) and acetonitrile on a C18 column was performed. For the separation of S- and R-enantiomers of KET, n-KET and DHNK, a gradient elution was applied using a mobile phase of ammonium acetate (pH 7.5; 10mM) and isopropanol on the CHIRAL-AGP(®) column. The enantioselective separation of the HNK metabolites was done on the chiral column Lux(®)-Amylose-2 with a gradient method using ammonium acetate (pH 9; 5mM) and a mixture of isopropanol and acetonitrile (4:1). The mass spectrometric detection monitored for each analyte 2-3 mass/charge transitions. D4-ketamine and D4-n-KET were used as internal standards. The assays were successfully validated according to current bioanalytical guidelines and applied to a pilot study in one healthy volunteer. Compared to previously published methods, our assays have superior analytical features such as a lower amount of required matrix, faster sample preparation, shorter analytical run time and higher sensitivity (LLOQ up to 0.1ng/ml). Moreover

  19. Quality assesment for the analysis of PCDDs/PCDFs in individual human serum samples

    Energy Technology Data Exchange (ETDEWEB)

    Perez, F. [IIQAB-CSIC, Barcelona (Spain). Dept. of Ecotechnologies, Lab. of Dioxins; Abad, E.; Llerena, J.J.; Caixach, J.; Rivera, J.

    2004-09-15

    The aim of this work was to optimise a relevant methodology for the ultratrace analysis of PCDDs/PCDFs in individual human serum samples. In order to carry out the study, different strategies including the elaboration of quality control samples, parallel sample analysis, control blanks and a number of quality assurance measures were implemented as analytical current practices. Some of the main drawbacks in the analysis of PCDDs/PCDFs in these kind of samples come from two conflicting aspects: the small sample size and the low levels expected to be found. Taking this into account, an unavoidable compromise between the sample amount and the minimum analytical requirements, mainly the detection limit (LOD), is mandatory. To reach this goal C{sub 18} solid phase extraction was used to remove the analytes from the matrix. Clean up was performed by solid-liquid adsorption chromatography using a variety of adsorbents. Instrumental analysis was achieved by high-resolution gas chromatography coupled to high-resolution mass spectrometry (HRGC/HRMS). Finally, the optimised methodology was applied to evaluate the potential impact in general population living in the surroundings of an obsolete municipal waste incinerator plant (MWI). Thus, more than 400 individuals serum samples potentially exposed to the emission of the incinerator and people not exposed were considered in this study.

  20. Barriers to acceptance of self-sampling for human papillomavirus across ethnolinguistic groups of women.

    Science.gov (United States)

    Howard, Michelle; Lytwyn, Alice; Lohfeld, Lynne; Redwood-Campbell, Lynda; Fowler, Nancy; Karwalajtys, Tina

    2009-01-01

    Immigrant and low socio-economic (SES) women in North America underutilize Papanicolaou screening. Vaginal swab self-sampling for oncogenic human papillomavirus (HPV) has the potential to increase cervical cancer screening participation. The purpose of this qualitative study was to understand the perceptions of lower SES and immigrant women regarding self-sampling for HPV. Eleven focus-group interviews were conducted: one with Canadian-born English-speaking lower SES women, and two groups each with Arabic, Cantonese, Dari (Afghani), Somali and Spanish (Latino)-speaking women (one group conducted in English, the other in the native language) recently immigrated to Canada. Five to nine women aged 35 to 65 years and married with children participated in each group. Themes included 1) who might use self-sampling and why; 2) aversion to self-sampling and reasons to prefer physician; 3) ways to improve the appeal of self-sampling. Women generally perceived benefits of self-sampling and a small number felt they might use the method, but all groups had some reservations. Reasons included: uncertainty over performing the sampling correctly; fear of hurting themselves; concern about obtaining appropriate material; and concerns about test accuracy. Women preferred testing by a health care professional because they were accustomed to pelvic examinations, it was more convenient, or they trusted the results. Perceptions of self-sampling for HPV were similar across cultures and pertained to issues of confidence in self-sampling and need for physician involvement in care. These findings can inform programs and studies planning to employ self-sampling as a screening modality for cervical cancer.

  1. Genomic studies of envelope gene sequences from mosquito and human samples from Bangkok, Thailand.

    Science.gov (United States)

    Pitaksajjakul, Pannamthip; Benjathummarak, Surachet; Son, Hyun Ngoc; Thongrungkiat, Supatra; Ramasoota, Pongrama

    2016-01-01

    Dengue virus (DENV) is an RNA virus showing a high degree of genetic variation as a consequence of its proofreading inability. This variation plays an important role in virus evolution and pathogenesis. Although levels of within-host genetic variation are similar following equilibrium, variation among different hosts is frequently different. To identify dengue quasispecies present among two hosts, we collected patient samples from six acute DENV cases and two pools of Aedes aegypti mosquitoes and analyzed the genetic variation of regions of the viral envelope gene. Among human and mosquito samples, we found three major clusters originating from two subpopulations. Although several shared lineages were observed in the two hosts, only one lineage showing evidence of neutral selection was observed among two hosts. Taken together, our data provide evidence for the existence of a DENV quasispecies, with less genetic variation observed in mosquitoes than humans and with circulating lineages found in both host types.

  2. Quantitative detection of fecal contamination with domestic poultry feces in environments in China.

    Science.gov (United States)

    Zhuang, Fang-Fang; Li, Hu; Zhou, Xin-Yuan; Zhu, Yong-Guan; Su, Jian-Qiang

    2017-12-01

    Poultry are an important source of fecal contamination in environments. However, tools for detecting and tracking this fecal contamination are in the early stages of development. In practice, we have found that source tracking methods targeting the 16S rRNA genes of poultry-specific microbiota are not sufficiently sensitive. We therefore developed two quantitative PCR assays for detection of poultry fecal contamination, by targeting chicken and duck mitochondrial genes: NADH dehydrogenase subunit 5 (ND5) and cytochrome b (cytb). The sensitivity of both assays was 100% when tested on 50 chicken and duck fecal samples from 10 provinces of China. These assays were also tested in field samples, including soil and water collected adjacent to duck farms, and soils fertilized with chicken manure. Poultry mitochondrial DNA was detected in most of these samples, indicating that the assays are a robust method for monitoring environmental contamination with poultry feces. Complemented with existing indicators of fecal contamination, these markers should improve the efficiency and accuracy of microbial source tracking.

  3. Characterization of Staphylococcus aureus in Goose Feces from State Parks in Northeast Ohio.

    Science.gov (United States)

    Thapaliya, Dipendra; Dalman, Mark; Kadariya, Jhalka; Little, Katie; Mansell, Victoria; Taha, Mohammed Y; Grenier, Dylan; Smith, Tara C

    2017-03-10

    Staphylococcus aureus can colonize a range of species. Although numerous studies have isolated pathogenic bacteria from wild birds, very little is known regarding S. aureus and their potential to spread methicillin-resistant (MRSA) strains. The objective of this study was to determine the presence and molecular characteristics of S. aureus in geese fecal samples collected from ten state parks across Northeast Ohio (NEO). A total of 182 fecal samples from Canada geese (Branta canadensis) were collected in April 2015. Isolates were characterized using multi-locus sequence (MLST) and spa typing, as well as PCR to detect the presence of Panton-Valentine leukocidin (PVL), mecA, and scn genes. Antibiotic susceptibility testing was done via Vitek-2 system. The overall contamination by S. aureus in fecal samples was 7.1% (13/182); 7/182 (3.8%) were MRSA and 6/182 (3.3%) were methicillin-susceptible S. aureus (MSSA). One isolate was positive for PVL. A total of eight different spa types were observed. MLST included ST5, ST8, ST291, ST298, and ST2111. One (7.7%) MSSA isolate was multi-drug resistant. The S. aureus contamination in NEO state parks ranged from 0% (park 1, 4, 8, 9) to 35% (7/20) (park 5). Parks 2, 3, 6, and 7 had 5% (1/20) positive. The results of this study indicate that the feces of geese collected at various state parks in NEO may harbor S. aureus.

  4. Load and failure behavior of human muscle samples in the context of proximal femur replacement

    OpenAIRE

    Schleifenbaum, Stefan; Schmidt, Michael; Möbius, Robert; Wolfskämpf, Thomas; Schröder, Christian; Grunert, Ronny; Hammer, Niels; Prietzel, Torsten

    2016-01-01

    Background: To ensure adequate function after orthopedic tumor reconstruction, it is important to reattach the remaining soft tissue to the implant. This study aimed at obtaining mechanical properties of textile muscle-implant and muscle-bone connections in a preliminary test. Methods: Two groups of soft-tissue attachment were mechanically tested and compared: Native bone-muscle samples obtained from human femora and muscles attached to a prosthetic implant by means of Trevira® attachment tu...

  5. Dioxin-like activity of environmental compounds in human blood and environmental samples

    DEFF Research Database (Denmark)

    Long, Manhai; Bonefeld-Jørgensen, Eva Cecilie

    2012-01-01

    R transactivation bioassay is utilized in an array of projects to study the AhR-mediated activities of individual chemicals and mixtures and for epidemiological purposes. This review summarizes a series of studies regarding the DL-activity of single compounds and complex compound mixtures in the environment...... a cost-effective and integrated screening tool for measurement of the DL-activity in human, environmental and commercial samples....

  6. Separation of proteins from human plasma by sample displacement chromatography in hydrophobic interaction mode

    OpenAIRE

    Josic, Djuro; Breen, Lucas; Clifton, James; Gajdosik, Martina Srajer; Gaso-Sokac, Dajana; Rucevic, Marijana; Müller, Egbert

    2012-01-01

    Sample displacement chromatography (SDC) in reversed-phase and ion-exchange modes was introduced approximately twenty years ago. This method was first used for the preparative purification of peptides and proteins. Recently, SDC in ion-exchange mode was also successfully used for enrichment of low abundance proteins from human plasma. In this paper, the use of SDC for the separation of plasma proteins in hydrophobic interaction mode is demonstrated. By use of two or more columns coupled in se...

  7. Same Exposure but Two Radically Different Responses to Antibiotics: Resilience of the Salivary Microbiome versus Long-Term Microbial Shifts in Feces.

    Science.gov (United States)

    Zaura, Egija; Brandt, Bernd W; Teixeira de Mattos, M Joost; Buijs, Mark J; Caspers, Martien P M; Rashid, Mamun-Ur; Weintraub, Andrej; Nord, Carl Erik; Savell, Ann; Hu, Yanmin; Coates, Antony R; Hubank, Mike; Spratt, David A; Wilson, Michael; Keijser, Bart J F; Crielaard, Wim

    2015-11-10

    Due to the spread of resistance, antibiotic exposure receives increasing attention. Ecological consequences for the different niches of individual microbiomes are, however, largely ignored. Here, we report the effects of widely used antibiotics (clindamycin, ciprofloxacin, amoxicillin, and minocycline) with different modes of action on the ecology of both the gut and the oral microbiomes in 66 healthy adults from the United Kingdom and Sweden in a two-center randomized placebo-controlled clinical trial. Feces and saliva were collected at baseline, immediately after exposure, and 1, 2, 4, and 12 months after administration of antibiotics or placebo. Sequences of 16S rRNA gene amplicons from all samples and metagenomic shotgun sequences from selected baseline and post-antibiotic-treatment sample pairs were analyzed. Additionally, metagenomic predictions based on 16S rRNA gene amplicon data were performed using PICRUSt. The salivary microbiome was found to be significantly more robust, whereas the antibiotics negatively affected the fecal microbiome: in particular, health-associated butyrate-producing species became strongly underrepresented. Additionally, exposure to different antibiotics enriched genes associated with antibiotic resistance. In conclusion, healthy individuals, exposed to a single antibiotic treatment, undergo considerable microbial shifts and enrichment in antibiotic resistance in their feces, while their salivary microbiome composition remains unexpectedly stable. The health-related consequences for the gut microbiome should increase the awareness of the individual risks involved with antibiotic use, especially in a (diseased) population with an already dysregulated microbiome. On the other hand, understanding the mechanisms behind the resilience of the oral microbiome toward ecological collapse might prove useful in combating microbial dysbiosis elsewhere in the body. Many health care professionals use antibiotic prophylaxis strategies to prevent

  8. Dioxin-like activity of environmental compounds in human blood and environmental samples

    DEFF Research Database (Denmark)

    Long, Manhai; Bonefeld-Jørgensen, Eva Cecilie

    2012-01-01

    and humans. We found that some pesticides, plasticizers and phytoestrogens can activate the AhR, and the combined effect of compounds with no or weak AhR potency cannot be ignored. The significant DL-activity in the wastewater effluent indicates the treatment is not sufficient to prevent contamination...... of surface waters with dioxins. Our results from human studies suggest that the serum DL-activity reflect the complex mixture of persistent organic pollutants (POPs). Greenlandic Inuit had lower serum DL-activity level compared to Europeans, probably due to long distance from the dioxin sources and UV...... a cost-effective and integrated screening tool for measurement of the DL-activity in human, environmental and commercial samples....

  9. Dioxin-like activity in environmental and human samples from Greenland and Denmark

    DEFF Research Database (Denmark)

    Long, Manhai; Bonefeld-Jørgensen, Eva Cecilie

    2012-01-01

    and humans. We found that some pesticides, plasticizers and phytoestrogens can activate the AhR, and the combined effect of compounds with no or weak AhR potency cannot be ignored. The significant DL-activity in the wastewater effluent indicates the treatment is not sufficient to prevent contamination...... of surface waters with dioxins. Our results from human studies suggest that the serum DL-activity reflect the complex mixture of persistent organic pollutants (POPs). Greenlandic Inuit had lower serum DL-activity level compared to Europeans, probably due to long distance from the dioxin sources and UV...... a cost-effective and integrated screening tool for measurement of the DL-activity in human, environmental and commercial samples....

  10. Least destructive sampling of human remains using laser drilling for Sr isotope analysis by TIMS

    Science.gov (United States)

    Willmes, Malte; Moffat, Ian; Grün, Rainer; Armstrong, Richard; Kinsley, Les; McMorrow, Linda

    2013-04-01

    Strontium isotope ratios (87Sr/86Sr) measured in ancient human remains can be used to reconstruct migration patterns of ancient human populations. This application is based on the fact that different geologic regions have distinct Sr isotope signatures that are cycled through the soils, plants and rivers, and eventually enter the food cycle. Sr isotope ratios measured in skeletal remains (bones and teeth) reflect the average of dietary Sr that was consumed when the tissue was formed, allowing the investigation of human migration between geologically distinct terrains. The analysis of human remains is always a sensitive topic requiring minimal damage to the sample, while at the same time providing highly precise and accurate results. Samples can be analysed either by solution methods like thermal ionisation mass spectrometry (TIMS), or by in-situ laser ablation MC-ICP-MS. For TIMS a drill is used to extract a small amount of sample, which is then digested in acid and Sr is separated out using ion exchange chromatography. This technique provides highly precise and accurate results, because any isobaric interferences are removed during chemical separation. The drawback is that drilling may cause visible damage to the sample, restricting access to precious human remains. LA-MC-ICP-MS analysis is very fast and nearly destruction free. However, the accuracy and precision of LA-MC-ICP-MS is limited by a number of factors including large instrumental mass discrimination, laser-induced isotopic and elemental fractionations and molecular interferences on 87Sr. Its application thus requires rigorous data reduction, which can introduce significant uncertainties into the analysis. This is especially true for samples with relatively low Sr concentrations such as human teeth (e.g., Woodhead et al., 2005; Horstwood et al., 2008; Vroon et al., 2008). In addition, LA-MC-ICP-MS has traditionally required a flat sample surface, thus an unbroken tooth needs to be cut, which is rather

  11. Laser induced breakdown spectroscopy of human liver samples with Wilson's disease

    Science.gov (United States)

    Grolmusová, Zuzana; Horňáčková, Michaela; Plavčan, Jozef; Kopáni, Martin; Babál, Pavel; Veis, Pavel

    2013-08-01

    Laser induced breakdown spectroscopy (LIBS) is an elemental analytical technique with various applications. The paper demonstrates the first LIBS measurements of human liver samples for the purpose of detecting the higher copper content related with the advanced stage of Wilson's disease. These measurements were implemented using a Nd:YAG laser working at the wavelength of 532 nm and an echelle type spectrometer equipped with an intensified CCD camera allowing for a wide spectral range coverage (200-950 nm) and rapid camera gating (minimum gating time of 5 ns). Seven liver samples with suspected Wilson's disease and five reference samples were investigated. The main parameter of interest was the Cu/C ratio obtained at first from spectra and secondly directly from an iCCD image. Our experiment is a pilot study, which shows LIBS analysis of human liver samples for the purpose of detecting the normal and higher copper content for the first time. The method proved to be a quick and a low-cost approach for the detection of pathological accumulation of copper in the affected tissue.

  12. DNA purification from crude samples for human identification using gradient elution isotachophoresis.

    Science.gov (United States)

    Strychalski, Elizabeth A; Konek, Christopher; Butts, Erica L R; Vallone, Peter M; Henry, Alyssa C; Ross, David

    2013-09-01

    Gradient elution isotachophoresis (GEITP) was demonstrated for DNA purification, concentration, and quantification from crude samples, represented here by soiled buccal swabs, with minimal sample preparation prior to human identification using STR analysis. During GEITP, an electric field applied across leading and trailing electrolyte solutions resulted in isotachophoretic focusing of DNA at the interface between these solutions, while a pressure-driven counterflow controlled the movement of the interface from the sample reservoir into a microfluidic capillary. This counterflow also prevented particulates from fouling or clogging the capillary and reduced or eliminated contamination of the delivered DNA by PCR inhibitors. On-line DNA quantification using laser-induced fluorescence compared favorably with quantitative PCR measurements and potentially eliminates the need for quantitative PCR prior to STR analysis. GEITP promises to address the need for a rapid and robust method to deliver DNA from crude samples to aid the forensic community in human identification. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Detection of Streptococcus mutans Genomic DNA in Human DNA Samples Extracted from Saliva and Blood

    Science.gov (United States)

    Vieira, Alexandre R.; Deeley, Kathleen B.; Callahan, Nicholas F.; Noel, Jacqueline B.; Anjomshoaa, Ida; Carricato, Wendy M.; Schulhof, Louise P.; DeSensi, Rebecca S.; Gandhi, Pooja; Resick, Judith M.; Brandon, Carla A.; Rozhon, Christopher; Patir, Asli; Yildirim, Mine; Poletta, Fernando A.; Mereb, Juan C.; Letra, Ariadne; Menezes, Renato; Wendell, Steven; Lopez-Camelo, Jorge S.; Castilla, Eduardo E.; Orioli, Iêda M.; Seymen, Figen; Weyant, Robert J.; Crout, Richard; McNeil, Daniel W.; Modesto, Adriana; Marazita, Mary L.

    2011-01-01

    Caries is a multifactorial disease, and studies aiming to unravel the factors modulating its etiology must consider all known predisposing factors. One major factor is bacterial colonization, and Streptococcus mutans is the main microorganism associated with the initiation of the disease. In our studies, we have access to DNA samples extracted from human saliva and blood. In this report, we tested a real-time PCR assay developed to detect copies of genomic DNA from Streptococcus mutans in 1,424 DNA samples from humans. Our results suggest that we can determine the presence of genomic DNA copies of Streptococcus mutans in both DNA samples from caries-free and caries-affected individuals. However, we were not able to detect the presence of genomic DNA copies of Streptococcus mutans in any DNA samples extracted from peripheral blood, which suggests the assay may not be sensitive enough for this goal. Values of the threshold cycle of the real-time PCR reaction correlate with higher levels of caries experience in children, but this correlation could not be detected for adults. PMID:21731912

  14. Novel genotype of Ehrlichia canis detected in samples of human blood bank donors in Costa Rica.

    Science.gov (United States)

    Bouza-Mora, Laura; Dolz, Gaby; Solórzano-Morales, Antony; Romero-Zuñiga, Juan José; Salazar-Sánchez, Lizbeth; Labruna, Marcelo B; Aguiar, Daniel M

    2017-01-01

    This study focuses on the detection and identification of DNA and antibodies to Ehrlichia spp. in samples of blood bank donors in Costa Rica using molecular and serological techniques. Presence of Ehrlichia canis was determined in 10 (3.6%) out of 280 blood samples using polymerase chain reaction (PCR) targeting the ehrlichial dsb conserved gene. Analysis of the ehrlichial trp36 polymorphic gene in these 10 samples revealed substantial polymorphism among the E. canis genotypes, including divergent tandem repeat sequences. Nucleotide sequences of dsb and trp36 amplicons revealed a novel genotype of E. canis in blood bank donors from Costa Rica. Indirect immunofluorescence assay (IFA) detected antibodies in 35 (35%) of 100 serum samples evaluated. Thirty samples showed low endpoint titers (64-256) to E. canis, whereas five sera yielded high endpoint titers (1024-8192); these five samples were also E. canis-PCR positive. These findings represent the first report of the presence of E. canis in humans in Central America.

  15. Pathogenic Vibrio Strains Isolated from Human Stool and Water Samples from Western Kenya

    Directory of Open Access Journals (Sweden)

    Roselida Achieng Owuor

    2016-03-01

    Full Text Available Objective: Investigate the type of pathogenic Vibrio strains from water and stool samples collected from Migori, SonduMiriu, Nyando and Yala regions in Western Kenya. Methods: A total of 811 samples (596 water and 215 stool samples were collected during the study periods of May to December 2013 and August to September 2014. Pathogenic Vibrio strains were identified through culturing in TCBS Agar, followed by oxidation, string and serological (polyvalent tests, respectively. The PCR analysis was done using combined primers targeting Vibrionaceae 16SrRNA and species specific primers for V. vulnificus and V. cholerae. Results: The results showed the presence of V. vulnificus and V. cholerae. However, V. parahaemolyticus was not found in any of the samples. The PCR results for 16SrRNA, Vib 1, and Vib 2 showed polymorphism in the genes, this was an indication of cross combination of genes from more than one strain in one isolate. Conclusion: The study showed the presence of V. cholerae (Ogawa and Inaba in water and human stool samples. Type B V. vulnificus was detected in the water sample collected from River Migori. This information is of essence in controlling and managing cholera in the western part of Kenya. J Microbiol Infect Dis 2016;6(1: 1-7

  16. Isolation and clinical sample typing of human leptospirosis cases in Argentina.

    Science.gov (United States)

    Chiani, Yosena; Jacob, Paulina; Varni, Vanina; Landolt, Noelia; Schmeling, María Fernanda; Pujato, Nazarena; Caimi, Karina; Vanasco, Bibiana

    2016-01-01

    Leptospira typing is carried out using isolated strains. Because of difficulties in obtaining them, direct identification of infective Leptospira in clinical samples is a high priority. Multilocus sequence typing (MLST) proved highly discriminatory for seven pathogenic species of Leptospira, allowing isolate characterization and robust assignment to species, in addition to phylogenetic evidence for the relatedness between species. In this study we characterized Leptospira strains circulating in Argentina, using typing methods applied to human clinical samples and isolates. Phylogenetic studies based on 16S ribosomal RNA gene sequences enabled typing of 8 isolates (6 Leptospira interrogans, one Leptospira wolffii and one Leptospira broomii) and 58 out of 85 (68.2%) clinical samples (55 L. interrogans, 2 Leptospira meyeri, and one Leptospira kirschneri). MLST results for the L. interrogans isolates indicated that five were probably Canicola serogroup (ST37) and one was probably Icterohaemorrhagiae serogroup (ST17). Eleven clinical samples (21.6%), provided MLST interpretable data: five were probably Pyrogenes serogroup (ST13), four Sejroe (ST20), one Autumnalis (ST22) and one Canicola (ST37). To the best of our knowledge this study is the first report of the use of an MLST typing scheme with seven loci to identify Leptospira directly from clinical samples in Argentina. The use of clinical samples presents the advantage of the possibility of knowing the infecting strain without resorting to isolates. This study also allowed, for the first time, the characterization of isolates of intermediate pathogenicity species (L. wolffii and L. broomii) from symptomatic patients.

  17. A single dose mass balance study of the Hedgehog pathway inhibitor vismodegib (GDC-0449) in humans using accelerator mass spectrometry.

    Science.gov (United States)

    Graham, Richard A; Lum, Bert L; Morrison, Glenn; Chang, Ilsung; Jorga, Karin; Dean, Brian; Shin, Young G; Yue, Qin; Mulder, Teresa; Malhi, Vikram; Xie, Minli; Low, Jennifer A; Hop, Cornelis E C A

    2011-08-01

    Vismodegib (GDC-0449), a small-molecule Hedgehog pathway inhibitor, was well tolerated in patients with solid tumors and showed promising efficacy in advanced basal cell carcinoma in a Phase I trial. The purpose of the study presented here was to determine routes of elimination and the extent of vismodegib metabolism, including assessment and identification of metabolites in plasma, urine, and feces. Six healthy female subjects of nonchildbearing potential were enrolled; each received a single 30-ml oral suspension containing 150 mg of vismodegib with 6.5 μg of [(14)C]vismodegib to yield a radioactivity dose of approximately 37 kBq (1000 nCi). Plasma, urine, and feces samples were collected over 56 days to permit sample collection for up to 5 elimination half-lives. Nonradioactive vismodegib was measured in plasma using liquid chromatographic-tandem mass spectrometry, and total radioactivity in plasma, urine, and feces was measured using accelerator mass spectrometry. Vismodegib was slowly eliminated by a combination of metabolism and excretion of parent drug, most of which was recovered in feces. The estimated excretion of the administered dose was 86.6% on average, with 82.2 and 4.43% recovered in feces and urine, respectively. Vismodegib was predominant in plasma, with concentrations representing >98% of the total circulating drug-related components. Metabolic pathways of vismodegib in humans included oxidation, glucuronidation, and uncommon pyridine ring cleavage. We conclude that vismodegib and any associated metabolic products are mainly eliminated through feces after oral administration in healthy volunteers.

  18. Development of Bacteroides 16S rRNA Gene TaqMan-Based Real-Time PCR Assays for Estimation of Total, Human, and Bovine Fecal Pollution in Water

    Science.gov (United States)

    Layton, Alice; McKay, Larry; Williams, Dan; Garrett, Victoria; Gentry, Randall; Sayler, Gary

    2006-01-01

    Bacteroides species are promising indicators for differentiating livestock and human fecal contamination in water because of their high concentration in feces and potential host specificity. In this study, a real-time PCR assay was designed to target Bacteroides species (AllBac) present in human, cattle, and equine feces. Direct PCR amplification (without DNA extraction) using the AllBac assay was tested on feces diluted in water. Fecal concentrations and threshold cycle were linearly correlated, indicating that the AllBac assay can be used to estimate the total amount of fecal contamination in water. Real-time PCR assays were also designed for bovine-associated (BoBac) and human-associated (HuBac) Bacteroides 16S rRNA genes. Assay specificities were tested using human, bovine, swine, canine, and equine fecal samples. The BoBac assay was specific for bovine fecal samples (100% true-positive identification; 0% false-positive identification). The HuBac assay had a 100% true-positive identification, but it also had a 32% false-positive rate with potential for cross-amplification with swine feces. The assays were tested using creek water samples from three different watersheds. Creek water did not inhibit PCR, and results from the AllBac assay were correlated with those from Escherichia coli concentrations (r2 = 0.85). The percentage of feces attributable to bovine and human sources was determined for each sample by comparing the values obtained from the BoBac and HuBac assays with that from the AllBac assay. These results suggest that real-time PCR assays without DNA extraction can be used to quantify fecal concentrations and provide preliminary fecal source identification in watersheds. PMID:16751534

  19. Dioxin and PCB levels in human samples from the Greek population

    Energy Technology Data Exchange (ETDEWEB)

    Leondiadis, L.; Vassiliadou, I.; Costopoulou, D.; Papadopoulos, A. [Mass Spectrometry and Dioxin Analysis Lab. - NCSR Demokritos, Athens (Greece)

    2004-09-15

    Polychlorinated biphenyls (PCBs) are commercial chemical substances produced in a large scale since 1930, with a wide range of applications in industry, such as for coolant fluids in transformers and dielectric fluids in capacitors. After their health effects became apparent, PCB production was banned in the late 1970s. However, humans are still exposed through PCB leakage of old capacitors and transformers and disposal of contaminated materials. Dioxins (polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzo-furans (PCDFs)), are formed as undesirable by-products mainly during the production of chlorinated chemicals and during the combustion of municipal and hazardous waste. Due to potential health hazard (dermal toxicity, immunotoxicity, reproductive effects, teratogenicity, endocrine disruption and carcinogenicity), their monitoring in humans is of high general concern. Enough information on POP presence in human tissues from industrialized countries is available to suggest that the concentration of these compounds has decreased during the last 10 years. Monitoring of human exposure to PCBs and dioxins, contaminants that accumulate in lipid tissue, is most conveniently performed by analysis of blood plasma or blood serum. Monitoring of dioxins in human milk is of also great importance, since it is especially feared that lactational exposure to dioxins and related compounds may adversely affect brain development and the immune system of infants and children. The present study includes the analyses of non-ortho, mono-ortho, indicator PCBs, and PCDD/Fs in human blood and human milk samples collected between November 2002 and February 2004 and is the first study of this kind to be undertaken in Greece.

  20. Genetic identification of missing persons: DNA analysis of human remains and compromised samples.

    Science.gov (United States)

    Alvarez-Cubero, M J; Saiz, M; Martinez-Gonzalez, L J; Alvarez, J C; Eisenberg, A J; Budowle, B; Lorente, J A

    2012-01-01

    Human identification has made great strides over the past 2 decades due to the advent of DNA typing. Forensic DNA typing provides genetic data from a variety of materials and individuals, and is applied to many important issues that confront society. Part of the success of DNA typing is the generation of DNA databases to help identify missing persons and to develop investigative leads to assist law enforcement. DNA databases house DNA profiles from convicted felons (and in some jurisdictions arrestees), forensic evidence, human remains, and direct and family reference samples of missing persons. These databases are essential tools, which are becoming quite large (for example the US Database contains 10 million profiles). The scientific, governmental and private communities continue to work together to standardize genetic markers for more effective worldwide data sharing, to develop and validate robust DNA typing kits that contain the reagents necessary to type core identity genetic markers, to develop technologies that facilitate a number of analytical processes and to develop policies to make human identity testing more effective. Indeed, DNA typing is integral to resolving a number of serious criminal and civil concerns, such as solving missing person cases and identifying victims of mass disasters and children who may have been victims of human trafficking, and provides information for historical studies. As more refined capabilities are still required, novel approaches are being sought, such as genetic testing by next-generation sequencing, mass spectrometry, chip arrays and pyrosequencing. Single nucleotide polymorphisms offer the potential to analyze severely compromised biological samples, to determine the facial phenotype of decomposed human remains and to predict the bioancestry of individuals, a new focus in analyzing this type of markers.

  1. Evaluation of the ELISA test for detection of Entamoeba histolytica in feces.

    Science.gov (United States)

    Merino, E; Glender, W; del Muro, R; Ortiz-Ortiz, L

    1990-01-01

    The clinical utility of an ELISA test with monoclonal antibodies to detect antigen of Entamoeba histolytica in feces was evaluated in 150 patients with gastrointestinal symptoms. Each subject was examined by rectosigmoidoscopy with rectal smear and/or a triple stool search for ova-bacteria-parasite (OBP); in addition, one stool sample was collected for the ELISA test. All the tests were independent and double blind. E. histolytica was detected by OBP and/or rectosigmoidoscopy in 66 patients; 61 patients had other parasites; and in 23, no parasites were identified. Of all patients, 116 were positive for the ELISA test. Of these, E. histolytica was identified in 52. In 47, other parasites were identified and in 17, no parasites were found. The ELISA test with a monoclonal antibody against E. histolytica antigen showed higher sensitivity than the standard diagnostic methods: the ability to detect the presence of E. histolytica antigen regardless of the destruction of the parasite or of the error due to misidentification of the parasite resulting from faulty preparation of the samples.

  2. The use of quantitative PCR for identification and quantification of Brachyspira pilosicoli, Lawsonia intracellularis and Escherichia coli fimbrial types F4 and F18 in pig feces

    DEFF Research Database (Denmark)

    Ståhl, Marie; Kokotovic, Branko; Hjulsager, Charlotte Kristiane

    2011-01-01

    than the earlier used method due to improvements in DNA extraction. In addition, as samples were not analysed for all four pathogen agents by traditional diagnostic methods, many samples were found positive for agents that were not expected on the basis of age and case history. The use of quantitative...... the spiking experiments were 102 bacteria/g feces for BpiloqPCR and Laws-qPCR, 103 CFU/g feces for F4-qPCR and F18-qPCR. The PCR efficiency for all four qPCR assays was between 0.91 and 1.01 with R2 above 0.993. Standard curves, slopes and elevation, varied between assays and between measurements from pure...... DNA from reference strains and feces spiked with the respective strains. The linear ranges found for spiked fecal samples differed both from the linear ranges from pure culture of the reference strains and between the qPCR tests. The linear ranges were five log units for F4- qPCR, and Laws-qPCR, six...

  3. A simple sample preparation method for measuring amoxicillin in human plasma by hollow fiber centrifugal ultrafiltration.

    Science.gov (United States)

    Dong, Wei-Chong; Hou, Zi-Li; Jiang, Xin-Hui; Jiang, Ye

    2013-02-01

    A simple sample preparation method has been developed for the determination of amoxicillin in human plasma by hollow fiber centrifugal ultrafiltration (HF-CF-UF). A 400-μL plasma sample was placed directly into the HF-CF-UF device, which consisited of a slim glass tube and a U-shaped hollow fiber. After centrifugation at 1.25 × 10(3) g for 10 min, the filtrate was withdrawn from the hollow fiber and 20 µL was directly injected into the high-performance liquid chromatography (HPLC) for analysis. The calibration curve was linear over the range of 0.1-20 µg/mL (r = 0.9996) and the limit of detection was as low as 0.025 µg/mL. The average recovery and absolute recovery were 99.9% and 84.5%, respectively. Both the intra-day and inter-day precisions (relative standard deviation) were less than 3.1% for three concentrations (0.25, 2.5 and 10 µg/mL). The sample preparation process was simplified. Only after a single centrifugal ultrafiltration can the filtrate be injected directly into HPLC. The present method is simple, sensitive and accurate. It could be effective for the analysis of biological samples with high protein contents, especially for the biopharmaceutical analysis of drugs that use traditional isolation techniques for sample preparation such as the protein precipitation method.

  4. Small Sample Kernel Association Tests for Human Genetic and Microbiome Association Studies.

    Science.gov (United States)

    Chen, Jun; Chen, Wenan; Zhao, Ni; Wu, Michael C; Schaid, Daniel J

    2016-01-01

    Kernel machine based association tests (KAT) have been increasingly used in testing the association between an outcome and a set of biological measurements due to its power to combine multiple weak signals of complex relationship with the outcome through the specification of a relevant kernel. Human genetic and microbiome association studies are two important applications of KAT. However, the classic KAT framework relies on large sample theory, and conservativeness has been observed for small sample studies, especially for microbiome association studies. The common approach for addressing the small sample problem relies on computationally intensive resampling methods. Here, we derive an exact test for KAT with continuous traits, which resolve the small sample conservatism of KAT without the need for resampling. The exact test has significantly improved power to detect association for microbiome studies. For binary traits, we propose a similar approximate test, and we show that the approximate test is very powerful for a wide range of kernels including common variant- and microbiome-based kernels, and the approximate test controls the type I error well for these kernels. In contrast, the sequence kernel association tests have slightly inflated genomic inflation factors after small sample adjustment. Extensive simulations and application to a real microbiome association study are used to demonstrate the utility of our method. © 2015 WILEY PERIODICALS, INC.

  5. A salting out and resin procedure for extracting Schistosoma mansoni DNA from human urine samples

    Directory of Open Access Journals (Sweden)

    Rodrigues Nilton B

    2010-04-01

    Full Text Available Abstract Background In this paper a simple and cheap salting out and resin (InstaGene matrix® resin - BioRad DNA extraction method from urine for PCR assays is introduced. The DNA of the fluke Schistosoma mansoni was chosen as the target since schistosomiasis lacks a suitable diagnostic tool which is sensitive enough to detect low worm burden. It is well known that the PCR technique provides high sensitivity and specificity in detecting parasite DNA. Therefore it is of paramount importance to take advantage of its excellent performance by providing a simple to handle and reliable DNA extraction procedure, which permits the diagnosis of the disease in easily obtainable urine samples. Findings The description of the extraction procedure is given. This extraction procedure was tested for reproducibility and efficiency in artificially contaminated human urine samples. The reproducibility reached 100%, showing positive results in 5 assay repetitions of 5 tested samples each containing 20 ng DNA/5 ml. The efficiency of the extraction procedure was also evaluated in a serial dilution of the original 20 ng DNA/5 ml sample. Detectable DNA was extracted when it was at a concentration of 1.28 pg DNA/mL, revealing the high efficiency of this procedure. Conclusions This methodology represents a promising tool for schistosomiasis diagnosis utilizing a bio-molecular technique in urine samples which is now ready to be tested under field conditions and may be applicable to the diagnosis of other parasitic diseases.

  6. Potential hazard of zoonotic parasites present in canine feces in Puerto Escondido, Oaxaca

    Directory of Open Access Journals (Sweden)

    León Vélez-Hernández

    2014-11-01

    Full Text Available Objective. To estimate the zoonotic parasites prevalence in feral dog feces in Puerto Escondido. Material and methods. The fecalism frecuency was estimated in ten zones. To identify the parasites parasitological flotation and direct smear methods were used. The parasitic prevalence was estimated in the canine feces. Results. All the zones presented canine fecalism. The parasitic prevalence in the feces was 73.33%. The parasites with the highest prevalence were Toxocara canis (47.78%, Ancylostoma caninum (17.88%, and Dipylidium caninum (13.89%. Conclusion. Canine fecalism comes from strayed and owned dogs. 66.66% of the parasites found in the dog feces are zoonotics. The factors associated to this problem are the suburban habitat, waste mishandling and nil tenure of stray dogs.

  7. Determination of total (IgG/IgM and specific (IgM antibodies to Hepatitis E Virus and molecular detection of the virus in feces of humans with or without occupational exposure to pigs in 10 municipalities of Antioquia, Colombia

    Directory of Open Access Journals (Sweden)

    Gutiérrez-Vergara, Cristian Camilo

    2015-07-01

    Full Text Available In 10 municipalities of Antioquia (Colombia the positivity rate in serum for total (IgG/IgM and specific (IgM antibodies to hepatitis E virus (HEV was determined, and tests were done for the presence of HEV RNA in the feces of individuals positive for IgM antibodies. According to previous exposure to pigs, two different groups were included, namely: exposed and unexposed. The latter group was subdivided into cohabitants of the exposed ones and general population. The frequency of total anti-HEV antibodies in the exposed group was 15.7%, and that of IgM, 2.5% (p<0.001. In the group of cohabitants, total antibodies were found in 5.9%, while IgM antibodies were not present. In the general population IgG/IgM antibodies were present in 7.2% and IgM, in 0.81% (p<0.001. None of the fecal specimens was positive for HEV RNA. These results indicate that individuals with occupational exposure to pigs have higher risk (RP: 2.42 of being positive for anti-HEV antibodies than the unexposed ones (95% CI: 1.66-3.53 (p<0.001. Also, that in Antioquia HEV is present regardless of the exposure to pigs. Further studies on HEV in Colombia should be done.

  8. Recognition of serous ovarian tumors in human samples by multimodal nonlinear optical microscopy

    Science.gov (United States)

    Adur, Javier; Pelegati, Vitor B.; Costa, Leverson F. L.; Pietro, Luciana; de Thomaz, Andre A.; Almeida, Diogo B.; Bottcher-Luiz, Fatima; Andrade, Liliana A. L. A.; Cesar, Carlos L.

    2011-09-01

    We used a multimodal nonlinear optics microscopy, specifically two-photon excited fluorescence (TPEF), second and third harmonic generation (SHG/THG) microscopies, to observe pathological conditions of ovarian tissues obtained from human samples. We show that strong TPEF + SHG + THG signals can be obtained in fixed samples stained with hematoxylin and eosin (H&E) stored for a very long time, and that H&E staining enhanced the THG signal. We then used the multimodal TPEF-SHG-THG microscopies in a stored file of H&E stained samples of human ovarian cancer to obtain complementary information about the epithelium/stromal interface, such as the transformation of epithelium surface (THG) and the overall fibrillary tissue architecture (SHG). This multicontrast nonlinear optics microscopy is able to not only differentiate between cancerous and healthy tissue, but can also distinguish between normal, benign, borderline, and malignant specimens according to their collagen disposition and compression levels within the extracellular matrix. The dimensions of the layers of epithelia can also be measured precisely and automatically. Our data demonstrate that optical techniques can detect pathological changes associated with ovarian cancer.

  9. Exploring the acceptability of human papillomavirus self-sampling among Muslim immigrant women.

    Science.gov (United States)

    Lofters, Aisha K; Vahabi, Mandana; Fardad, Mitra; Raza, Afrah

    2017-01-01

    With appropriate screening (ie, the Papanicolaou [Pap] test), cervical cancer is highly preventable, and high-income countries, including Canada, have observed significant decreases in cervical cancer mortality. However, certain subgroups, including immigrants from countries with large Muslim populations, experience disparities in cervical cancer screening. Little is known about the acceptability of human papillomavirus (HPV) self-sampling as a screening strategy among Muslim immigrant women in Canada. This study assessed cervical cancer screening practices, knowledge and attitudes, and acceptability of HPV self-sampling among Muslim immigrant women. A convenience sample of 30 women was recruited over a 3-month period (June-August 2015) in the Greater Toronto Area. All women were between 21 and 69 years old, foreign-born, and self-identified as Muslim, and had good knowledge of English. Data were collected through a self-completed questionnaire. More than half of the participants falsely indicated that Pap tests may cause cervical infection, and 46.7% indicated that the test is an intrusion on privacy. The majority of women reported that they would be willing to try HPV self-sampling, and more than half would prefer this method to provider-administered sampling methods. Barriers to self-sampling included confidence in the ability to perform the test and perceived cost, and facilitators included convenience and privacy being preserved. The results demonstrate that HPV self-sampling may provide a favorable alternative model of care to the traditional provider-administered Pap testing. These findings add important information to the literature related to promoting cancer screening among women who are under or never screened for cervical cancer.

  10. Human-parathormone assay for use in dogs: validation, sample handling studies, and parathyroid function testing.

    Science.gov (United States)

    Torrance, A G; Nachreiner, R

    1989-07-01

    Ten commercially available parathormone (PTH) assays were competitively validated, using dilutional parallelism, intra-assay and interassay coefficients of variation, and sensitivity and measured responses of 2 dogs to calcium and EDTA infusions. A 2-site immunoradiometric assay for intact human-PTH was superior to the others for estimating canine-PTH, met the criteria for validity, and was further investigated. A series of sample-handling studies was performed. Serum and plasma samples stored at 24 C lost 15% (n = 5; P less than 0.05) of PTH between 2 and 24 hours. This did not occur at 6 C. The mean PTH concentration of sera from blood samples clotted at 24 C was 6% (P less than 0.05) higher than equivalent EDTA samples. Serum samples stored at 6 and 37 C deteriorated 35% and 100% (n = 5; P less than 0.05), respectively, after 1 week, whereas samples stored at -20 and -70 C for 4 weeks did not deteriorate. There was no significant deterioration of PTH in samples frozen (-40 C) and thawed up to 7 times (n = 5). Parathyroid function testing was investigated by use of 2-hour infusions of disodium EDTA (25 mg/kg/h), 10-minute infusions of calcium gluconate (3 mg of elemental calcium/kg/10 min), and physiologic saline controls (n = 8). Renal function was monitored before and after EDTA infusion by exogenous creatinine clearance. Infusion of disodium EDTA increased mean PTH concentration from 67 (time 0) to 317 and 235 pg/ml at 90 and 180 minutes, respectively (P less than 0.001). Infusion of calcium gluconate decreased mean PTH concentration from 84 (time 0) to 14 and 12 pg/ml at 15 and 60 minutes, respectively (P less than 0.005). There were no observable side effects of the infusions in normal conscious dogs and no differences in exogenous creatinine clearance after EDTA infusion.

  11. Freezing adversely affects measurement of vascular endothelial growth factor levels in human aqueous samples

    Directory of Open Access Journals (Sweden)

    Sankarathi Balaiya

    2011-01-01

    Full Text Available Sankarathi Balaiya Sandeep Grover Ravi K Murthy Kakarla V ChalamDepartment of Ophthalmology, University of Florida College of Medicine, Jacksonville, FL, USAPurpose: Aqueous levels of vascular endothelial growth factor (VEGF can be a surrogate marker of intraocular VEGF activity and a measure of efficacy of anti-VEGF treatment in a variety of vasoproliferative retinal disorders, including diabetic retinopathy, age-related macular degeneration, and central retinal vein occlusion. Measurement of the VEGF level may be adversely affected by premeasurement variables, such as freezing and delay, in sample analysis. We aim to evaluate the effect of storage and delayed measurement of human aqueous VEGF levels in these conditions.Methods: Aqueous samples collected from patients receiving intravitreal injection of bevacizumab for various retinal diseases were divided into two groups. In Group 1, the VEGF levels were analyzed on the same day; in Group 2, the VEGF levels were analyzed after 21 days of freezer storage (-80°C using immunobead assay. Statistical comparison using a paired t-test was performed between the two groups.Results: Thirty-one aqueous humor samples were collected, and the VEGF concentration for fresh samples was 7.8 ± 5.9 pg/mL (mean ± SD compared to 6.5 ± 6.0 pg/mL in frozen samples, resulting in a statistically significant difference (P = 0.03.Conclusions: Accurate measurement of the VEGF level is a vital component of clinical decision-making. Delayed analysis of VEGF levels in aqueous samples may result in significant sample degradation and lower levels of measured VEGF.Keywords: VEGF level, aqueous humor, immunobead assay, VEGF storage

  12. Study of microtip-based extraction and purification of DNA from human samples for portable devices

    Science.gov (United States)

    Fotouhi, Gareth

    DNA sample preparation is essential for genetic analysis. However, rapid and easy-to-use methods are a major challenge to obtaining genetic information. Furthermore, DNA sample preparation technology must follow the growing need for point-of-care (POC) diagnostics. The current use of centrifuges, large robots, and laboratory-intensive protocols has to be minimized to meet the global challenge of limited access healthcare by bringing the lab to patients through POC devices. To address these challenges, a novel extraction method of genomic DNA from human samples is presented by using heat-cured polyethyleneimine-coated microtips generating a high electric field. The microtip extraction method is based on recent work using an electric field and capillary action integrated into an automated device. The main challenges to the method are: (1) to obtain a stable microtip surface for the controlled capture and release of DNA and (2) to improve the recovery of DNA from samples with a high concentration of inhibitors, such as human samples. The present study addresses these challenges by investigating the heat curing of polyethyleneimine (PEI) coated on the surface of the microtip. Heat-cured PEI-coated microtips are shown to control the capture and release of DNA. Protocols are developed for the extraction and purification of DNA from human samples. Heat-cured PEI-coated microtip methods of DNA sample preparation are used to extract genomic DNA from human samples. It is discovered through experiment that heat curing of a PEI layer on a gold-coated surface below 150°C could inhibit the signal of polymerase chain reaction (PCR). Below 150°C, the PEI layer is not completely cured and dissolved off the gold-coated surface. Dissolved PEI binds with DNA to inhibit PCR. Heat curing of a PEI layer above 150°C on a gold-coated surface prevents inhibition to PCR and gel electrophoresis. In comparison to gold-coated microtips, the 225°C-cured PEI-coated microtips improve the

  13. Paenibacillus spp. isolated from human and environmental samples in Spain: detection of 11 new species

    Directory of Open Access Journals (Sweden)

    J.A. Sáez-Nieto

    2017-09-01

    Full Text Available One hundred thirty-six isolates, 88 human and 48 environmental, that met the requirements to belong to the genus Paenibacillus were identified using a polyphasic taxonomic approach known as 16S rRNA plus phenotypic traits. Thirty-seven Paenibacillus species were identified; some had not been previously reported from clinical samples. The main species were P. pabuli (13 isolates, P. provencensis (11, P. phoenicis (9 and P. lautus (8. P. pabuli (11/13 and P. provencensis (8/11 were mainly environmental isolates, while P. phoenicis (9/9 and P. lautus (6/8 were mainly human isolates. Despite the difficulties in assigning to human Paenibacillus isolates a role as a pathogen or contaminant, here 25% of the isolates were involved in true infections, especially in those cases that affected abscesses, wound exudates, ocular infections and diverse fluids. In addition, 15 isolates were identified as 11 ‘Candidatus’ to a new species, all of them from human specimens except one that was obtained from laboratory air. The antimicrobial susceptibility testing showed 95.6% of isolates were resistant to ampicillin, 44% were resistant to cotrimoxazole, 20 to 30% were resistant to cefotaxime and vancomycin and 13% were resistant to rifampicin and erythromycin.

  14. Applicability of the CALUX bioassay for screening of dioxin levels in human milk samples

    DEFF Research Database (Denmark)

    Laier, P.; Cederberg, Tommy Licht; Larsen, John Christian;

    2003-01-01

    . The results obtained with the bioassay when testing milk extracts fractionated into dioxins/furans, non-ortho PCB and mono/di-ortho PCB fractions indicated that the correlation between the bioassay and the chemical analyses depends primarily on the A receptor activity observed in the mono/di-ortho PCB......The CALUX (chemically activated luciferase expression) bioassay based on rat hepatoma (H4IIE) cells is a sensitive assay for the detection of Ah receptor agonists like 2,3,7,8-substituted chlorinated dibenzo-p-dioxins and dibenzofurans and related PCBs. In this paper, the assay was optimized...... and applied for monitoring levels of dioxins in human milk samples. Combination effects of dioxin-like compounds were evaluated by testing potential mechanisms of interaction between seven of the major dioxin-like compounds in human milk using the isobole method. Results showed that the compounds acted...

  15. Dioxin-like activity in environmental and human samples from Greenland and Denmark

    DEFF Research Database (Denmark)

    Long, Manhai; Bonefeld-Jørgensen, Eva Cecilie

    2012-01-01

    of surface waters with dioxins. Our results from human studies suggest that the serum DL-activity reflect the complex mixture of persistent organic pollutants (POPs). Greenlandic Inuit had lower serum DL-activity level compared to Europeans, probably due to long distance from the dioxin sources and UV...... degradation of the high potent dioxin and/or the inhibitory effect of the high level of non-DL POPs. Selective bioaccumulation of PCBs in the food chain may contribute to the negative correlation between serum POPs and DL-activity observed in Greenlandic Inuit. Hence the AhR transactivation bioassay provides...... a cost-effective and integrated screening tool for measurement of the DL-activity in human, environmental and commercial samples....

  16. Nondestructive sampling of human skeletal remains yields ancient nuclear and mitochondrial DNA.

    Science.gov (United States)

    Bolnick, Deborah A; Bonine, Holly M; Mata-Míguez, Jaime; Kemp, Brian M; Snow, Meradeth H; LeBlanc, Steven A

    2012-02-01

    Museum curators and living communities are sometimes reluctant to permit ancient DNA (aDNA) studies of human skeletal remains because the extraction of aDNA usually requires the destruction of at least some skeletal material. Whether these views stem from a desire to conserve precious materials or an objection to destroying ancestral remains, they limit the potential of aDNA research. To help address concerns about destructive analysis and to minimize damage to valuable specimens, we describe a nondestructive method for extracting DNA from ancient human remains. This method can be used with both teeth and bone, but it preserves the structural integrity of teeth much more effectively than that of bone. Using this method, we demonstrate that it is possible to extract both mitochondrial and nuclear DNA from human remains dating between 300 BC and 1600 AD. Importantly, the method does not expose the remains to hazardous chemicals, allowing them to be safely returned to curators, custodians, and/or owners of the samples. We successfully amplified mitochondrial DNA from 90% of the individuals tested, and we were able to analyze 1-9 nuclear loci in 70% of individuals. We also show that repeated nondestructive extractions from the same tooth can yield amplifiable mitochondrial and nuclear DNA. The high success rate of this method and its ability to yield DNA from samples spanning a wide geographic and temporal range without destroying the structural integrity of the sampled material may make possible the genetic study of skeletal collections that are not available for destructive analysis. Copyright © 2011 Wiley Periodicals, Inc.

  17. An Improved Method for High Quality Metagenomics DNA Extraction from Human and Environmental Samples

    DEFF Research Database (Denmark)

    Bag, Satyabrata; Saha, Bipasa; Mehta, Ojasvi

    2016-01-01

    and human origin samples. We introduced a combination of physical, chemical and mechanical lysis methods for proper lysis of microbial inhabitants. The community microbial DNA was precipitated by using salt and organic solvent. Both the quality and quantity of isolated DNA was compared with the existing...... methodologies and the supremacy of our method was confirmed. Maximum recovery of genomic DNA in the absence of substantial amount of impurities made the method convenient for nucleic acid extraction. The nucleic acids obtained using this method are suitable for different downstream applications. This improved...

  18. Modular Sampling and Analysis Techniques for the Real-Time Analysis of Human Breath

    Energy Technology Data Exchange (ETDEWEB)

    Frank, M; Farquar, G; Adams, K; Bogan, M; Martin, A; Benner, H; Spadaccini, C; Steele, P; Davis, C; Loyola, B; Morgan, J; Sankaran, S

    2007-07-09

    At LLNL and UC Davis, we are developing several techniques for the real-time sampling and analysis of trace gases, aerosols and exhaled breath that could be useful for a modular, integrated system for breath analysis. Those techniques include single-particle bioaerosol mass spectrometry (BAMS) for the analysis of exhaled aerosol particles or droplets as well as breath samplers integrated with gas chromatography mass spectrometry (GC-MS) or MEMS-based differential mobility spectrometry (DMS). We describe these techniques and present recent data obtained from human breath or breath condensate, in particular, addressing the question of how environmental exposure influences the composition of breath.

  19. Genetic Characterization of Atypical Mansonella (Mansonella) ozzardi Microfilariae in Human Blood Samples from Northeastern Peru

    Science.gov (United States)

    Marcos, Luis A.; Arrospide, Nancy; Recuenco, Sergio; Cabezas, Cesar; Weil, Gary J.; Fischer, Peter U.

    2012-01-01

    DNA sequence comparisons are useful for characterizing proposed new parasite species or strains. Microfilariae with an atypical arrangement of nuclei behind the cephalic space have been recently described in human blood samples from the Amazon region of Peru. Three blood specimens containing atypical microfilariae were genetically characterized using three DNA markers (5S ribosomal DNA, 12S ribosomal DNA, and cytochrome oxidase I). All atypical microfilariae were clustered into the Mansonella group and indistinguishable from M. ozzardi based on these DNA markers. PMID:22826497

  20. [The quality control of preanalytical variations for the determination of lead in samples of human origin].

    Science.gov (United States)

    Zhong, Kun; Wang, Wei; He, Falin; Wang, Zhiguo

    2015-02-01

    The aims of this article was to provide the quality control requirements of preanalytical variation for the determination of lead in samples of human origin, reduce the influence of preanalytical variation on the test results. According to the Clinical and Laboratory Standards Institute documents, control of preanalytical variation in trace element determinations, analytical procedures for the determination of lead in blood and urine and other references and guidelines, the methods of quality control of lead determination had been made, including: the factors needed to be considered before collection, preservation, transportation and other preanalytical factors, the abilities and considerations of laboratory staff, etc.

  1. Calcium isolation from large-volume human urine samples for 41Ca analysis by accelerator mass spectrometry.

    Science.gov (United States)

    Miller, James J; Hui, Susanta K; Jackson, George S; Clark, Sara P; Einstein, Jane; Weaver, Connie M; Bhattacharyya, Maryka H

    2013-08-01

    Calcium oxalate precipitation is the first step in preparation of biological samples for (41)Ca analysis by accelerator mass spectrometry. A simplified protocol for large-volume human urine samples was characterized, with statistically significant increases in ion current and decreases in interference. This large-volume assay minimizes cost and effort and maximizes time after (41)Ca administration during which human samples, collected over a lifetime, provide (41)Ca:Ca ratios that are significantly above background.

  2. Calcium Isolation from Large-Volume Human Urine Samples for 41Ca Analysis by Accelerator Mass Spectrometry

    Science.gov (United States)

    Miller, James J; Hui, Susanta K; Jackson, George S; Clark, Sara P; Einstein, Jane; Weaver, Connie M; Bhattacharyya, Maryka H

    2013-01-01

    Calcium oxalate precipitation is the first step in preparation of biological samples for 41Ca analysis by accelerator mass spectrometry. A simplified protocol for large-volume human urine samples was characterized, with statistically significant increases in ion current and decreases in interference. This large-volume assay minimizes cost and effort and maximizes time after 41Ca administration during which human samples, collected over a lifetime, provide 41Ca:Ca ratios that are significantly above background. PMID:23672965

  3. Human Cytomegalovirus and Human Umbilical Vein Endothelial Cells: Restriction of Primary Isolation to Blood Samples and Susceptibilities of Clinical Isolates from Other Sources to Adaptation

    OpenAIRE

    2002-01-01

    In immunocompromised patients with disseminated infection, human cytomegalovirus (HCMV) is widespread in the microvascular endothelium of multiple organs. Human umbilical vein endothelial cells (HUVEC) were used in parallel to human embryonic lung fibroblasts (HELF) to recover HCMV from blood samples of immunocompromised patients. Using the shell vial technique, comparable median numbers of p72-positive HUVEC and HELF cells were found with the 26 HCMV-positive buffy coat samples out of 150 ex...

  4. Optimization of a Pre-MEKC Separation SPE Procedure for Steroid Molecules in Human Urine Samples

    Directory of Open Access Journals (Sweden)

    Ilona Olędzka

    2013-11-01

    Full Text Available Many steroid hormones can be considered as potential biomarkers and their determination in body fluids can create opportunities for the rapid diagnosis of many diseases and disorders of the human body. Most existing methods for the determination of steroids are usually time- and labor-consuming and quite costly. Therefore, the aim of analytical laboratories is to develop a new, relatively low-cost and rapid implementation methodology for their determination in biological samples. Due to the fact that there is little literature data on concentrations of steroid hormones in urine samples, we have made attempts at the electrophoretic determination of these compounds. For this purpose, an extraction procedure for the optimized separation and simultaneous determination of seven steroid hormones in urine samples has been investigated. The isolation of analytes from biological samples was performed by liquid-liquid extraction (LLE with dichloromethane and compared to solid phase extraction (SPE with C18 and hydrophilic-lipophilic balance (HLB columns. To separate all the analytes a micellar electrokinetic capillary chromatography (MECK technique was employed. For full separation of all the analytes a running buffer (pH 9.2, composed of 10 mM sodium tetraborate decahydrate (borax, 50 mM sodium dodecyl sulfate (SDS, and 10% methanol was selected. The methodology developed in this work for the determination of steroid hormones meets all the requirements of analytical methods. The applicability of the method has been confirmed for the analysis of urine samples collected from volunteers—both men and women (students, amateur bodybuilders, using and not applying steroid doping. The data obtained during this work can be successfully used for further research on the determination of steroid hormones in urine samples.

  5. Three dimensional imaging of paraffin embedded human lung tissue samples by micro-computed tomography.

    Directory of Open Access Journals (Sweden)

    Anna E Scott

    Full Text Available Understanding the three-dimensional (3-D micro-architecture of lung tissue can provide insights into the pathology of lung disease. Micro computed tomography (µCT has previously been used to elucidate lung 3D histology and morphometry in fixed samples that have been stained with contrast agents or air inflated and dried. However, non-destructive microstructural 3D imaging of formalin-fixed paraffin embedded (FFPE tissues would facilitate retrospective analysis of extensive tissue archives of lung FFPE lung samples with linked clinical data.FFPE human lung tissue samples (n = 4 were scanned using a Nikon metrology µCT scanner. Semi-automatic techniques were used to segment the 3D structure of airways and blood vessels. Airspace size (mean linear intercept, Lm was measured on µCT images and on matched histological sections from the same FFPE samples imaged by light microscopy to validate µCT imaging.The µCT imaging protocol provided contrast between tissue and paraffin in FFPE samples (15 mm x 7 mm. Resolution (voxel size 6.7 µm in the reconstructed images was sufficient for semi-automatic image segmentation of airways and blood vessels as well as quantitative airspace analysis. The scans were also used to scout for regions of interest, enabling time-efficient preparation of conventional histological sections. The Lm measurements from µCT images were not significantly different to those from matched histological sections.We demonstrated how non-destructive imaging of routinely prepared FFPE samples by laboratory µCT can be used to visualize and assess the 3D morphology of the lung including by morphometric analysis.

  6. High-throughput miRNA profiling of human melanoma blood samples

    Directory of Open Access Journals (Sweden)

    Rass Knuth

    2010-06-01

    Full Text Available Abstract Background MicroRNA (miRNA signatures are not only found in cancer tissue but also in blood of cancer patients. Specifically, miRNA detection in blood offers the prospect of a non-invasive analysis tool. Methods Using a microarray based approach we screened almost 900 human miRNAs to detect miRNAs that are deregulated in their expression in blood cells of melanoma patients. We analyzed 55 blood samples, including 20 samples of healthy individuals, 24 samples of melanoma patients as test set, and 11 samples of melanoma patients as independent validation set. Results A hypothesis test based approch detected 51 differentially regulated miRNAs, including 21 miRNAs that were downregulated in blood cells of melanoma patients and 30 miRNAs that were upregulated in blood cells of melanoma patients as compared to blood cells of healthy controls. The tets set and the independent validation set of the melanoma samples showed a high correlation of fold changes (0.81. Applying hierarchical clustering and principal component analysis we found that blood samples of melanoma patients and healthy individuals can be well differentiated from each other based on miRNA expression analysis. Using a subset of 16 significant deregulated miRNAs, we were able to reach a classification accuracy of 97.4%, a specificity of 95% and a sensitivity of 98.9% by supervised analysis. MiRNA microarray data were validated by qRT-PCR. Conclusions Our study provides strong evidence for miRNA expression signatures of blood cells as useful biomarkers for melanoma.

  7. Detection of Human Papillomavirus 18 in Cervical Cancer Samples Using PCR-ELISA (DIAPOPS

    Directory of Open Access Journals (Sweden)

    KN Tafreshi

    2011-12-01

    Full Text Available Background and Objectives: Human Papillomavirus (HPV infection is a major risk factor for adenocarcinoma of the cervix. The high-risk types of the virus such as HPV16 and HPV18, which possess the E6 and E7 oncogenes, are responsible for approximately 50% of all cervical cancers. A rapid, sensitive and specific test has been proposed for detection of HPV to improve cervical cancer screening programs.Objectives: The aim of this study was to develop a fast PCR-ELISA assay designated as DIAPOPS (Detection of Immobilized Amplified Products in a One Phase Systemfor detection of HPV16 and HPV18 types in SCC samples and Pap smears. The type specific primers and probes were designed for PCR and PCR-ELISA. The amplified products were hybridized with a specific biotin-labeled probe for HPV18 inner amplicons. The hybrids were detected with peroxidase conjugated avidin. The test was performed on the paraffin block and Pap smear samples from the cervical cancer patients, and the results of DIAPOPS were compared with conventional PCR assay.Results: The 70 samples (SCC and Pap smear samples were collected from Imam Khomeini and Mirzakoochak Khan Hospitals in Tehran. The PCR-based method detected six HPV16 positive, three HPV18 positive and Two HPV33 positive samples. DIAPOPS results were compared with the conventional PCR results and they showed an increase in sensitivity of the DIAPOPS test. Not only all of them were confirmed by PCR-ELISA but also three samples that conventional PCR showed negative for HPV18, were demonstrated positive by the PCR-ELISA method.Conclusion: The results of the study show that modified PCR-ELISA assay is more sensitive to detect HPV types and can be used for diagnostic purposes.

  8. Quantitation of enniatins in biological samples of Wistar rats after oral administration by LC-MS/MS.

    Science.gov (United States)

    Escrivá, Laura; Font, Guillermina; Manyes, Lara

    2015-01-01

    The emerging Fusarium mycotoxins enniatins (ENNs) have diverse biological properties, mainly due to their ionophoric activity, and represent a potential risk to human and animal health since they are commonly found in food and feed. In vivo toxicity studies are scarce and limited to the major mycotoxins. Until now, any method for the simultaneous analysis of these compounds in plasma, serum and feces from rat has been reported. A method for the extraction and determination of ENNs A, A1, B and B1 from Wistar rat samples by liquid chromatography tandem mass spectrometry has been developed. The method was successfully validated with satisfactory recoveries (70-106%), good intraday (rat samples that were administered a mixture of ENNs containing 1.19, 2.16, 1.03 and 1.41 mg/kg body weight of ENN A, A1, B and B1, respectively. Blood, urine and feces samples collected every 2 h during the 8-h duration of the experiment were analyzed. The administered dose of the mixture of ENNs did not cause observable adverse effects on the animals. ENNs concentrations detected in serum and urine were below LOQs. The four ENNs were detected in feces reaching the maximum concentration at 6 h after administration.

  9. Multi-elemental imaging of paraffin-embedded human samples by laser-induced breakdown spectroscopy

    Science.gov (United States)

    Moncayo, S.; Trichard, F.; Busser, B.; Sabatier-Vincent, M.; Pelascini, F.; Pinel, N.; Templier, I.; Charles, J.; Sancey, L.; Motto-Ros, V.

    2017-07-01

    Chemical elements play central roles for physiological homeostasis in human cells, and their dysregulation might lead to a certain number of pathologies. Novel imaging techniques that improve the work of pathologists for tissue analysis and diagnostics are continuously sought. We report the use of Laser-Induced Breakdown Spectroscopy (LIBS) to perform multi-elemental images of human paraffin-embedded skin samples on the entire biopsy scale in a complementary and compatible way with microscope histopathological examination. A specific instrumental configuration is proposed in order to detect most of the elements of medical interest (i.e. P, Al, Mg, Na, Zn, Si, Fe, and Cu). As an example of medical application, we selected and analysed skin biopsies, including healthy skin tissue, cutaneous metastasis of melanoma, Merkel-cell carcinoma and squamous cell carcinoma. Clear distinctions in the distribution of chemical elements are observed from the different samples investigated. This study demonstrates the high complementarity of LIBS elemental imaging with conventional histopathology, opening new opportunities for any medical application involving metals.

  10. Quantitative second-harmonic generation imaging to detect osteogenesis imperfecta in human skin samples

    Science.gov (United States)

    Adur, J.; Ferreira, A. E.; D'Souza-Li, L.; Pelegati, V. B.; de Thomaz, A. A.; Almeida, D. B.; Baratti, M. O.; Carvalho, H. F.; Cesar, C. L.

    2012-03-01

    Osteogenesis Imperfecta (OI) is a genetic disorder that leads to bone fractures due to mutations in the Col1A1 or Col1A2 genes that affect the primary structure of the collagen I chain with the ultimate outcome in collagen I fibrils that are either reduced in quantity or abnormally organized in the whole body. A quick test screening of the patients would largely reduce the sample number to be studied by the time consuming molecular genetics techniques. For this reason an assessment of the human skin collagen structure by Second Harmonic Generation (SHG) can be used as a screening technique to speed up the correlation of genetics/phenotype/OI types understanding. In the present work we have used quantitative second harmonic generation (SHG) imaging microscopy to investigate the collagen matrix organization of the OI human skin samples comparing with normal control patients. By comparing fibril collagen distribution and spatial organization, we calculated the anisotropy and texture patterns of this structural protein. The analysis of the anisotropy was performed by means of the two-dimensional Discrete Fourier Transform and image pattern analysis with Gray-Level Co-occurrence Matrix (GLCM). From these results, we show that statistically different results are obtained for the normal and disease states of OI.

  11. Goblet cells of the normal human bulbar conjunctiva and their assessment by impression cytology sampling.

    Science.gov (United States)

    Doughty, Michael J

    2012-07-01

    Goblet cells of the conjunctiva are the main source of mucus for the ocular surface. The objectives of this review are to consider the goblet cells as assessed by various histological, cytological and electron microscopy methods, and to assess the consistency of published reports (over more than 25 years) of goblet cell density (GCD) from impression cytology specimens from nominally healthy human subjects. Reported GCD values have been notably variable, with a range from 24 to 2226 cells/mm² for average values. Data analysis suggests that a high density of goblet cells should be expected for the healthy human conjunctiva, with a tendency toward higher values in samples taken from normally covered locations (inferior and superior bulbar conjunctiva) of the open eye (at 973 +/- 789 cells/ mm²) than in samples taken from exposed (interpalpebral) locations (at 427 +/- 376 cells/mm²). No obvious change in GCD was found with respect to age, perhaps because the variability of the data did not allow detection of any age-related decline in GCD. Analyses of published data from 33 other sources indicated a trend for GCD to be lower than normal across a spectrum of ocular surface diseases.

  12. Metagenomic Survey of Viral Diversity Obtained from Feces of Subantarctic and South American Fur Seals

    Science.gov (United States)

    Kluge, Mariana; Campos, Fabrício Souza; Tavares, Maurício; de Amorim, Derek Blaese; Valdez, Fernanda Pedone; Giongo, Adriana; Roehe, Paulo Michel; Franco, Ana Claudia

    2016-01-01

    The Brazilian South coast seasonally hosts numerous marine species, observed particularly during winter months. Some animals, including fur seals, are found dead or debilitated along the shore and may harbor potential pathogens within their microbiota. In the present study, a metagenomic approach was performed to evaluate the viral diversity in feces of fur seals found deceased along the coast of the state of Rio Grande do Sul. The fecal virome of two fur seal species was characterized: the South American fur seal (Arctocephalus australis) and the Subantarctic fur seal (Arctocephalus tropicalis). Fecal samples from 10 specimens (A. australis, n = 5; A. tropicalis, n = 5) were collected and viral particles were purified, extracted and amplified with a random PCR. The products were sequenced through Ion Torrent and Illumina platforms and assembled reads were submitted to BLASTx searches. Both viromes were dominated by bacteriophages and included a number of potentially novel virus genomes. Sequences of picobirnaviruses, picornaviruses and a hepevirus-like were identified in A. australis. A rotavirus related to group C, a novel member of the Sakobuvirus and a sapovirus very similar to California sea lion sapovirus 1 were found in A. tropicalis. Additionally, sequences of members of the Anelloviridae and Parvoviridae families were detected in both fur seal species. This is the first metagenomic study to screen the fecal virome of fur seals, contributing to a better understanding of the complexity of the viral community present in the intestinal microbiota of these animals. PMID:26986573

  13. Metagenomic Survey of Viral Diversity Obtained from Feces of Subantarctic and South American Fur Seals.

    Directory of Open Access Journals (Sweden)

    Mariana Kluge

    Full Text Available The Brazilian South coast seasonally hosts numerous marine species, observed particularly during winter months. Some animals, including fur seals, are found dead or debilitated along the shore and may harbor potential pathogens within their microbiota. In the present study, a metagenomic approach was performed to evaluate the viral diversity in feces of fur seals found deceased along the coast of the state of Rio Grande do Sul. The fecal virome of two fur seal species was characterized: the South American fur seal (Arctocephalus australis and the Subantarctic fur seal (Arctocephalus tropicalis. Fecal samples from 10 specimens (A. australis, n = 5; A. tropicalis, n = 5 were collected and viral particles were purified, extracted and amplified with a random PCR. The products were sequenced through Ion Torrent and Illumina platforms and assembled reads were submitted to BLASTx searches. Both viromes were dominated by bacteriophages and included a number of potentially novel virus genomes. Sequences of picobirnaviruses, picornaviruses and a hepevirus-like were identified in A. australis. A rotavirus related to group C, a novel member of the Sakobuvirus and a sapovirus very similar to California sea lion sapovirus 1 were found in A. tropicalis. Additionally, sequences of members of the Anelloviridae and Parvoviridae families were detected in both fur seal species. This is the first metagenomic study to screen the fecal virome of fur seals, contributing to a better understanding of the complexity of the viral community present in the intestinal microbiota of these animals.

  14. Diarrhea-associated pathogens, lactobacilli and cellulolytic bacteria in equine feces: responses to antibiotic challenge.

    Science.gov (United States)

    Harlow, Brittany E; Lawrence, Laurie M; Flythe, Michael D

    2013-09-27

    Antibiotics are important to equine medicine, but antibiotic-associated diarrhea (AAD) can lead to poor performance and even mortality. AAD is attributed to disruption of the hindgut microbiota, which permits proliferation of pathogenic microbes. The goal of this study was to evaluate the effects of common antibiotics on cellulolytic bacteria, lactobacilli, and AAD-associated pathogens in the feces of healthy horses. Fifteen horses were assigned to three treatment groups (blocked by age and sex): control (no antibiotics), trimethoprim-sulfadiazine (PO), or ceftiofur (IM). Fecal samples (n=8 per horse) were taken during dietary adaptation (3 weeks), antibiotic challenge (1 week), and withdrawal (1 week). Bacteria were enumerated by serial dilution and viable count. Cellulolytic bacteria decreased by >99% during administration of either antibiotic (Pantibiotic challenge period (PAntibiotic challenged horses also shed more salmonella than control horses (PAntibiotics had no effect on the number of Clostridium perfringens isolates. There was no detectable Clostridium difficile during adaptation or in any control horse. C. difficile increased (Pantibiotics, and were still detectable 1 week after withdrawal. These results indicate that antibiotics can disrupt the normal gastrointestinal microbiota and allow proliferation of Salmonella spp. and C. difficile.

  15. Deep RNA sequencing analysis of readthrough gene fusions in human prostate adenocarcinoma and reference samples

    Directory of Open Access Journals (Sweden)

    Nacu Serban

    2011-01-01

    Full Text Available Abstract Background Readthrough fusions across adjacent genes in the genome, or transcription-induced chimeras (TICs, have been estimated using expressed sequence tag (EST libraries to involve 4-6% of all genes. Deep transcriptional sequencing (RNA-Seq now makes it possible to study the occurrence and expression levels of TICs in individual samples across the genome. Methods We performed single-end RNA-Seq on three human prostate adenocarcinoma samples and their corresponding normal tissues, as well as brain and universal reference samples. We developed two bioinformatics methods to specifically identify TIC events: a targeted alignment method using artificial exon-exon junctions within 200,000 bp from adjacent genes, and genomic alignment allowing splicing within individual reads. We performed further experimental verification and characterization of selected TIC and fusion events using quantitative RT-PCR and comparative genomic hybridization microarrays. Results Targeted alignment against artificial exon-exon junctions yielded 339 distinct TIC events, including 32 gene pairs with multiple isoforms. The false discovery rate was estimated to be 1.5%. Spliced alignment to the genome was less sensitive, finding only 18% of those found by targeted alignment in 33-nt reads and 59% of those in 50-nt reads. However, spliced alignment revealed 30 cases of TICs with intervening exons, in addition to distant inversions, scrambled genes, and translocations. Our findings increase the catalog of observed TIC gene pairs by 66%. We verified 6 of 6 predicted TICs in all prostate samples, and 2 of 5 predicted novel distant gene fusions, both private events among 54 prostate tumor samples tested. Expression of TICs correlates with that of the upstream gene, which can explain the prostate-specific pattern of some TIC events and the restriction of the SLC45A3-ELK4 e4-e2 TIC to ERG-negative prostate samples, as confirmed in 20 matched prostate tumor and normal

  16. Exploring the acceptability of human papillomavirus self-sampling among Muslim immigrant women

    Directory of Open Access Journals (Sweden)

    Lofters AK

    2017-07-01

    Full Text Available Aisha K Lofters,1–4 Mandana Vahabi,5,6 Mitra Fardad,7 Afrah Raza8 1Centre for Urban Health Solutions, Li Ka Shing Knowledge Institute, St. Michael’s Hospital, 2Department of Family and Community Medicine, University of Toronto, 3Department of Family and Community Medicine, St. Michael’s Hospital, 4Institute for Clinical Evaluative Sciences, 5Faculty of Community Services, Daphne Cockwell School of Nursing, 6Graduate Program in Immigration and Settlement Studies, Ryerson University, 7Faculty of Community Service, Daphne Cockwell School of Nursing, Ryerson University, Toronto, ON, Canada; 8University of Michigan Medical School, Ann Arbor, MI, USA Background: With appropriate screening (ie, the Papanicolaou [Pap] test, cervical cancer is highly preventable, and high-income countries, including Canada, have observed significant decreases in cervical cancer mortality. However, certain subgroups, including immigrants from countries with large Muslim populations, experience disparities in cervical cancer screening. Little is known about the acceptability of human papillomavirus (HPV self-sampling as a screening strategy among Muslim immigrant women in Canada. This study assessed cervical cancer screening practices, knowledge and attitudes, and acceptability of HPV self-sampling among Muslim immigrant women. Methods: A convenience sample of 30 women was recruited over a 3-month period (June–August 2015 in the Greater Toronto Area. All women were between 21 and 69 years old, foreign-born, and self-identified as Muslim, and had good knowledge of English. Data were collected through a self-completed questionnaire. Results: More than half of the participants falsely indicated that Pap tests may cause cervical infection, and 46.7% indicated that the test is an intrusion on privacy. The majority of women reported that they would be willing to try HPV self-sampling, and more than half would prefer this method to provider-administered sampling methods

  17. Variation in the Extraction Efficiency of Estradiol and Progesterone in Moist and Lyophilized Feces of the Black Howler Monkey (Alouatta pigra): Alternative Methods

    Science.gov (United States)

    Torres-Pelayo, Vianey del R.; Rovirosa-Hernández, M. J.; García-Orduña, F.; Chavira-Ramírez, R. D.; Boeck, L.; Canales-Espinosa, D.; Rodríguez-Landa, J. F.

    2011-01-01

    Several fecal steroid extraction techniques have been developed to measure the ovary function in different species of mammals. However, regardless of the method of extraction and the sample type chosen, it has been observed that they can yield results with different percentages of recuperation. The objective of this study was to determine whether the type of substratum, solvent and extraction method used have any influence on the extraction efficiency in the feces of Alouatta pigra (black howler monkey). For this purpose we used two methods: agitation and ebullition. With each method, we utilized moist and lyophilized feces. The validation of radioimmunoassay method was accurate and precise for quantify estradiol and progesterone in lyophilized feces of A. pigra. To both of which ethanol and methanol, absolute and at 80%, were added, besides the hormones 125I-Estradiol and 125I-Progesterone. The extraction efficiency for 125I-Estradiol was from 87.72 ± 3.97 to 41.24 ± 2.67%, and for 125I-Progesterone from 71.15 ± 4.24 to 42.30 ± 1.19% when we used the agitation method. Whereas with the ebullition method, the extraction efficiency for 125I-Estradiol ranged from 86.89 ± 2.66 to 71.68 ± 3.02% and for 125I-Progesterone from 98.31 ± 1.26 to 85.40 ± 1.98%. Due to the differences found in these assays, which depend on the method used, the type of feces employed and the type of solvent added to them, we recommend the ebullition method and the lyophilized feces of A. pigra for extracting the hormones, since in moist feces there may exist variables which might interfere in the quantification of 125I-Estradiol and 125I-Progesterone. PMID:22194723

  18. Prevalence of human papillomavirus infection in a clinic sample of transsexuals in Italy.

    Science.gov (United States)

    Loverro, Giuseppe; Di Naro, Edoardo; Caringella, Anna Maria; De Robertis, Anna Lisa; Loconsole, Daniela; Chironna, Maria

    2016-02-01

    Detectable human papillomavirus (HPV) DNA is the most common sexually transmitted infection. Reports on the prevalence of detectable HPV DNA among transsexuals (not sex workers) are scarce. The objective of the study was to determine the prevalence of detectable HPV DNA in a clinic sample of transsexuals and to assess the relationship between detectable HPV DNA and cytological outcomes. Clinical samples (oral, anal, vaginal, cervicovaginal and penile scraped cells) from 35 transsexuals (surgically treated and surgically untreated) who attended the outpatient Clinic of Gender Identity Dysphoria of the Department of Obstetrics and Gynecology of Policlinico Hospital (Bari, Italy) were collected for cytological analysis and HPV DNA detection and typing. All enrolled subjects answered an anonymous structured questionnaire about their sexual habits. Serological status for other sexually transmitted diseases (hepatitis B virus (HBV), hepatitis C virus (HCV), HIV and syphilis) was also evaluated. HPV DNA was detected in 14 of 35 patients (40.0%). The prevalence of detectable HPV DNA was 38.2% (13/34) in tested anal samples, 9.1% (2/22) in vaginal samples and 8.3% (1/12) in penile samples. Oncogenic HPV genotypes have been detected in 93% of HPV-positive transsexuals. More than one-third (35.7%) of HPV-positive transsexuals were infected with at least one of the four vaccine-preventable genotypes, 6, 11, 16 and 18. The high rate of detectable HPV DNA by oncogenic types suggests that periodic cytological screening and clinical evaluation may be necessary since transsexuals are at high risk of anogenital cancer. Also promoting HPV vaccination in younger subjects may be advisable. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

  19. Norovirus in feces and nasopharyngeal swab of children with and without acute gastroenteritis symptoms: First report of GI.5 in Brazil and GI.3 in nasopharyngeal swab.

    Science.gov (United States)

    Dábilla, Nathânia; Nunes Vieira Almeida, Tâmera; Carvalho Rebouças Oliveira, Anniely; Kipnis, André; Neres Silva, Thairiny; Souza Fiaccadori, Fabíola; Teixeira de Sousa, Teresinha; de Paula Cardoso, Divina das Dôres; Souza, Menira

    2017-02-01

    Noroviruses (NoVs) are an important cause of acute gastroenteritis (AGE), worldwide. To evaluate the frequency, viral load and molecular profile of NoV in fecal and nasopharyngeal swab samples from hospitalized children, and to determine children's secretor status. From May 2014 to May 2015, 219 children were included in the study, 96 with gastroenteric symptoms and 123 without gastroenteric symptoms. All fecal and nasopharyngeal swab samples were screened by TaqMan RT-qPCR duplex (GI/GII NoV) and quality samples were characterized by genomic sequencing. Norovirus positivity rate in feces was 15.4% in asymptomatic and 18.8% in the symptomatic group. The median viral loads in feces were 2.69×10(8)GC/g and 4.32×10(7)GC/g from children with or without AGE symptoms, respectively. In nasopharyngeal swab samples, the NoV positivity was 11.4% in symptomatic children, with a median viral load of 2.20×10(7)GC/mL and 6.5% in asymptomatic children, with an average viral load of 1.73×10(6)GC/mL. In only two cases NoV was detected in both samples. A considerable genomic variability was observed in feces, with six genotypes being detected, as follows: GII.4, GII.6, GI.3 and GII.3, GI.2 and GI.5. Two GI.3 was detected in nasopharyngeal swab. Our data reveal considerable NoV frequencies in both nasopharyngeal and fecal samples from symptomatic and asymptomatic children. Higher viral loads were detected in samples from AGE symptomatic children, when compared to asymptomatic children. High genomic variability was observed, with this being the first report of GI.5 NoV in Brazil and of GI.3 in nasopharyngeal swab samples. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Comparison of sequencing platforms for single nucleotide variant calls in a human sample.

    Science.gov (United States)

    Ratan, Aakrosh; Miller, Webb; Guillory, Joseph; Stinson, Jeremy; Seshagiri, Somasekar; Schuster, Stephan C

    2013-01-01

    Next-generation sequencings platforms coupled with advanced bioinformatic tools enable re-sequencing of the human genome at high-speed and large cost savings. We compare sequencing platforms from Roche/454(GS FLX), Illumina/HiSeq (HiSeq 2000), and Life Technologies/SOLiD (SOLiD 3 ECC) for their ability to identify single nucleotide substitutions in whole genome sequences from the same human sample. We report on significant GC-related bias observed in the data sequenced on Illumina and SOLiD platforms. The differences in the variant calls were investigated with regards to coverage, and sequencing error. Some of the variants called by only one or two of the platforms were experimentally tested using mass spectrometry; a method that is independent of DNA sequencing. We establish several causes why variants remained unreported, specific to each platform. We report the indel called using the three sequencing technologies and from the obtained results we conclude that sequencing human genomes with more than a single platform and multiple libraries is beneficial when high level of accuracy is required.

  1. Accumulation levels and characteristics of some pesticides in human adipose tissue samples from Southeast China.

    Science.gov (United States)

    Wang, Na; Shi, Lili; Kong, Deyang; Cai, Daoji; Cao, Yanzhong; Liu, Yongming; Pang, Guofang; Yu, Rongbin

    2011-08-01

    This paper presents a comprehensive study of pesticide levels and bio-accumulation characteristics in human adipose tissues among residents of Southeast China. A large number of adipose samples (n=633) were selected for 58 pesticides and were analyzed by high sensitive Gas Chromatography-Tandem Mass Spectrometry (GC-MS/MS). The results showed that POPs pesticides were frequently detected, including 2,4'-DDD, 2,4'-DDE, 2,4'-DDT, 4,4'-DDD, 4,4'-DDE, 4,4'-DDT, α-HCH, β-HCH, γ-HCH, δ-HCH, hexachlorobenzene (HCB), and mirex. Other detected pesticide species were dicofol, methamidophos and chlordimeform, which have rarely been reported. Comparing to different countries, the concentrations of total DDT and HCH in these three Chinese southeastern sites were in the middle range, whereas the HCB and mirex were in the lower end. A significant correlation was observed between region as well as age and POPs pesticide levels. Some pesticide residue levels were also found significantly correlated to occupation. However, there was no significant correlation between gender and pesticides. Meanwhile, it is interesting to find that mortality of malignant tumors tends to associate with the pesticides levels in human adipose tissue. More importantly, the measured data presented in this study provide realistic information which is useful for assessing human exposure to pesticides in the general population of Southeast China. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. Tracking human footprints in Antarctica through passive sampling of polycyclic aromatic hydrocarbons in inland lakes.

    Science.gov (United States)

    Yao, Yao; Meng, Xiang-Zhou; Wu, Chen-Chou; Bao, Lian-Jun; Wang, Feng; Wu, Feng-Chang; Zeng, Eddy Y

    2016-06-01

    Freely dissolved polycyclic aromatic hydrocarbons (PAHs) were monitored in seven inland lakes of Antarctica by a polyethylene (PE)-based passive sampling technique, with the objective of tracking human footprints. The measured concentrations of PAHs were in the range of 14-360 ng L(-1) with the highest values concentrated around the Russian Progress II Station, indicating the significance of human activities to the loading of PAHs in Antarctica. The concentrations of PAHs in the inland lakes were in the upper part of the PAHs levels in aquatic environments from remote and background regions across the globe. The composition profiles of PAHs indicated that PAHs in the inland lakes were derived mainly from local oil spills, which was corroborated by a large number of fuel spillage reports from ship and plane crash incidents in Antarctica during recent years. Clearly, local human activities, rather than long-range transport, are the dominant sources of PAH contamination to the inland lakes. Finally, the present study demonstrates the efficacy of PE-based passive samplers for investigating PAHs in the aquatic environment of Antarctica under complex field conditions.

  3. Detection of Mycobacterium tuberculosis and Mycobacterium bovis from human sputum samples through multiplex PCR.

    Science.gov (United States)

    Jabbar, Abdul; Khan, Jafar; Ullah, Aman; Rehman, Hazir; Ali, Ijaz

    2015-07-01

    Tuberculosis (TB) has a long history and being present even before the start of recording history. It has left detrimental effects on all aspect of the life and geared the developments in the science of health. TB is caused by Mycobacterium tuberculosis complex (MTBC) including five species M. tuberculosis, M. bovis, M. africanum, M. canetti, and M. microti. M. tuberculosis and M. bovis infect both animals and humans. Therefore, differentiation of these two closely related species is very important for epidemiological and management purpose. We undertook the present study to characterize mycobacteria isolated from sputum of known TB patients by conventional methods and further, by multiplex PCR (mPCR) to detect the prevalence of Zoonotic TB (TB caused by M. bovis). Sputum samples from TB patient were collected from two tertiary care hospitals in Peshawar i.e. Lady Reading Hospital and Hayatabad Medical Complex. All the samples were subjected to Ziehl Neelsen (ZN) stain, culture on Lowenstein Jensen (LJ) and Stone Brink medium, Nitrate reduction test and multiplex PCR. A total of hundred mycobacterial strains were isolated from these samples on the basis of ZN staining, cultural and biochemical methods. Later on, these isolates were subjected to multiplex PCR by using pncATB-1.2 and pncAMT-2 primers specific to M. tuberculosis and JB21, JB22 primers specific to M. bovis. By means of conventional method, these hundred cultures isolates were differentiated into M. tuberculosis (ninety six) and M. bovis (four). Furthermore, by mPCR, it was determined that out of hundred isolates, ninety-eight were identified as M. tuberculosis and two isolates as M. bovis. This molecular method enables to differentiate M. bovis from M. tuberculosis in human sputum.

  4. A universal genome sequencing method for rotavirus A from human fecal samples which identifies segment reassortment and multi-genotype mixed infection.

    Science.gov (United States)

    Dung, Tran Thi Ngoc; Duy, Pham Thanh; Sessions, October M; Sangumathi, Uma K; Phat, Voong Vinh; Tam, Pham Thi Thanh; To, Nguyen Thi Nguyen; Phuc, Tran My; Hong Chau, Tran Thi; Chau, Nguyen Ngoc Minh; Minh, Ngoc Nguyen; Thwaites, Guy E; Rabaa, Maia A; Baker, Stephen

    2017-04-24

    Genomic characterization of rotavirus (RoV) has not been adopted at large-scale due to the complexity of obtaining sequences for all 11 segments, particularly when feces are used as starting material. To overcome these limitations, we developed a novel RoV capture and genome sequencing method combining commercial enzyme immunoassay plates and a set of routinely used reagents. Our approach had a 100% success rate, producing >90% genome coverage for diverse RoV present in fecal samples (Ct RoV characterization and could be scaled-up for use in global RoV surveillance systems. Current Controlled Trials ISRCTN88101063 . Date of registration: 14/06/2012.

  5. DNA isolation protocols affect the detection limit of PCR approaches of bacteria in samples from the human gastrointestinal tract

    NARCIS (Netherlands)

    Zoetendal, E.G.; Ben-Amor, K.; Akkermans, A.D.L.; Abee, T.; Vos, de W.M.

    2001-01-01

    A major concern in molecular ecological studies is the lysis efficiency of different bacteria in a complex ecosystem. We used a PCR-based 16S rDNA approach to determine the effect of two DNA isolation protocols (i.e. the bead beating and Triton-X100 method) on the detection limit of seven feces-asso

  6. Human genomic DNA analysis using a semi-automated sample preparation, amplification, and electrophoresis separation platform.

    Science.gov (United States)

    Raisi, Fariba; Blizard, Benjamin A; Raissi Shabari, Akbar; Ching, Jesus; Kintz, Gregory J; Mitchell, Jim; Lemoff, Asuncion; Taylor, Mike T; Weir, Fred; Western, Linda; Wong, Wendy; Joshi, Rekha; Howland, Pamela; Chauhan, Avinash; Nguyen, Peter; Petersen, Kurt E

    2004-03-01

    The growing importance of analyzing the human genome to detect hereditary and infectious diseases associated with specific DNA sequences has motivated us to develop automated devices to integrate sample preparation, real-time PCR, and microchannel electrophoresis (MCE). In this report, we present results from an optimized compact system capable of processing a raw sample of blood, extracting the DNA, and performing a multiplexed PCR reaction. Finally, an innovative electrophoretic separation was performed on the post-PCR products using a unique MCE system. The sample preparation system extracted and lysed white blood cells (WBC) from whole blood, producing DNA of sufficient quantity and quality for a polymerase chain reaction (PCR). Separation of multiple amplicons was achieved in a microfabricated channel 30 microm x 100 microm in cross section and 85 mm in length filled with a replaceable methyl cellulose matrix operated under denaturing conditions at 50 degrees C. By incorporating fluorescent-labeled primers in the PCR, the amplicons were identified by a two-color (multiplexed) fluorescence detection system. Two base-pair resolution of single-stranded DNA (PCR products) was achieved. We believe that this integrated system provides a unique solution for DNA analysis.

  7. [Phenotypic characterization and distribution of Yersinia in human and environmental samples].

    Science.gov (United States)

    Javier Castillo, F; Larraz, V; Asunción Lafarga, M; Navarro, M; Gómez-Lus, R

    1994-01-01

    The distribution of species and phenotypes of Yersinia isolated from environmental samples over an eight year period are compared to that of stool cultures obtained from patients of the same geographical location (Zaragoza, Spain). The number of samples and the percentage contamination were as follows: wastewater 362, 67.4%, freshwater 523, 13.4%, raw food 607, 24.5% and cooked food 1134, 7.9%. Yersinia enterocolitica was isolated significantly more frequently than other species in wastewater, while Yersinia intermedia was the most significant species found in freshwater. Significant differences between the percentage isolates of identified species in raw and cooked foods were not found. Fifteen different serogroups were identified from faeces, thirteen of which were also isolated from environmental samples. Three serogroups of Y. enterocolitica associated with human disease were isolated from the patients faeces as follows: O:3, 145 cases; O:8, 3 cases and O:5,27, 1 case. A low proportion were isolated from food: O:3, 3 strains; O:8, 2 strains and O:5,27, 5 strains. Only one isolate from serogroup O:3 was obtained from freshwater.

  8. [Detection and typing by molecular biology of human papillomavirus in genital samples].

    Science.gov (United States)

    Suárez Moya, A; Esquivias Gómez, J I; Vidart Aragón, J A; Picazo de la Garza, J J

    2006-06-01

    Recently, there has been a marked increase in human papillomavirus (HPV) infection, and the etiological relationship between some HPV genotypes and genital cancer has been confirmed. Therefore, we used current molecular biology techniques to evaluate the prevalence of these viruses and their genotype in genital samples. We processed 401 genital samples from 281 women and 120 men, all with a diagnosis compatible with HPV infection. Virus was detected using PCR, and positive samples were typed using an array technique which enabled us to detect the 35 most common types of mucous-associated HPV. Of the 401 patients studied, 185 (46.1%) were positive, and only one type of HPV was detected in 133 cases. We found that 41.6% of the women and 56.7% of the men were positive. A total of 260 HPVs were typed; 154 were high oncogenic risk. They infected 16 men (23.5%) and 88 women (75.2%). The difference was statistically significant (pHVP 16 in 52 cases. We found a 46% prevalence of HPV infection. More than half of these patients were infected by high-risk HPV. The presence of high-risk HPV was significantly higher in women.

  9. Polychlorinated biphenyls and organochlorine pesticides in human milk samples from two regions in Croatia.

    Science.gov (United States)

    Klinčić, D; Herceg Romanić, S; Matek Sarić, M; Grzunov, J; Dukić, B

    2014-03-01

    We analyzed 20 polychlorinated biphenyls (PCBs) and seven organochlorine pesticides (OCPs) in milk samples collected during 2009-2011 from primiparae living in two different regions in Croatia. p,p'-DDE is the dominant organochlorine pesticide. α-HCH/γ-HCH and p,p'-DDE/p,p'-DDT ratios indicate that there is fresh input of γ-HCH in investigated population on both locations, while this is not applicable to p,p'-DDT. The PCB profile was dominated by higher chlorinated congeners. Non-ortho PCB congeners which have the highest TEF values were not detected in any of individual samples. Toxic equivalents for mono-ortho substituted PCB congeners indicated higher exposure to toxic PCBs in Zadar, but estimated daily intakes for both locations indicate that infants consuming mother's milk are not at risk of adverse effects caused by PCBs and OCPs. Our study builds on the previous research of human milk samples collected in Zagreb and reveals that over 10-year period, levels of investigated organochlorine compounds decreased significantly. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Analysis of inflammatory response in human plasma samples by an automated multicapillary electrophoresis system.

    Science.gov (United States)

    Larsson, Anders; Hansson, Lars-Olof

    2004-01-01

    A new automated multicapillary zone electrophoresis instrument with a new high-resolution (HR) buffer (Capillarys with HR buffer) for analysis of human plasma proteins was evaluated. Albumin, alpha(1)-antitrypsin, alpha(1)-acid glycoprotein, haptoglobin, fibrinogen, immunoglobulin (Ig)A, IgG and IgM were determined nephelometrically in 200 patient plasma samples. The same samples were then analyzed on the Capillarys system (Sebia, Paris, France). The albumin concentration from the nephelometric determination was used for quantification of the individual peaks in the capillary electrophoresis (CE) electropherogram. There was strong linear correlation between the nephelometric and electrophoretic determination of alpha(1)-antitrypsin (R(2) = 0.906), alpha(1)-acid glycoprotein (R(2) =0.894) and haptoglobin (R(2) = 0.913). There was also good correlation between the two determinations of gamma-globulins (R(2) = 0.883), while the correlation was weaker for fibrinogen (R(2) = 0.377). The Capillarys instrument is a reliable system for plasma protein analysis, combining the advantages of full automation, good analytical performance and high throughput. The HR buffer in combination with albumin quantification allows the simultaneous quantification of inflammatory markers in plasma samples without the need for nephelometric determination of these proteins.

  11. PERT: a method for expression deconvolution of human blood samples from varied microenvironmental and developmental conditions.

    Directory of Open Access Journals (Sweden)

    Wenlian Qiao

    Full Text Available The cellular composition of heterogeneous samples can be predicted using an expression deconvolution algorithm to decompose their gene expression profiles based on pre-defined, reference gene expression profiles of the constituent populations in these samples. However, the expression profiles of the actual constituent populations are often perturbed from those of the reference profiles due to gene expression changes in cells associated with microenvironmental or developmental effects. Existing deconvolution algorithms do not account for these changes and give incorrect results when benchmarked against those measured by well-established flow cytometry, even after batch correction was applied. We introduce PERT, a new probabilistic expression deconvolution method that detects and accounts for a shared, multiplicative perturbation in the reference profiles when performing expression deconvolution. We applied PERT and three other state-of-the-art expression deconvolution methods to predict cell frequencies within heterogeneous human blood samples that were collected under several conditions (uncultured mono-nucleated and lineage-depleted cells, and culture-derived lineage-depleted cells. Only PERT's predicted proportions of the constituent populations matched those assigned by flow cytometry. Genes associated with cell cycle processes were highly enriched among those with the largest predicted expression changes between the cultured and uncultured conditions. We anticipate that PERT will be widely applicable to expression deconvolution strategies that use profiles from reference populations that vary from the corresponding constituent populations in cellular state but not cellular phenotypic identity.

  12. PERT: a method for expression deconvolution of human blood samples from varied microenvironmental and developmental conditions.

    Science.gov (United States)

    Qiao, Wenlian; Quon, Gerald; Csaszar, Elizabeth; Yu, Mei; Morris, Quaid; Zandstra, Peter W

    2012-01-01

    The cellular composition of heterogeneous samples can be predicted using an expression deconvolution algorithm to decompose their gene expression profiles based on pre-defined, reference gene expression profiles of the constituent populations in these samples. However, the expression profiles of the actual constituent populations are often perturbed from those of the reference profiles due to gene expression changes in cells associated with microenvironmental or developmental effects. Existing deconvolution algorithms do not account for these changes and give incorrect results when benchmarked against those measured by well-established flow cytometry, even after batch correction was applied. We introduce PERT, a new probabilistic expression deconvolution method that detects and accounts for a shared, multiplicative perturbation in the reference profiles when performing expression deconvolution. We applied PERT and three other state-of-the-art expression deconvolution methods to predict cell frequencies within heterogeneous human blood samples that were collected under several conditions (uncultured mono-nucleated and lineage-depleted cells, and culture-derived lineage-depleted cells). Only PERT's predicted proportions of the constituent populations matched those assigned by flow cytometry. Genes associated with cell cycle processes were highly enriched among those with the largest predicted expression changes between the cultured and uncultured conditions. We anticipate that PERT will be widely applicable to expression deconvolution strategies that use profiles from reference populations that vary from the corresponding constituent populations in cellular state but not cellular phenotypic identity.

  13. Unsafe Child Feces Disposal is Associated with Environmental Enteropathy and Impaired Growth.

    Science.gov (United States)

    George, Christine Marie; Oldja, Lauren; Biswas, Shwapon; Perin, Jamie; Sack, R Bradley; Ahmed, Shahnawaz; Shahnaij, Mohammad; Haque, Rashidul; Parvin, Tahmina; Azmi, Ishrat J; Bhuyian, Sazzadul Islam; Talukder, Kaisar A; Faruque, Abu G

    2016-09-01

    To investigate the relationship between unsafe child feces disposal, environmental enteropathy, and impaired growth, we conducted a prospective cohort study of 216 young children in rural Bangladesh. Using a prospective cohort study design in rural Bangladesh, unsafe child feces disposal, using the Joint Monitoring Program definition, was assessed using 5-hour structured observation by trained study personnel as well as caregiver reports. Anthropometric measurements were collected at baseline and at a 9-month follow-up. Stool was analyzed for fecal markers of environmental enteropathy: alpha-1-antitrypsin, myeloperoxidase, neopterin (combined to form an environmental enteropathy disease activity score), and calprotectin. Among 216 households with young children, 84% had an unsafe child feces disposal event during structured observation and 75% had caregiver reported events. There was no significant difference in observed unsafe child feces disposal events for households with or without an improved sanitation option (82% vs 85%, P = .72) or by child's age (P = .96). Children in households where caregivers reported unsafe child feces disposal had significantly higher environmental enteropathy scores (0.82-point difference, 95% CI 0.11-1.53), and significantly greater odds of being wasted (weight-for-height z score <-2 SDs) (9% vs 0%, P = .024). In addition, children in households with observed unsafe feces disposal had significantly reduced change in weight-for-age z-score (-0.34 [95% CI -0.68, -0.01] and weight-for-height z score (-0.52 [95% CI -0.98, -0.06]). Unsafe child feces disposal was significantly associated with environmental enteropathy and impaired growth in a pediatric population in rural Bangladesh. Interventions are needed to reduce this high-risk behavior to protect the health of susceptible pediatric populations. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Analysis of persistence of human papillomavirus infection in men evaluated by sampling multiple genital sites.

    Science.gov (United States)

    Capra, G; Nyitray, A G; Lu, B; Perino, A; Marci, R; Schillaci, R; Matranga, D; Firenze, A; Caleca, M; Bellavia, C; Guarneri, F; Giuliano, A; Giovannelli, L

    2015-11-01

    Although human papillomavirus (HPV) infection has been studied extensively in women, data on male infection are limited. The purpose of this study was to investigate persistence of HPV infection at multiple genital sites in men and to define potential associations with socio-behavioural characteristics. Penile, urethral and seminal specimens were tested by the INNO-LiPA HPV system (Innogenetics) and a PCR assay. Persistence was defined as the detection of same HPV type at ≥ 2 consecutive visits. The Kaplan-Meier method and the log-rank test were applied to estimate the likelihood of persistence. A total of 50 men (median age: 33 years) were followed for a median of 14.7 months. Altogether, 49%, 36%, 26% and 11% of baseline HPV-positive men had 6-, 12-, 18- and 24-month persistent infection with any HPV type, respectively. The 6-, 12- and 18- month persistence was more common for oncogenic HPV infections; 24-month persistence was similar. The median duration of persistence was 21.7 months for any HPV. The median duration of persistence for any HPV type was significantly longer in the penile sample (22.5 months, 95% CI: 18.3-26.7) than the semen sample (15.3 months, 95% CI: 14.5-16.1). Over a third of type-specific HPV infections in men remained persistent over a 24-month period. The median duration of HPV infection was longer in penile samples compared to seminal samples. As being increasing the attention of HPV vaccination as a potential preventive approach also for men, it is imperative to obtain additional insight on natural history of HPV infection in men, particularly as far as incidence and duration are concerned.

  15. Comparison of two adult mosquito sampling methods with human landing catches in south-central Ethiopia.

    Science.gov (United States)

    Kenea, Oljira; Balkew, Meshesha; Tekie, Habte; Gebre-Michael, Teshome; Deressa, Wakgari; Loha, Eskindir; Lindtjørn, Bernt; Overgaard, Hans J

    2017-01-13

    The human landing catch (HLC) is the standard reference method for measuring human exposure to mosquito bites. However, HLC is labour-intensive, exposes collectors to infectious mosquito bites and is subjected to collector bias. These necessitate local calibration and application of alternative methods. This study was undertaken to determine the relative sampling efficiency (RSE) of light traps with or without yeast-produced carbon dioxide bait vs. HLC in south-central Ethiopia. The experiment was conducted for 39 nights in a 3 × 3 Latin square randomized design with Anopheles arabiensis as the target species in the period between July and November 2014 in Edo Kontola village, south-central Ethiopia. Center for Disease Control and Prevention light trap catches (LTC) and yeast-generated carbon dioxide-baited light trap catches (CB-LTC) were each evaluated against HLC. The total nightly mosquito catches for each Anopheles species in either method was compared with HLC by Pearson correlation and simple linear regression analysis on log-transformed [log10(x + 1)] values. To test if the RSE of each alternative method was affected by mosquito density, the ratio of the number of mosquitoes in each method to the number of mosquitoes in HLC was plotted against the average mosquito abundance. Overall, 7606 Anopheles females were collected by the three sampling methods. Among these 5228 (68.7%) were Anopheles ziemanni, 1153 (15.2%) An. arabiensis, 883 (11.6%) Anopheles funestus s.l., and 342 (4.5%) Anopheles pharoensis. HLC yielded 3392 (44.6%), CB-LTC 2150 (28.3%), and LTC 2064 (27.1%) Anopheles females. The RSEs of LTC and HLC for An. arabiensis were significantly correlated (p method for sampling An. arabiensis. LTC can be used for large-scale indoor An. arabiensis surveillance and monitoring when it is difficult to use HLC. CB-LTC does not substantially improve sampling of this major vector compared to LTC in this setting. Trial registration PACTR201411000882128

  16. In Vitro Efficient Expansion of Tumor Cells Deriving from Different Types of Human Tumor Samples

    Directory of Open Access Journals (Sweden)

    Ilaria Turin

    2014-03-01

    Full Text Available Obtaining human tumor cell lines from fresh tumors is essential to advance our understanding of antitumor immune surveillance mechanisms and to develop new ex vivo strategies to generate an efficient anti-tumor response. The present study delineates a simple and rapid method for efficiently establishing primary cultures starting from tumor samples of different types, while maintaining the immuno-histochemical characteristics of the original tumor. We compared two different strategies to disaggregate tumor specimens. After short or long term in vitro expansion, cells analyzed for the presence of malignant cells demonstrated their neoplastic origin. Considering that tumor cells may be isolated in a closed system with high efficiency, we propose this methodology for the ex vivo expansion of tumor cells to be used to evaluate suitable new drugs or to generate tumor-specific cytotoxic T lymphocytes or vaccines.

  17. Determination of Human Chorionic Gonadotropin in Postmortem Samples in Ectopic Pregnancies.

    Science.gov (United States)

    Palmiere, Cristian; Lesta, Maria del Mar; Fanton, Laurent; Ventura, Francesco; Bonsignore, Alessandro; Reggiani Bonetti, Luca

    2016-01-01

    Increased human chorionic gonadotropin levels (HCG) can be detected in femoral blood, bile, and vitreous humor collected during autopsy of pregnant women using a standard kit designed for living patients. In the study herein, the concentrations of HCG were measured in postmortem serum, vitreous, bile, cerebrospinal, and pericardial fluids in 4 cases of fatal ectopic pregnancy and 40 controls using a quantitative electrochemiluminescence immunoassay designed for living patients. No false-negative cases were identified in any of the analyzed samples in any of the ectopic pregnancy cases. No correlations were found between total HCG levels in postmortem serum and the other tested specimens. The results of this study would suggest that higher HCG in bile, vitreous, pericardial, and cerebrospinal fluids may confirm the existence of ectopic pregnancy and therefore identify other situations in which this hormone is increased, although gestational age cannot be reliably estimated using these values. © 2015 American Academy of Forensic Sciences.

  18. Ultrasensitive PCR and real-time detection from human genomic samples using a bidirectional flow microreactor.

    Science.gov (United States)

    Chen, Lin; West, Jonathan; Auroux, Pierre-Alain; Manz, Andreas; Day, Philip J R

    2007-12-01

    In this paper we present a reliable bidirectional flow DNA amplification microreactor for processing real-world genomic samples. This system shares the low-power thermal responsiveness of a continuous flow reactor with the low surface area to volume ratio character of stationary reactors for reducing surface inhibitory effects. Silanization with dimethyldichlorosilane in combination with dynamic surface passivation was used to enhance PCR compatibility and enable efficient amplification. For real-time fragment amplification monitoring we have implemented an epimodal fluorescent detection capability. The passivated bidirectional flow system was ultrasensitive, achieving an RNase P gene detection limit of 24 human genome copies with a reaction efficiency of 77%. This starts to rival the performance of a conventional real-time PCR instrument with a reaction efficiency of 93% and revitalizes flow-through PCR as a viable component of lab on a chip DNA analysis formats.

  19. Analyses of robotic traverses and sample sites in the Schrödinger basin for the HERACLES human-assisted sample return mission concept

    Science.gov (United States)

    Steenstra, Edgar S.; Martin, Dayl J. P.; McDonald, Francesca E.; Paisarnsombat, Sarinya; Venturino, Christian; O'Hara, Sean; Calzada-Diaz, Abigail; Bottoms, Shelby; Leader, Mark K.; Klaus, Kurt K.; van Westrenen, Wim; Needham, Debra H.; Kring, David A.

    2016-09-01

    The International Space Exploration Coordination Group (ISECG) developed an integrated Global Exploration Roadmap (GER) that outlines plans for human-assisted sample return from the lunar surface in ∼2024 and for human presence on the lunar surface in ∼2028. Previous studies have identified the Schrödinger basin, situated on the far side of the Moon, as a prime target for lunar science and exploration where a significant number of the scientific concepts reviewed by the National Research Council (NRC, 2007) can be addressed. In this study, two robotic mission traverses within the Schrödinger basin are proposed based on a 3 year mission plan in support of the HERACLES human-assisted sample return mission concept. A comprehensive set of modern remote sensing data (LROC imagery, LOLA topography, M3 and Clementine spectral data) has been integrated to provide high-resolution coverage of the traverses and to facilitate identification of specific sample localities. We also present a preliminary Concept of Operations (ConOps) study based on a set of notional rover capabilities and instrumental payload. An extended robotic mission to the Schrödinger basin will allow for significant sample return opportunities from multiple distinct geologic terrains and will address multiple high-priority NRC (2007) scientific objectives. Both traverses will offer the first opportunity to (i) sample pyroclastic material from the lunar farside, (ii) sample Schrödinger impact melt and test the lunar cataclysm hypothesis, (iii) sample deep crustal lithologies in an uplifted peak ring and test the lunar magma ocean hypothesis and (iv) explore the top of an impact melt sheet, enhancing our ability to interpret Apollo samples. The shorter traverse will provide the first opportunity to sample farside mare deposits, whereas the longer traverse has significant potential to collect SPA impact melt, which can be used to constrain the basin-forming epoch. These robotic missions will revalidate

  20. Grazing behavior and spatial distribution of feces of Young bulls in silvopastoral systems and Marandu monoculture in the Pre-Amazon region

    Directory of Open Access Journals (Sweden)

    Ricardo Alves de Araújo

    2017-02-01

    Full Text Available The objective was to evaluate the grazing behavior and the spatial distribution of feces of F1 young bulls from the cross between Nellore and Guzera on pastures of Urochloa brizantha cv. Marandu in silvopastoral systems composed of babassu palm (Attalea speciosa and Marandu monoculture in the Pre-Amazon region of the state of Maranhão. Animals were evaluated in four systems consisting of 0, 80, 131, 160 palms ha-1, characterizing monoculture (mono, low density of palm trees (LD, medium density of palm trees (MD and high density of palm trees (HD during the rainy (RE and dry (DE periods. Five animals (repetitions were used in each system, with 231-303 days of age and 180±15 kg body weight. Determinations of behavioral patterns were made by instant sampling, at 10 min intervals. In each system, we demarcated 50 squares of 100 m2, which served as useful area to evaluate the dispersion of feces. The grazing behavior was influenced by the sward structure, which, in turn, was influenced by densities of palm trees, due to shading. The distribution of feces was affected by both the presence of babassu plantations and periods. The silvopastoral systems made the environment more pleasant to animals, since activities considered more stressful and avoided during the daytime were performed by animals of these environments, unlike animals in the monoculture system.

  1. Comparison of Droplet Digital PCR and qPCR for the Quantification of Shiga Toxin-Producing Escherichia coli in Bovine Feces.

    Science.gov (United States)

    Verhaegen, Bavo; De Reu, Koen; De Zutter, Lieven; Verstraete, Karen; Heyndrickx, Marc; Van Coillie, Els

    2016-05-18

    Cattle are considered to be the main reservoir for Shiga toxin-producing Escherichia coli (STEC) and are often the direct or indirect source of STEC outbreaks in humans. Accurate measurement of the concentration of shed STEC in cattle feces could be a key answer to questions concerning transmission of STEC, contamination sources and efficiency of treatments at farm level. Infected animals can be identified and the contamination level quantified by real-time quantitative PCR (qPCR), which has its specific limitations. Droplet digital PCR (ddPCR) has been proposed as a method to overcome many of the drawbacks of qPCR. This end-point amplification PCR is capable of absolute quantification independent from any reference material and is less prone to PCR inhibition than qPCR. In this study, the qPCR-based protocol described by Verstraete et al. (2014) for Shiga toxin genes stx1 and stx2 and the intimin gene eae quantification was optimized for ddPCR analysis. The properties of ddPCR and qPCR using two different mastermixes (EMM: TaqMan(®) Environmental Master Mix 2.0; UMM: TaqMan(®) Universal PCR Master Mix) were evaluated, using standard curves and both artificial and natural contaminated cattle fecal samples. In addition, the susceptibility of these assays to PCR-inhibitors was investigated. Evaluation of the standard curves and both artificial and natural contaminated cattle fecal samples suggested a very good agreement between qPCR using EMM and ddPCR. Furthermore, similar sensitivities and no PCR inhibition were recorded for both assays. On the other hand, qPCR using UMM was clearly prone to PCR inhibition. In conclusion, the ddPCR technique shows potential for the accurate absolute quantification of STEC on the farms, without relying on standardized reference material.

  2. Comparison of Droplet Digital PCR and qPCR for the Quantification of Shiga Toxin-Producing Escherichia coli in Bovine Feces

    Directory of Open Access Journals (Sweden)

    Bavo Verhaegen

    2016-05-01

    Full Text Available Cattle are considered to be the main reservoir for Shiga toxin-producing Escherichia coli (STEC and are often the direct or indirect source of STEC outbreaks in humans. Accurate measurement of the concentration of shed STEC in cattle feces could be a key answer to questions concerning transmission of STEC, contamination sources and efficiency of treatments at farm level. Infected animals can be identified and the contamination level quantified by real-time quantitative PCR (qPCR, which has its specific limitations. Droplet digital PCR (ddPCR has been proposed as a method to overcome many of the drawbacks of qPCR. This end-point amplification PCR is capable of absolute quantification independent from any reference material and is less prone to PCR inhibition than qPCR. In this study, the qPCR-based protocol described by Verstraete et al. (2014 for Shiga toxin genes stx1 and stx2 and the intimin gene eae quantification was optimized for ddPCR analysis. The properties of ddPCR and qPCR using two different mastermixes (EMM: TaqMan® Environmental Master Mix 2.0; UMM: TaqMan® Universal PCR Master Mix were evaluated, using standard curves and both artificial and natural contaminated cattle fecal samples. In addition, the susceptibility of these assays to PCR-inhibitors was investigated. Evaluation of the standard curves and both artificial and natural contaminated cattle fecal samples suggested a very good agreement between qPCR using EMM and ddPCR. Furthermore, similar sensitivities and no PCR inhibition were recorded for both assays. On the other hand, qPCR using UMM was clearly prone to PCR inhibition. In conclusion, the ddPCR technique shows potential for the accurate absolute quantification of STEC on the farms, without relying on standardized reference material.

  3. Comparison of stool containment in cloth and single-use diapers using a simulated infant feces.

    Science.gov (United States)

    Kubiak, M; Kressner, B; Raynor, W; Davis, J; Syverson, R E

    1993-03-01

    Single-use diapers and cloth diapers with vinyl pants were compared for their relative abilities to contain stool within the diaper. Artificial feces with carbon black as an additive allowed a quantitative measure of fecal containment by image analysis in 60 infants. This method showed complete containment of feces in the diaper in 50% of the single-use diapers whereas only 10% of the cloth diapers showed complete containment. In infants where the border of the vinyl pants was used as the boundary of containment with the cloth diapers, complete containment occurred only 33% of the time. Fluorescein dye ratings for containment/leakage in 69 infants showed that 83% of single-use diapers and 30% of the cloth diapers were rated as having no or minor leakage of feces. Cultures were taken of laundered vinyl pants that had previously been used over cloth diapers to determine microbial contamination. Thirty-nine percent of the pants contained Gram-negative, lactose-fermenting bacilli indicating fecal contamination. This study comparing single-use and cloth diapers for containment of artificial feces by use of image analysis and fluorescein dye ratings showed better containment by single-use diapers. The study also raises the question of possible spread of feces-borne pathogens by the vinyl pants used over cloth diapers, particularly in a day-care center.

  4. Spore dispersal of fetid Lysurus mokusin by feces of mycophagous insects.

    Science.gov (United States)

    Chen, Gao; Zhang, Rui-Rui; Liu, Yang; Sun, Wei-Bang

    2014-08-01

    The ecological roles and biological mechanisms of zoochory in plants have long been foci in studies of co-evolutionary processes between plants and animals. However, the dispersal of fungal spores by animals has received comparatively little attention. In this study, the dispersal of spores of a selected fetid fungus, Lysurus mokusin, via feces of mycophagous insects was explored by: collecting volatiles emitted by the fungus using dynamic headspace extraction and analyzing them by GC-MS; testing the capacity of mycophagous insects to disperse its spores by counting spores in their feces; comparing the germinability of L. mokusin spores extracted from feces of nocturnal earwigs and natural gleba of the fungus; and assessing the ability of L. mokusin volatiles to attract insects in bioassays with synthetic scent mixtures. Numerous spores were detected in insects' feces, the bioassays indicated that L. mokusin odor (similar to that of decaying substances) attracts diverse generalist mycophagous insects, and passage through the gut of Anisolabis maritima earwigs significantly enhanced the germination rate of L. mokusin spores. Therefore, nocturnal earwigs and diurnal flies probably play important roles in dispersal of L. mokusin spores, and dispersal via feces may be an important common dispersal mechanism for fungal reproductive tissue.

  5. Integrating Multiple Analytical Datasets to Compare Metabolite Profiles of Mouse Colonic-Cecal Contents and Feces.

    Science.gov (United States)

    Zeng, Huawei; Grapov, Dmitry; Jackson, Matthew I; Fahrmann, Johannes; Fiehn, Oliver; Combs, Gerald F

    2015-09-11

    The pattern of metabolites produced by the gut microbiome comprises a phenotype indicative of the means by which that microbiome affects the gut. We characterized that phenotype in mice by conducting metabolomic analyses of the colonic-cecal contents, comparing that to the metabolite patterns of feces in order to determine the suitability of fecal specimens as proxies for assessing the metabolic impact of the gut microbiome. We detected a total of 270 low molecular weight metabolites in colonic-cecal contents and feces by gas chromatograph, time-of-flight mass spectrometry (GC-TOF) and ultra-high performance liquid chromatography, quadrapole time-of-flight mass spectrometry (UPLC-Q-TOF). Of that number, 251 (93%) were present in both types of specimen, representing almost all known biochemical pathways related to the amino acid, carbohydrate, energy, lipid, membrane transport, nucleotide, genetic information processing, and cancer-related metabolism. A total of 115 metabolites differed significantly in relative abundance between both colonic-cecal contents and feces. These data comprise the first characterization of relationships among metabolites present in the colonic-cecal contents and feces in a healthy mouse model, and shows that feces can be a useful proxy for assessing the pattern of metabolites to which the colonic mucosum is exposed.

  6. Integrating Multiple Analytical Datasets to Compare Metabolite Profiles of Mouse Colonic-Cecal Contents and Feces

    Directory of Open Access Journals (Sweden)

    Huawei Zeng

    2015-09-01

    Full Text Available The pattern of metabolites produced by the gut microbiome comprises a phenotype indicative of the means by which that microbiome affects the gut. We characterized that phenotype in mice by conducting metabolomic analyses of the colonic-cecal contents, comparing that to the metabolite patterns of feces in order to determine the suitability of fecal specimens as proxies for assessing the metabolic impact of the gut microbiome. We detected a total of 270 low molecular weight metabolites in colonic-cecal contents and feces by gas chromatograph, time-of-flight mass spectrometry (GC-TOF and ultra-high performance liquid chromatography, quadrapole time-of-flight mass spectrometry (UPLC-Q-TOF. Of that number, 251 (93% were present in both types of specimen, representing almost all known biochemical pathways related to the amino acid, carbohydrate, energy, lipid, membrane transport, nucleotide, genetic information processing, and cancer-related metabolism. A total of 115 metabolites differed significantly in relative abundance between both colonic-cecal contents and feces. These data comprise the first characterization of relationships among metabolites present in the colonic-cecal contents and feces in a healthy mouse model, and shows that feces can be a useful proxy for assessing the pattern of metabolites to which the colonic mucosum is exposed.

  7. Bacterial populations and metabolites in the feces of free roaming and captive grizzly bears.

    Science.gov (United States)

    Schwab, Clarissa; Cristescu, Bogdan; Boyce, Mark S; Stenhouse, Gordon B; Gänzle, Michael

    2009-12-01

    Gut physiology, host phylogeny, and diet determine the composition of the intestinal microbiota. Grizzly bears (Ursus arctos horribilis) belong to the Order Carnivora, yet feed on an omnivorous diet. The role of intestinal microflora in grizzly bear digestion has not been investigated. Microbiota and microbial activity were analysed from the feces of wild and captive grizzly bears. Bacterial composition was determined using culture-dependent and culture-independent methods. The feces of wild and captive grizzly bears contained log 9.1 +/- 0.5 and log 9.2 +/- 0.3 gene copies x g(-1), respectively. Facultative anaerobes Enterobacteriaceae and enterococci were dominant in wild bear feces. Among the strict anaerobes, the Bacteroides-Prevotella-Porphyromonas group was most prominent. Enterobacteriaceae were predominant in the feces of captive grizzly bears, at log 8.9 +/- 0.5 gene copies x g(-1). Strict anaerobes of the Bacteroides-Prevotella-Porphyromonas group and the Clostridium coccoides cluster were present at log 6.7 +/- 0.9 and log 6.8 +/- 0.8 gene copies x g(-1), respectively. The presence of lactate and short-chain fatty acids (SCFAs) verified microbial activity. Total SCFA content and composition was affected by diet. SCFA composition in the feces of captive grizzly bears resembled the SCFA composition of prey-consuming wild animals. A consistent data set was obtained that associated fecal microbiota and metabolites with the distinctive gut physiology and diet of grizzly bears.

  8. Trace samples of human blood in mosquitoes as a forensic investigation tool.

    Science.gov (United States)

    Rabêlo, K C N; Albuquerque, C M R; Tavares, V B; Santos, S M; Souza, C A; Oliveira, T C; Oliveira, N C L; Crovella, S

    2015-11-23

    Investigations of any type of crime invariably starts at the crime scene by collecting evidence. Thus, the purpose of this research was to collect and analyze an entomological trace from an environment that is similar to those of indoor crime scenes. Hematophagous mosquitoes were collected from two residential units; saliva of volunteers that were residents in the units was also collected for genetic analysis as reference samples. We examined the allele frequencies of 15 short tandem repeat loci (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, vWA, TPOX, D18S51, D5S818, and FGA) and amelogenin. A total of 26 female hematophagous mosquitoes were identified as Aedes aegypti, Aedes albopictus, and Culex quinquefasciatus; we were able to obtain 11 forensically valid genetic profiles, with a minimum of 0.028203 ng/μL of human DNA. Thus, the results of this study showed that it was possible to correlate human genetic information from mosquitoes with the volunteer reference samples, which validates the use of this information as forensic evidence. Furthermore, we observed mixed genetic profiles from one mosquito. Therefore, it is clearly important to collect these insects indoors where crimes were committed, because it may be possible to find intact genetic profiles of suspects in the blood found in the digestive tract of hematophagous mosquitoes for later comparison to identify an offender and/or exclude suspects.

  9. Assessment of pregnancy status of Asian elephants (Elephas maximus) by measurement of progestagen and glucocorticoid and their metabolite concentrations in serum and feces, using enzyme immunoassay (EIA).

    Science.gov (United States)

    Kajaysri, Jatuporn; Nokkaew, Weerapun

    2014-03-01

    The study was to find patterns of progestagen (progesterone and its metabolite) and glucocorticoid and their metabolite concentrations in serum and feces of pregnant Asian elephants (Elephas maximus). The 5 female Asian domestic elephants were naturally mated until pregnancy. After that, blood and feces samples were collected monthly during pregnancy for progestagen, glucocorticoid and their metabolites analysis by enzyme immunoassay (EIA). The results showed the serum progestagen concentration during gestation was 2.11 ± 0.60 to 18.44 ± 2.28 ng/ml. Overall, serum progestagen concentration rose from the 1st month to reach peak in the 11th month, after which it declined to its lowest level in the 22nd month of pregnancy. Fecal progestagen concentration varied from 1.18 ± 0.54 to 3.35 ± 0.45 µg/g during pregnancy. In general, fecal progestagen concentration increased from the 1st month to its highest level in the 12th month. After this, it declined reaching its lowest point in the 22nd month of pregnancy. Glucocorticoid hormones and their metabolite concentrations both in serum and feces fluctuated from low to medium throughout almost the entire pregnancy period and then rapidly increased around the last week before calving. Our study suggests that this profile of progestagen and glucocorticoid hormones and their metabolite concentration levels in serum and feces can be used to assess the pregnancy status of Asian elephants. If serum and fecal progestagen concentrations were found in very low levels and glucocorticoid and their metabolite concentrations were found in very high levels, it was indicated that the cow elephant would calve within 7 days.

  10. Characterization of quinolone resistance in Salmonella spp. isolates from food products and human samples in Brazil

    Science.gov (United States)

    Pribul, Bruno Rocha; Festivo, Marcia Lima; de Souza, Miliane Moreira Soares; dos Prazeres Rodrigues, Dalia

    2016-01-01

    Non-typhoidal salmonellosis is an important zoonotic disease caused by Salmonella enterica. The aim of this study was to investigate the prevalence of plasmid-mediated quinolone resistance in Salmonella spp. and its association with fluoroquinolone susceptibility in Brazil. A total of 129 NTS isolates (samples from human origin, food from animal origin, environmental, and animal) grouped as from animal (n = 62) and human (n = 67) food were evaluated between 2009 and 2013. These isolates were investigated through serotyping, antimicrobial susceptibility testing, and the presence of plasmid-mediated quinolone resistance (PMQR) genes (qnr, aac(6′)-Ib) and associated integron genes (integrase, and conserved integron region). Resistance to quinolones and/or fluoroquinolones, from first to third generations, was observed. Fifteen isolates were positive for the presence of qnr genes (8 qnrS, 6 qnrB, and 1 qnrD) and twenty three of aac(6′)-Ib. The conserved integron region was detected in 67 isolates as variable regions, from ±600 to >1000 pb. The spread of NTS involving PMQR carriers is of serious concern and should be carefully monitored. PMID:26887245

  11. Assessment of malathion and its effects on leukocytes in human blood samples

    Science.gov (United States)

    Sharma, Amit Kumar; Tiwari, Udita; Gaur, Mulayam Singh; Tiwari, Rajeev Kumar

    2016-01-01

    Abstract In the present paper, we report a reproducible, cost effective, fast response method for detection of malathion and its effects on leukocytes in different human blood groups. Spectroscopic methods (UV-Vis spectrometry) and Fourier transform infrared coupled with solid phase extraction were applied for analyzing malathion content in human blood plasma. The spiking levels of malathion in the range of 0.1-1.7 µg/mL were extracted from blood plasma samples using SPE. The present active functional groups (C = O; P-O-C; -OH; P = S) were also characterized. The recovery rate of malathion was 80%±4.5%. The calculated correlation coefficient was 0.9799, indicating the linearity of the results. The limit of detection (LOD) and limit of quantification (LOQ) were (0.1-1.7) µg/mL and (0.3-1.5) µg/mL, respectively. Malathion <1.0 µg/mL showed no significant change while higher levels of malathion exposure (1.5 µg/mL and 3.0 µg/mL) reduced the number of white blood cells. In conclusion, the spectroscopic results may be useful to understand the mechanism of other pesticides such as methyl parathion and parathion.

  12. Characterization of quinolone resistance in Salmonella spp. isolates from food products and human samples in Brazil.

    Science.gov (United States)

    Pribul, Bruno Rocha; Festivo, Marcia Lima; de Souza, Miliane Moreira Soares; Rodrigues, Dalia dos Prazeres

    2016-01-01

    Non-typhoidal salmonellosis is an important zoonotic disease caused by Salmonella enterica. The aim of this study was to investigate the prevalence of plasmid-mediated quinolone resistance in Salmonella spp. and its association with fluoroquinolone susceptibility in Brazil. A total of 129 NTS isolates (samples from human origin, food from animal origin, environmental, and animal) grouped as from animal (n=62) and human (n=67) food were evaluated between 2009 and 2013. These isolates were investigated through serotyping, antimicrobial susceptibility testing, and the presence of plasmid-mediated quinolone resistance (PMQR) genes (qnr, aac(6')-Ib) and associated integron genes (integrase, and conserved integron region). Resistance to quinolones and/or fluoroquinolones, from first to third generations, was observed. Fifteen isolates were positive for the presence of qnr genes (8 qnrS, 6 qnrB, and 1 qnrD) and twenty three of aac(6')-Ib. The conserved integron region was detected in 67 isolates as variable regions, from ±600 to >1000pb. The spread of NTS involving PMQR carriers is of serious concern and should be carefully monitored.

  13. Determination of human albumin in serum and urine samples by constant-energy synchronous fluorescence method.

    Science.gov (United States)

    Madrakian, Tayyebeh; Bagheri, Habibollah; Afkhami, Abbas

    2015-08-01

    A sensitive spectrofluorimetric method using constant-energy synchronous fluorescence technique is proposed for the determination of human albumin without separation. In this method, no reagent was used for enhancement of the fluorescence signal of albumin in the solution. Effects of some parameters, such as energy difference between excitation and emission monochromators (ΔE), emission and excitation slit widths and scan rate of wavelength were studied and the optimum conditions were established. For this purpose factorial design and response surface method were employed for optimization of the effective parameters on the fluorescence signal. The results showed that the scan rate of the wavelength has no significant effect on the analytical signal. The calibration curve was linear in the range 0.1-220.0 µg mL(-1) of albumin with a detection limit of 7.0 × 10(-3)  µg mL(-1). The relative standard deviations (RSD) for six replicate measurements of albumin were calculated as 2.2%, 1.7% and 1.3% for 0.5, 10.0 and 100.0 µg mL(-1) albumin, respectively. Furthermore the proposed method has been employed for the determination of albumin in human serum and urine samples.

  14. Characterization of a Hemoglobin Adduct from Ethyl Vinyl Ketone Detected in Human Blood Samples.

    Science.gov (United States)

    Carlsson, Henrik; Motwani, Hitesh V; Osterman Golkar, Siv; Törnqvist, Margareta

    2015-11-16

    Electrophiles have the ability to form adducts to nucleophilic sites in proteins and DNA. Internal exposure to such compounds thus constitutes a risk for toxic effects. Screening of adducts using mass spectrometric methods by adductomic approaches offers possibilities to detect unknown electrophiles present in tissues. Previously, we employed untargeted adductomics to detect 19 unknown adducts to N-terminal valine in hemoglobin (Hb) in human blood. This article describes the characterization of one of these adducts, which was identified as the adduct from ethyl vinyl ketone (EVK). The mean adduct level was 40 ± 12 pmol/g Hb in 12 human blood samples; adduct levels from acrylamide (AA) and methyl vinyl ketone (MVK) were quantified for comparison. Using l-valine p-nitroanilide (Val-pNA), introduced as a model of the N-terminal valine, the rate of formation of the EVK adduct was studied, and the rate constant determined to 200 M(-1)h(-1) at 37 °C. In blood, the reaction rate was too fast to be feasibly measured, EVK showing a half-life adduct was found to be unstable, with a half-life of 7.6 h. From the mean adduct level measured in human blood, a daily dose (area under the concentration-time-curve, AUC) of 7 nMh EVK was estimated. The AUC of AA from intake via food is about 20 times higher. EVK is naturally present in a wide range of foods and is also used as a food additive. Most probably, naturally formed EVK is a major source to observed adducts. Evaluation of available toxicological data and information on occurrence of EVK indicate that further studies of EVK are motivated. This study illustrates a quantitative strategy in the initial evaluation of the significance of an adduct detected through adduct screening.

  15. Inactivation of human immunodeficiency virus type 1 in blood samples stored as high-salt lysates.

    Science.gov (United States)

    Zolg, J W; Lanciotti, R S; Wendlinger, M; Meyer, W A

    1990-09-01

    Blood samples to be tested for the presence of parasite DNA by using specific DNA probes are routinely stored in our laboratory as high-salt lysates (HSL). To safeguard against the risk of accidental infection with etiological agents such as the human immunodeficiency virus type 1 (HIV-1) while manipulating large numbers of blood samples in preparation for DNA probing, we determined the residual infectivity of HIV-1 after exposure to HSL components. Both high-titer virus stocks or provirus-carrying cells, suspended either in tissue culture medium or freshly drawn blood, were completely inactivated upon contact with the HSL components. This was verified by the absence of any detectable HIV-1-specific antigen in the supernatants of long-term cultures and the absence of virus-specific DNA fragments after amplification by polymerase chain reaction with DNA from such cultures as target DNA. These results support the conclusion that the virus is in fact completely inactivated by contact with the HSL components, rendering blood specimens stored as HSL noninfectious in regard to HIV-1.

  16. Aqueous two phase system based on ionic liquid for isolation of quinine from human plasma sample.

    Science.gov (United States)

    Flieger, J; Czajkowska-Żelazko, A

    2015-01-01

    Aqueous two phase system was applied for selective extraction of quinine from human plasma. Bi-phase was constructed from ionic liquid: butyl-methyl-imidazolium chloride after addition kosmotropic salts K₃PO₄ or KH₂PO₄. Quinine was determined in plasma samples after drinking of tonic containing quinine. Determination was performed by HPLC on 5-μm Zorbax SB-CN column and eluent containing 40% acetonitrile (v/v), 20 mM phosphate buffer at pH 3 and 40 mM NaPF₆ using external standard method. The spectrophotometric detection was set λ=214 nm. Selective fluorescence detection was performed at excitation of 325 nm and emission of 375 nm. Proposed strategy provides suitable sample purification and gives extraction yields in the range of 89-106%. The determination coefficient (R(2)) has a value ≥0.997 in the range of 50-800 ng/ml quinine concentration. The limit of quantification was set at 27.9 ng/ml and the detection limit was found to be 8.4 ng/ml under fluorescence detection.

  17. Analysis of Fluconazole in Human Urine Sample by High Performance Liquid Chromatography Method

    Science.gov (United States)

    Hermawan, D.; Ali, N. A. Md; Ibrahim, W. A. Wan; Sanagi, M. M.

    2013-04-01

    A method for determination of fluconazole, antifungal drug in human urine by using reversed-phased high performance liquid chromatography (RP-HPLC) with ultraviolet (UV) detector was developed. Optimization HPLC conditions were carried out by changing the flow rate and composition of mobile phase. The optimum separation conditions at a flow rate 0.85 mL/min with a composition of mobile phase containing methanol:water (70:30, v/v) with UV detection at a wavelength 254 nm was able to analyze fluconazole within 3 min. The excellent linearity was obtained in the range of concentration 1 to 10 μg/mL with r2 = 0.998. The limit of detection (LOD) and limit of quantitation (LOQ) were 0.39 μg/mL and 1.28 μg/mL, respectively. Solid phase extraction (SPE) method using octadecylsilane (C18) as a sorbent was used to clean-up and pre-concentrated of the urine sample prior to HPLC analysis. The average recoveries of fluconazole in spiked urine sample was 72.4% with RSD of 3.21% (n=3).

  18. Proteomic and oxidative stress analysis in human brain samples of Huntington disease.

    Science.gov (United States)

    Sorolla, Ma Alba; Reverter-Branchat, Gemma; Tamarit, Jordi; Ferrer, Isidre; Ros, Joaquim; Cabiscol, Elisa

    2008-09-01

    Huntington disease (HD) is a neurodegenerative disorder caused by expansion of CAG repeats in exon 1 of the huntingtin gene, affecting initially the striatum and progressively the cortex. This work reports a proteomic analysis of human brain postmortem samples obtained from striatum and cortex of patients with HD compared to samples of age- and sex-matched controls. Antioxidant defense proteins that were strongly induced in striatum, but also detectable in cortex, were identified as peroxiredoxins 1, 2, and 6, as well as glutathione peroxidases 1 and 6. The activities of other antioxidant enzymes such as mitochondrial superoxide dismutase and catalase were also increased in HD. Aconitase, a protein involved in energy metabolism, showed decreased activities in striatum of HD patients. Protein carbonyls, used as markers of oxidative stress, were increased in HD, and glial fibrillary acidic protein, aconitase, gamma-enolase, and creatine kinase B were identified as the main targets. Taken together, these results indicate that oxidative stress and damage to specific macromolecules would participate in the disease progression. Also, these data support the rationale for therapeutic strategies that either potentiate antioxidant defenses or avoid oxidative stress generation to delay disease progression.

  19. Prevalence and genotyping of high risk human papillomavirus in cervical cancer samples from Punjab, Pakistan.

    Science.gov (United States)

    Siddiqa, Abida; Zainab, Maidah; Qadri, Ishtiaq; Bhatti, Muhammad Faraz; Parish, Joanna L

    2014-07-17

    Cervical cancer is the third most common cause of cancer-related death in women worldwide. Infection with high-risk human papillomavirus (HPV) is established as the cause of cervical carcinoma, therefore, high risk HPV detection may have prognostic significance for the women who are at increased risk of disease progression. The paucity of data on the incidence of cervical cancer in Pakistan makes it difficult to determine disease burden. Even less information is available regarding the prevalent HPV strains in cervical specimens collected from this region. Cervical cancer is a neglected disease in Pakistan in terms of screening, prevention, and vaccination. Identification and accurate genotyping of the virus burden in cancer specimens is important to inform intervention policies for future management of HPV associated disease and to potentially stratify patients dependent on HPV status. In this study, detection and genotyping of HPV types 16 and 18 from 77 cervical specimens were carried out. Consensus primers GP5+/GP6+, which detect 44 genital HPV types, and type specific primers (TS16 and TS18) were used in conjunction with newly designed type specific primers. Using a combination of these methods of detection, a total of 94.81% (95% CI ±4.95) of cervical lesions were positive for HPV. Single infections of HPV16 were detected in 24.68% (95% CI ±9.63) of total samples and HPV18 was found in 25.97% (95% CI ±9.79) samples. Interestingly, a high proportion of samples (40.26%, 95% CI ±10.95) was positive for both HPV16 and 18, indicating a higher incidence of co-infection than previously reported for similar ethnic regions. The HPV genotype of 3.90% of HPV positive samples remained undetected, although these samples were positive with the GP5+/GP6+ primer set indicating infection with an HPV type other than 16 or 18. These data indicate that the overall incidence of high risk HPV infection in cervical cancer and intraepithelial neoplasia specimens in Punjab

  20. Fungal propagules and DNA in feces of two detritus-feeding amphipods.

    Science.gov (United States)

    Sridhar, Kandikere Ramaiah; Beaton, Margaret; Bärlocher, Felix

    2011-01-01

    Aquatic shredders (leaf-eating invertebrates) preferentially ingest and digest leaves colonized by aquatic hyphomycetes (fungi). This activity destroys leaf-associated fungal biomass and detritial resources in streams. Fungal counter-adaptations may include the ability to survive passage through the invertebrate's digestive tract. When fecal pellets of Gammarus tigrinus and Hyalella azteca were incubated with sterile leaves, spores of nine (G. tigrinus) and seven (H. azteca) aquatic hyphomycete species were subsequently released from the leaves, indicating the presence of viable fungal structures in the feces. Extraction, amplification, and sequencing of DNA from feces revealed numerous fungal phylotypes, two of which could be assigned unequivocally to an aquatic hyphomycete. The estimated contributions of major fungal groups varied depending on whether 18S or ITS sequences were amplified and cloned. We conclude that a variable proportion of fungal DNA in the feces of detritivores may originate from aquatic hyphomycetes. Amplified DNA may be associated with metabolically active, dormant, or dead fungal cells.

  1. Evaluation of extraction methods for progesterone determination in rabbit (Oryctolagus cuniculus) feces by radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Korndoerfer, Clotilde Maria; Meirelles, Cyro Ferreira; Bueno, Ives Claudio da Silva; Abdalla, Adibe Luiz [Centro de Energia Nuclear na Agricultura (CENA), Piracicaba, SP (Brazil). Secao de Ciencias Animais]. E-mail: cfmeirel@esalq.usp.br

    1998-07-01

    The purpose of this study was to find a practical procedure for the extraction of progesterone (P{sub 4}) from feces and to determine if the P4 plasma profiles during pregnancy were reflected in total fecal P4 of pregnant rabbits. The rabbit was used as model for the techniques. Plasma and feces were collected from 11 rabbits during a period of 42 days. Three different methods of P4 extraction were used. The total P4 was measured by solid-phase radioimmunoassay (RIA) with {sup 125} I-P4 as the tracer. Results suggested that it was possible to extract total P4 from rabbit feces with methanol and petroleum ether. Plasma and fecal P4 profiles were compared for both pregnant and ovariectomized rabbits. It was possible to differentiate total P4 extracted from day two through 28 after breeding (p<0.01). (author)

  2. Evaluation of acid digestion techniques to estimate chromium contents in cattle feces

    Directory of Open Access Journals (Sweden)

    Gabriel Cipriano Rocha

    2015-01-01

    Full Text Available The objective of this work was to evaluate the accuracy of digestion techniques using nitric and perchloric acid at the ratios of 2:1, 3:1, and 4:1 v v-1, in one- or two-step digestion, to estimate chromium contents in cattle feces, using sodium molybdate as a catalyst. Fecal standards containing known chromium contents (0, 2, 4, 6, 8, and 10 g kg-1 were produced from feces of five animals. The chromium content in cattle feces is accurately estimated using digestion techniques based on nitric and perchloric acids, at a 3:1 v v-1 ratio, in one-step digestion, with sodium molybdate as a catalyst.

  3. SEROTYPING AND ANTIMICROBIAL DRUG RESISTANCE OF SALMONELLA ISOLATED FROM LETTUCE AND HUMAN DIARRHEA SAMPLES IN BURKINA FASO.

    Science.gov (United States)

    Siourimè, Somda Namwin; Isidore, Bonkoungou Ouindgueta Juste; Oumar, Traoré; Nestor, Bassolé Ismael Henri; Yves, Traoré; Nicolas, Barro; Aly, Savadogo

    2017-01-01

    Background: In Burkina Faso dirty water in particular those of the stoppings and the gutter ones are used for vegetables irrigation in the gardens. The aim of this study was to determine the prevalence and antibiotic susceptibility of Salmonella serotypes from humans and lettuce samples inBurkina Faso. Materials and Methods:Salmonella strains isolated from patients in 2009 to 2015 and lettuce samples in 2014 in Burkina Faso were serotyped using specific antisera. All strains were subjected to a set of 14 antibiotics to study their antibiogram by using Baeur–Kirby disk diffusion method. Results: Out of 154 Salmonella isolated, 60 were from human and 94 from lettuce samples. Serotyping revealed four different serotypes and 39% (60) untypeable strains from human and lettuce (14 and 46 strains). Salmonella serotypes from human and lettuce samples were: Paratyphi A (10% and 22%), Paratyphi B (34% and 8%), Paratyphi C (14% and 18%) and Typhi (21% and 1%). A high resistance of Salmonella Paratyphi B and Salmonella spp to tetracycline were 70% from human and 35 % from lettuce samples. Multiresistance was observed to tetracycline, chloramphenicol and amoxicillin/clavulanic-acid or ampicillin with Salmonella ParatyphiB 35% and Salmonella Typhi 33% from human samples and Salmonella spp 4% from lettuce samples. Conclusion: This study showed the diversity of Salmonella serotypes from both clinical and environmental samples and emergence of multiresistant Salmonella to antibiotics in Burkina Faso. A lettuce is a potential source of transmission of Salmonella causing diarrhea among human in Burkina Faso. List of non-standard Abbreviations : HDB: Hôpital du District de Bogodogo, LNSP: Laboratoire National de Santé Publique, DSG : District Sanitaire de Gourcy, DSB : District Sanitaire de Boromo PMID:28670637

  4. Isolation and serotyping of Streptococcus mutans from teeth and feces of children.

    OpenAIRE

    Hamada, S; Masuda, N; Kotani, S

    1980-01-01

    Streptococcus mutans were detected in the feces from 10 of 29 caries-active patients, aged 4 to 9 years. The percentage of S. mutans to the total counts of facultatively anaerobic streptococci on mitis salivarius agar (Difco Laboratories) varied from 0 to 72.5%. S. mutans were then isolated from dental plaque of sound teeth and carious dentin of the 10 subjects known to harbor S. mutans in the feces. The frequency distribution of various serotypes of these dental and fecal isolates of S. muta...

  5. Tracking human activity and well-being in natural environments using wearable sensors and experience sampling.

    Science.gov (United States)

    Doherty, Sean T; Lemieux, Christopher J; Canally, Culum

    2014-04-01

    A growing range of studies have begun to document the health and well-being benefits associated with contact with nature. Most studies rely on generalized self-reports following engagement in the natural environment. The actual in-situ experience during contact with nature, and the environmental features and factors that evoke health benefits have remained relatively unexplored. Smartphones offer a new opportunity to monitor and interact with human subjects during everyday life using techniques such as Experience Sampling Methods (ESM) that involve repeated self-reports of experiences as they occur in-situ. Additionally, embedded sensors in smartphones such as Global Positioning Systems (GPS) and accelerometers can accurately trace human activities. This paper explores how these techniques can be combined to comprehensively explore the perceived health and well-being impacts of contact with nature. Custom software was developed to passively track GPS and accelerometer data, and actively prompt subjects to complete an ESM survey at regular intervals throughout their visit to a provincial park in Ontario, Canada. The ESM survey includes nine scale questions concerning moods and emotions, followed by a series of open-ended experiential questions that subjects provide recorded audio responses to. Pilot test results are used to illustrate the nature, quantity and quality of data obtained. Participant activities were clearly evident from GPS maps, including especially walking, cycling and sedate activities. From the ESM surveys, participants reported an average of 25 words per question, taking an average of 15 s to record them. Further qualitative analysis revealed that participants were willing to provide considerable insights into their experiences and perceived health impacts. The combination of passive and interactive techniques is sure to make larger studies of this type more affordable and less burdensome in the future, further enhancing the ability to understand

  6. Molecular epidemiology of human papillomavirus infections in cervical samples from cuban women older than 30 years.

    Science.gov (United States)

    Soto, Yudira; Torres, Griselda; Kourí, Vivian; Limia, Celia María; Goicolea, Adibel; Capó, Virginia; Pérez, Lissette; de la Torre, Ana Isabel; López, Ledy Xiomara; Govín, Anamays; Correa, Consuelo Beatriz; Alemán, Yoan; Alvarez, Alina Ana; Manzano, Blanca Rosa

    2014-07-01

    This study aimed to provide information about the molecular epidemiology of human papillomavirus (HPV) in a group of Cuban women. DNA from cervical samples was analyzed using a quantitative real-time polymerase chain reaction (PCR), which detects 6 of the clinically most relevant high-risk HPV types. Furthermore, end point PCR and sequencing were performed. Three hundred twenty-two women (211 with positive and 111 with negative cytologic results) aged between 30 and 69 years were enrolled. Risk factors associated with HPV infections and premalignant lesions were also investigated. HPV DNA was detected in 76.1% (245/322) of the studied population, and 34 different genotypes were found. There was an association between HPV infection and low educational level, history of oral contraceptives, menopausal stage, as well as cigarette and/or alcohol consumption. Besides, in a multivariate analysis, previous positive Pap test result and positive colposcopy finding were both predictor variables for HPV infections and for premalignant lesions. Human papillomavirus infection was found in 94.3% of women (199/211) with positive cytologic result and in 41.4% (46/111) of those with negative results, being more likely that the first group was infected with any HPV (odds ratio = 23.43; 95% CI = 11.70-46.92; p = .000). The most common genotypes were HPV types 16, 18, 31, 58, 33, and 45. All the cases with HPV positive findings had at least 1 high-risk HPV genotype. This is the first report of the molecular epidemiology of HPV in Cuban women, based on results from a DNA sequence and quantitative PCR. Most individuals were infected with high-risk HPV types. These findings support the inclusion of HPV vaccine in Cuba.

  7. Human papillomavirus testing by self-sampling: assessment of accuracy in an unsupervised clinical setting

    Science.gov (United States)

    Szarewski, Anne; Cadman, Louise; Mallett, Susan; Austin, Janet; Londesborough, Philip; Waller, Jo; Wardle, Jane; Altman, Douglas G; Cuzick, Jack

    2007-01-01

    Objectives: To compare the performance and acceptability of unsupervised self-sampling with clinician sampling for high-risk human papillomavirus (HPV) types for the first time in a UK screening setting. Setting: Nine hundred and twenty women, from two demographically different centres, attending for routine cervical smear testing Methods: Women performed an unsupervised HPV self-test. Immediately afterwards, a doctor or nurse took an HPV test and cervical smear. Women with an abnormality on any test were offered colposcopy. Results: Twenty-one high-grade and 39 low-grade cervical intraepithelial neoplasias (CINs) were detected. The sensitivity for high-grade disease (CIN2+) for the self HPV test was 81% (95% confidence interval [CI] 60–92), clinician HPV test 100% (95% CI 85–100), cytology 81% (95% CI 60–92). The sensitivity of both HPV tests to detect high- and low-grade cervical neoplasia was much higher than that of cytology (self-test 77% [95%CI 65–86], clinician test 80% [95% CI 68–88], cytology 48% [95% CI 36–61]). For both high-grade alone, and high and low grades together, the specificity was significantly higher for cytology (greater than 95%) than either HPV test (between 82% and 87%). The self-test proved highly acceptable to women and they reported that the instructions were easy to understand irrespective of educational level. Conclusions: Our results suggest that it would be reasonable to offer HPV self-testing to women who are reluctant to attend for cervical smears. This approach should now be directly evaluated among women who have been non-attenders in a cervical screening programme. PMID:17362570

  8. Levels and complexity of IgA antibody against oral bacteria in samples of human colostrum.

    Science.gov (United States)

    Petrechen, L N; Zago, F H; Sesso, M L T; Bertoldo, B B; Silva, C B; Azevedo, K P; de Lima Pereira, S A; Geraldo-Martins, V R; Ferriani, V P L; Nogueira, R D

    2015-01-01

    Streptococcus mutans (SM) have three main virulence antigens: glucan binding protein B (gbpB), glucosyltransferase (Gtf) and antigens I/II (Ag I/II) envolved in the capacity of those bacteria to adhere and accumulate in the dental biofilm. Also, the glycosyltransferases 153 kDa of Streptococcus gordonii (SGO) and 170kDa of Streptococcus sanguinis (SSA) were important antigens associated with the accumulation of those bacterias. Streptococcus mitis (SMI) present IgA1 protease of 202 kDa. We investigated the specificity and levels IgA against those antigens of virulence in samples of human colostrum. This study involved 77 samples of colostrum that were analyzed for levels of immunoglobulian A, M and G by Elisa. The specificity of IgA against extracts of SM and initials colonizators (SSA, SMI, SGO) were analyzed by the Western blot. The mean concentration of IgA was 2850.2 (±2567.2) mg/100 mL followed by IgM and IgG (respectively 321.8±90.3 and 88.3±51.5), statistically different (pbacteria antigens and theirs virulence antigens. To SM, the GbpB was significantly lower detected than others antigens of SM (p0.4). So, the breast milk from first hours after birth presented significant levels of IgA specific against important virulence of antigens those oral streptococci, which can disrupt the installation and accumulation process of these microorganisms in the oral cavity. Copyright © 2014 Elsevier GmbH. All rights reserved.

  9. Novel encapsulation improves recovery of probiotic strains in fecal samples of human volunteers.

    Science.gov (United States)

    Mai, Volker; Waugh, Sheldon; Byrd, Doratha; Simpson, Damion; Ukhanova, Maria

    2017-02-01

    Probiotic supplements can contribute to maintaining health and ameliorating various disease symptoms. Probiotics can be delivered in many forms with crucial differences in their survival during gastrointestinal (GI) passage. Previously, a novel encapsulation, Probiotic Pearls™ Acidophilus, Integrative Therapeutics, LLC, USA (Pearls), was shown to increase survival in vitro after exposure to gastric conditions. Here, we compare fecal recovery in human volunteers consuming Pearls or a conventional hard-shelled gelatin capsule. We performed a randomized double-blinded, two-armed trial, with six healthy subjects in each 12-day study arm. In fecal samples collected at baseline, twice during the intervention period, and after washout, we compared colony counts between the two encapsulation methods. The identity of the colonies was confirmed by colony morphology, strain-specific PCR, and 16S rRNA gene sequencing. We further performed a comprehensive 16S rRNA gene sequencing-based analysis to identify differential effects on overall microbiota composition. We detected an average log increase in bifidobacteria of 0.152 cfu/g with gelatin and 0.651 cfu/g with Pearls capsules (p > 0.05). Total lactobacilli counts increased in both groups with no difference between the groups. However, the supplemented Lactobacillus acidophilus NCFM decreased to baseline levels within 7 days after end of supplementation with gelatin capsules while 3.11 log cfu/g higher counts compared to baseline (p = 0.05) remained for Pearls. Targeted qPCR largely confirmed the trends observed by viable plate counts. Protecting the probiotic strains by Pearls encapsulation results in higher recovery rates of the supplemented lactobacilli and bifidobacteria in fecal samples and increased persistence, suggesting an improved survival and viability that might increase efficacy towards achieving desired health benefits.

  10. Detection of nandrolone, testosterone, and their esters in rat and human hair samples.

    Science.gov (United States)

    Höld, K M; Borges, C R; Wilkins, D G; Rollins, D E; Joseph, R E

    1999-10-01

    Nandrolone and testosterone are anabolic androgenic steroids occasionally abused by athletes. A sensitive, specific, and reproducible gas chromatography-mass spectrometry method for the quantitative determination of nandrolone, testosterone, and their esters in hair has been developed. The limits of quantitation of this method, based on 20 mg of hair, were 50 pg/mg for nandrolone and testosterone, 100 pg/mg for testosterone acetate, and 200 pg/mg for nandrolone-decanoate. Nandrolone-d3 and testosterone-d3 were used as internal standards. This method has been applied to the analysis of these compounds incorporated into rat and human hair. Male Long-Evans rats were given nandrolone decanoate 60 mg/kg intraperitoneally (i.p.) once daily for 10 days over a time period of 14 days. Two of the three rats contained nandrolone in the pigmented hair collected at day 21 at a concentration of 63 and 76 pg/mg, respectively. No drug was found in the corresponding nonpigmented hair. The rat hair samples that tested positive for nandrolone contained also nandrolone decanoate in concentrations of 0.9 and 1.2 ng/mg, respectively. In a separate experiment rats were given testosterone acetate 10 mg/kg i.p. once daily for five days. No testosterone or testosterone acetate was detected in the rat hair samples. Hair specimens were also obtained from four self-reported steroid users. The hair of two subjects were determined to be positive for testosterone in concentrations of 54 and 81 pg/mg. These data demonstrate that it is possible to detect the steroids nandrolone, testosterone, and nandrolone decanoate in hair after systemic administration.

  11. Prevalence of Human Papillomavirus Infection in Unselected SurePath Samples Using the APTIMA HPV mRNA Assay

    DEFF Research Database (Denmark)

    Rebolj, Matejka; Preisler, Sarah; Ejegod, Ditte M;

    2013-01-01

    The APTIMA Human Papillomavirus (HPV) Assay detects E6/E7 mRNA from 14 human papillomavirus genotypes. Horizon was a population-based split-sample study among well-screened women, with an aim to compare APTIMA, Hybrid Capture 2 (HC2), and liquid-based cytology (LBC) using SurePath samples. APTIMA...... agreement between APTIMA and HC2. This is the first APTIMA study using SurePath samples on the PANTHER platform. The trends in positivity rates on SurePath samples for APTIMA, HC2, and LBC were consistent with studies based on PreservCyt samples, and the agreement between the two HPV assays was substantial....... The high proportions of women testing positive suggest that in countries with a high HPV prevalence, caution will be needed if HPV tests, including mRNA-based tests, are to replace LBC....

  12. Sensitivity of PCR assays for murine gammaretroviruses and mouse contamination in human blood samples.

    Directory of Open Access Journals (Sweden)

    Li Ling Lee

    Full Text Available Gammaretroviruses related to murine leukemia virus (MLV have variously been reported to be present or absent in blood from chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME patients and healthy controls. Using subjects from New York State, we have investigated by PCR methods whether MLV-related sequences can be identified in nucleic acids isolated from whole blood or from peripheral blood mononuclear cells (PBMCs or following PBMC culture. We have also passaged the prostate cancer cell line LNCaP following incubation with plasma from patients and controls and assayed nucleic acids for viral sequences. We have used 15 sets of primers that can effectively amplify conserved regions of murine endogenous and exogenous retrovirus sequences. We demonstrate that our PCR assays for MLV-related gag sequences and for mouse DNA contamination are extremely sensitive. While we have identified MLV-like gag sequences following PCR on human DNA preparations, we are unable to conclude that these sequences originated in the blood samples.

  13. Gene expression profiling of human breast tissue samples using SAGE-Seq.

    Science.gov (United States)

    Wu, Zhenhua Jeremy; Meyer, Clifford A; Choudhury, Sibgat; Shipitsin, Michail; Maruyama, Reo; Bessarabova, Marina; Nikolskaya, Tatiana; Sukumar, Saraswati; Schwartzman, Armin; Liu, Jun S; Polyak, Kornelia; Liu, X Shirley

    2010-12-01

    We present a powerful application of ultra high-throughput sequencing, SAGE-Seq, for the accurate quantification of normal and neoplastic mammary epithelial cell transcriptomes. We develop data analysis pipelines that allow the mapping of sense and antisense strands of mitochondrial and RefSeq genes, the normalization between libraries, and the identification of differentially expressed genes. We find that the diversity of cancer transcriptomes is significantly higher than that of normal cells. Our analysis indicates that transcript discovery plateaus at 10 million reads/sample, and suggests a minimum desired sequencing depth around five million reads. Comparison of SAGE-Seq and traditional SAGE on normal and cancerous breast tissues reveals higher sensitivity of SAGE-Seq to detect less-abundant genes, including those encoding for known breast cancer-related transcription factors and G protein-coupled receptors (GPCRs). SAGE-Seq is able to identify genes and pathways abnormally activated in breast cancer that traditional SAGE failed to call. SAGE-Seq is a powerful method for the identification of biomarkers and therapeutic targets in human disease.

  14. Haematospirillum jordaniae gen. nov., sp. nov., isolated from human blood samples.

    Science.gov (United States)

    Humrighouse, B W; Emery, B D; Kelly, A J; Metcalfe, M G; Mbizo, J; McQuiston, J R

    2016-04-01

    A Gram-negative, aerobic, motile, spiral-shaped bacterium, strain H5569(T), was isolated from a human blood sample. Phenotypic and molecular characteristics of the isolate were investigated. Optimal growth was found to occur at 35 °C under aerobic conditions on Heart Infusion Agar supplemented with 5 % rabbit blood. The major fatty acids present in the cells were identified as C16:0, C16:1ω7c and C18:1ω7c. The predominant respiratory quinone was found to be ubiquinone-Q10. The G+C content of genomic DNA for strain H5569(T) was found to be 49.9 %. Based on 16S rRNA gene sequence analysis results, 13 additional isolates were also analysed in this study. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the organism, represented by strain H5569(T), forms a distinct lineage within the family Rhodospirillaceae, closely related to two Novispirillum itersonii subspecies (93.9-94.1 %) and two Caenispirillum sp. (91.2-91.6 %). Based on these results, the isolate H5569(T) is concluded to represent a new genus and species for which the name Haematospirillum jordaniae gen. nov., sp. nov. is proposed. The type strain is H5569(T) (=DSM(T) 28903 = CCUG 66838(T)).

  15. Sources of technical variability in quantitative LC-MS proteomics: human brain tissue sample analysis.

    Science.gov (United States)

    Piehowski, Paul D; Petyuk, Vladislav A; Orton, Daniel J; Xie, Fang; Moore, Ronald J; Ramirez-Restrepo, Manuel; Engel, Anzhelika; Lieberman, Andrew P; Albin, Roger L; Camp, David G; Smith, Richard D; Myers, Amanda J

    2013-05-03

    To design a robust quantitative proteomics study, an understanding of both the inherent heterogeneity of the biological samples being studied as well as the technical variability of the proteomics methods and platform is needed. Additionally, accurately identifying the technical steps associated with the largest variability would provide valuable information for the improvement and design of future processing pipelines. We present an experimental strategy that allows for a detailed examination of the variability of the quantitative LC-MS proteomics measurements. By replicating analyses at different stages of processing, various technical components can be estimated and their individual contribution to technical variability can be dissected. This design can be easily adapted to other quantitative proteomics pipelines. Herein, we applied this methodology to our label-free workflow for the processing of human brain tissue. For this application, the pipeline was divided into four critical components: Tissue dissection and homogenization (extraction), protein denaturation followed by trypsin digestion and SPE cleanup (digestion), short-term run-to-run instrumental response fluctuation (instrumental variance), and long-term drift of the quantitative response of the LC-MS/MS platform over the 2 week period of continuous analysis (instrumental stability). From this analysis, we found the following contributions to variability: extraction (72%) > instrumental variance (16%) > instrumental stability (8.4%) > digestion (3.1%). Furthermore, the stability of the platform and its suitability for discovery proteomics studies is demonstrated.

  16. Sources of Technical Variability in Quantitative LC-MS Proteomics: Human Brain Tissue Sample Analysis.

    Energy Technology Data Exchange (ETDEWEB)

    Piehowski, Paul D.; Petyuk, Vladislav A.; Orton, Daniel J.; Xie, Fang; Moore, Ronald J.; Ramirez Restrepo, Manuel; Engel, Anzhelika; Lieberman, Andrew P.; Albin, Roger L.; Camp, David G.; Smith, Richard D.; Myers, Amanda J.

    2013-05-03

    To design a robust quantitative proteomics study, an understanding of both the inherent heterogeneity of the biological samples being studied as well as the technical variability of the proteomics methods and platform is needed. Additionally, accurately identifying the technical steps associated with the largest variability would provide valuable information for the improvement and design of future processing pipelines. We present an experimental strategy that allows for a detailed examination of the variability of the quantitative LC-MS proteomics measurements. By replicating analyses at different stages of processing, various technical components can be estimated and their individual contribution to technical variability can be dissected. This design can be easily adapted to other quantitative proteomics pipelines. Herein, we applied this methodology to our label-free workflow for the processing of human brain tissue. For this application, the pipeline was divided into four critical components: Tissue dissection and homogenization (extraction), protein denaturation followed by trypsin digestion and SPE clean-up (digestion), short-term run-to-run instrumental response fluctuation (instrumental variance), and long-term drift of the quantitative response of the LC-MS/MS platform over the 2 week period of continuous analysis (instrumental stability). From this analysis, we found the following contributions to variability: extraction (72%) >> instrumental variance (16%) > instrumental stability (8.4%) > digestion (3.1%). Furthermore, the stability of the platform and its’ suitability for discovery proteomics studies is demonstrated.

  17. Association of human papillomavirus and Chlamydia trachomatis with intraepithelial alterations in cervix samples

    Directory of Open Access Journals (Sweden)

    Denise Wohlmeister

    2016-02-01

    Full Text Available The influence of different infectious agents and their association with human papillomavirus (HPV in cervical carcinogenesis have not been completely elucidated. This study describes the association between cytological changes in cervical epithelium and the detection of the most relevant aetiological agents of sexually transmitted diseases. Samples collected from 169 patients were evaluated by conventional cytology followed by molecular analysis to detect HPV DNA, Chlamydia trachomatis, herpes simplex virus 1 and 2,Neisseria gonorrhoeae, Mycoplasma genitalium, Trichomonas vaginalis, andTreponema pallidum, besides genotyping for most common high-risk HPV. An association between cytological lesions and different behavioural habits such as smoking and sedentariness was observed. Intraepithelial lesions were also associated with HPV and C. trachomatis detection. An association was also found between both simple and multiple genotype infection and cytological changes. The investigation of HPV and C. trachomatisproved its importance and may be considered in the future for including in screening programs, since these factors are linked to the early diagnosis of patients with precursor lesions of cervical cancer.

  18. Human sexuality education in the middle grades classroom: A review of curricula in a sample of Florida school districts

    Science.gov (United States)

    Myrick, Melinda D.

    2007-12-01

    This study examined the extent to which human sexuality topics are covered in Florida middle school science classrooms and the process by which curricular decisions are made regarding human sexuality education on a county-wide basis. Primary data included interviews with county-level administrators who oversee curricular decisions related to the middle-grades science curriculum or health curriculum in twelve school districts within the state. These districts represented four geographic locations and districts of various sizes. Administrators from four of the twelve studies in the sample chose to provide information regarding their human sexuality education curriculum. In two cases, teacher leads were identified and were interviewed to understand the implementation of the curriculum within the classroom. Additional data were collected from the district curriculum guides for human sexuality education and the adopted middle-grades science textbook for each county. The interview and documentary data were analyzed by comparison to established criteria for a comprehensive human sexuality education curriculum. The analysis revealed that the scope of human sexuality education varied considerably within the sample and that much of the curricula in place failed to include topics and activities that have been identified as important in a successful human sexuality education program. These findings are limited because few counties chose to fully participate. Additional research is clearly needed to examine the effectiveness of existing human sexuality education curricula in Florida. In addition, research is needed to understand the characteristics, values, and beliefs of successful human sexuality education instructors across the state.

  19. Use of feces to attract insects by a Glittering-bellied Emerald, Chlorostilbon lucidus (Shaw, 1812 (Apodiformes: Trochilidae

    Directory of Open Access Journals (Sweden)

    Fábio André Facco Jacomassa

    2014-09-01

    Full Text Available This study describes the occurrence of a female Glittering-bellied Emerald, Chlorostilbon lucidus, using feces to attract insects to the nesting site for predation. This is the first report of a hummingbird using feces to attract insects.

  20. Accurate measurement of circulating mitochondrial DNA content from human blood samples using real-time quantitative PCR.

    Science.gov (United States)

    Ajaz, Saima; Czajka, Anna; Malik, Afshan

    2015-01-01

    We describe a protocol to accurately measure the amount of human mitochondrial DNA (MtDNA) in peripheral blood samples which can be modified to quantify MtDNA from other body fluids, human cells, and tissues. This protocol is based on the use of real-time quantitative PCR (qPCR) to quantify the amount of MtDNA relative to nuclear DNA (designated the Mt/N ratio). In the last decade, there have been increasing numbers of studies describing altered MtDNA or Mt/N in circulation in common nongenetic diseases where mitochondrial dysfunction may play a role (for review see Malik and Czajka, Mitochondrion 13:481-492, 2013). These studies are distinct from those looking at genetic mitochondrial disease and are attempting to identify acquired changes in circulating MtDNA content as an indicator of mitochondrial function. However, the methodology being used is not always specific and reproducible. As more than 95 % of the human mitochondrial genome is duplicated in the human nuclear genome, it is important to avoid co-amplification of nuclear pseudogenes. Furthermore, template preparation protocols can also affect the results because of the size and structural differences between the mitochondrial and nuclear genomes. Here we describe how to (1) prepare DNA from blood samples; (2) pretreat the DNA to prevent dilution bias; (3) prepare dilution standards for absolute quantification using the unique primers human mitochondrial genome forward primer (hMitoF3) and human mitochondrial genome reverse primer(hMitoR3) for the mitochondrial genome, and human nuclear genome forward primer (hB2MF1) and human nuclear genome reverse primer (hB2MR1) primers for the human nuclear genome; (4) carry out qPCR for either relative or absolute quantification from test samples; (5) analyze qPCR data; and (6) calculate the sample size to adequately power studies. The protocol presented here is suitable for high-throughput use.

  1. A divergent clade of circular single-stranded DNA viruses from pig feces

    Science.gov (United States)

    Using metagenomics and molecular cloning methods, we characterized five novel small circular viral genomes from pig feces distantly related to chimpanzee and porcine stool-associated circular viruses, (ChiSCV and PoSCV1). Phylogenetic analysis placed these viruses into a new, highly divergent, clade...

  2. Pb-Al relationships in waterfowl feces discriminate between sources of Pb exposure

    Energy Technology Data Exchange (ETDEWEB)

    Martinez-Haro, M. [Instituto de Investigacion en Recursos Cinegeticos, IREC (CSIC-UCLM-JCCM), Ronda de Toledo, 13071 Ciudad Real (Spain); Department of Wetland Ecology, Estacion Biologica de Donana - CSIC, Americo Vespucio, 41092 Sevilla (Spain); Taggart, M.A. [Instituto de Investigacion en Recursos Cinegeticos, IREC (CSIC-UCLM-JCCM), Ronda de Toledo, 13071 Ciudad Real (Spain); Mateo, R., E-mail: rafael.mateo@uclm.e [Instituto de Investigacion en Recursos Cinegeticos, IREC (CSIC-UCLM-JCCM), Ronda de Toledo, 13071 Ciudad Real (Spain)

    2010-07-15

    Hunted mallards (n = 56) were collected in the Ebro Delta (Spain) in 2007-08 to evaluate the use of feces as a non-invasive monitoring method to study lead shot ingestion. The prevalence of Pb shot ingestion in these birds was 28.6%, and similar to that reported before a ban on Pb shot use in 2003. Lead concentrations in terminal intestinal contents were higher in mallards with Pb shot in their gizzard. Feces Pb concentrations >34 {mu}g/g d.w were indicative of Pb shot ingestion, and background Pb levels were <12.5 {mu}g/g d.w. To discriminate between birds ingesting soil Pb, and those ingesting Pb shot, we correlated Al and Pb levels and showed that mallards with ingested Pb shot in the gizzard stood out as outliers within the regression. Feces Pb-Al relationships can be used as a simple non-invasive tool in monitoring programs regarding Pb shot ingestion in birds. - By analysing Pb-Al levels in feces, a simple non-invasive method can help determine sources of Pb exposure in waterfowl.

  3. Degradation of Biomacromolecules during High-rate Composting of Wheat Straw-Amended Pig Feces.

    NARCIS (Netherlands)

    Veeken, A.H.M.; Adani, F.; Nierop, K.G.J.; Jager, de P.A.; Hamelers, H.V.M.

    2001-01-01

    Pig (Sus scrofa) feces, separately collected and amended with wheat straw, was composted in a tunnel reactor connected with a cooler. The composting process was monitored for 4 wk and the degradation of organic matter was studied by two chemical extraction methods, 13C cross polarization magic angle

  4. Vesicular-arbuscular mycorrhizae established with Glomus fasciculatus spores isolated from the feces of cricetine mice

    Science.gov (United States)

    Frederick M. Rothwell; Coleman Holt

    1978-01-01

    Cricetine mice were trapped on two revegetated surface-mined areas - one with a freshly seeded grass-legume cover and one with an early successional grass-forb cover. Chlamydospores of Glomus fasciculatus isolated from the feces of these animals produced representative endomycorrhizae with corn under greenhouse conditions.

  5. Specific detection and analysis of a probiotic Bifidobacterium strain in infant feces

    NARCIS (Netherlands)

    Kok, RG; DeWaal, A; Schut, F; Welling, GW; Weenk, G; Hellingwerf, KJ

    1996-01-01

    For specific detection of the probiotic Bifidobacterium sp. strain LW420 in infant feces and for rapid quality control of this strain in culture, three strain-specific 16S rRNA gene-targeted primers have been developed. These primers allow specific detection of the organism via PCR. Specificity of t

  6. Life cycle assessment of segregating fattening pig urine and feces compared to conventional liquid manure management

    NARCIS (Netherlands)

    Vries, de J.W.; Aarnink, A.J.A.; Groot Koerkamp, P.W.G.; Boer, de I.J.M.

    2013-01-01

    Gaseous emissions from in-house storage of liquid animal manure remain a major contributor to the environmental impact of manure management. Our aim was to assess the life cycle environmental consequences and reduction potential of segregating fattening pig urine and feces with an innovative V-belt

  7. Decay of Fecal Indicator Bacteria and Microbial Source Tracking Markers in Cattle Feces

    Science.gov (United States)

    The survival of fecal indicator bacteria (FIB) and microbial source tracking (MST) markers in water microcosms and manure amended soils has been well documented; however, little is known about the survival of MST markers in bovine feces deposited on pastures. We conducted a study...

  8. Congenital Chloride Diarrhea: Diagnosis by Easy-Accessible Chloride Measurement in Feces

    Directory of Open Access Journals (Sweden)

    C. Gils

    2016-01-01

    Full Text Available Background. Congenital chloride diarrhea (CCD is an autosomal recessive disorder caused by mutations in the genes encoding the intestinal Cl−/HCO3- exchanger and is clinically characterized by watery, profound diarrhea, electrolyte disturbances, and metabolic alkalosis. The CCD diagnosis is based on the clinical symptoms and measurement of high chloride concentration in feces (>90 mmol/L and is confirmed by DNA testing. Untreated CCD is lethal, while long-term clinical outcome improves when treated correctly. Case Presentation. A 27-year-old woman had an emergency caesarian due to pain and discomfort in gestational week 36 + 4. The newborn boy had abdominal distension and yellow fluid per rectum. Therapy with intravenous glucose and sodium chloride decreased his stool frequency and improved his clinical condition. A suspicion of congenital chloride diarrhea was strongly supported using blood gas analyzer to measure an increased chloride concentration in the feces; the diagnosis was confirmed by DNA testing. Discussion. Measurement of chloride in feces using an ordinary blood gas analyzer can serve as a preliminary analysis when congenital chloride diarrhea is suspected. This measurement can be easily performed with a watery feces composition. An easy-accessible chloride measurement available will facilitate the diagnostics and support the initial treatment if CCD is suspected.

  9. Interferon-λ4 (IFNL4 transcript expression in human liver tissue samples.

    Directory of Open Access Journals (Sweden)

    Ahmad Amanzada

    Full Text Available Eradication of hepatitis C virus (HCV infection, both spontaneous and treatment-induced, is marked by the wildtype allele C of a single nucleotide polymorphism upstream of the IL28B gene, rs12979860. This favorable allele was recently described to be in linkage disequilibrium with the wildtype allele TT of a dinucleotide polymorphism, ss469415590, located within a new protein-coding gene. While the TT allele introduces a frame-shift and disrupts the open reading frame, only the variant allele, ΔG, creates a novel type III interferon (IFN protein, IFN-λ4/IFNL4. Absence of IFNL4 is thus supposed to favor resolution of HCV infection. As to date IFNL4 mRNA transcription has only been investigated in polyI:C-stimulated primary human hepatocytes and not yet in HCV infection in vivo, this study analyzed IFNL4 mRNA expression in human liver biopsy specimens. Samples were obtained from patients with a broad panel of disorders including no liver disease, liver diseases of non-viral etiology, chronic hepatitis B and chronic hepatitis C. Hepatic IFNL4 transcripts were detectable exclusively in a subgroup of chronic hepatitis C patients (24/45. Their amounts were positively related to liver HCV RNA copy numbers (p = 0.0023, r = 0.56 suggesting that the hepatic viral load influences IFNL4 transcription irrespective of IFNL4 governing genotype. Both, the IFNL4 creating allele ΔG (p<0.0001 and actual IFNL4 transcription (p = 0.0015 were found to be correlated to the activation of IFN stimulatory genes (ISGs. By contrast, IFNL4 ss469415590 genotypes were not found to be related to IFN-λ2/3/IL28 or IFN-λ1/IL29 gene expression. In conclusion, this study is the first report on intrahepatic transcript levels of the recently discovered IFNL4 gene. Data indicate that HCV infection in particular might activate IFNL4 transcription in the liver. It provides a possible explanation as to why hepatitis C patients show ISG stimulation in their livers in the

  10. A study on the levels of a polybrominated biphenyl in Chinese human milk samples collected in 2007 and 2011.

    Science.gov (United States)

    Liu, Xiao; Wen, Sheng; Li, Jingguang; Zhang, Lei; Zhao, Yunfeng; Wu, Yongning

    2016-09-01

    The levels of a 2,2',4,4',5,5'-hexabromobiphenyl (BB-153) were measured in human milk samples collected in 2007 and 2011 from residents in China by high-resolution gas chromatography-high-resolution mass chromatography (HRGC-HRMS) with isotope dilution. The median concentrations of BB-153 from the samples collected in 2007 and 2011 were 8.3 and 7.2 pg/g lipid weight, respectively. The levels of BB-153 in the human milk collected from rural areas were not significantly different to those collected from the urban areas in China. Meanwhile, significant positive correlations were found between the levels of BB-153 in human milk and the consumption of animal-origin foods. In the present study, the mean levels of BB-153 in human milk from Chinese mothers were found to be lower than those from European and American mothers.

  11. Multiple types of human papillomavirus in cervical samples in women in Campo Grande, MS, Brazil

    Directory of Open Access Journals (Sweden)

    Inês Aparecida Tozetti

    2006-10-01

    Full Text Available We analyzed 87 cervical samples from Campo Grande, Mato Grosso do Sul, with a PGMY/GP+ nested PCR system. Positive samples were typed using E7 type-specific primer pairs for HPV 6/11, 16, 18, 45 and 66. Eighteen samples (22% were infected with HPV6/11, 18 samples (22% with HPV66, 13 samples (15.9% with HPV45, 8 samples (9.8% with HPV18 and 7 samples (8.5% with HPV16. Seventeen samples (20.7% were infected by two HPV types, and five samples (6.1% by three HPV types. We conclude that infection with multiple types is present at a high frequency in our population and that there is a relation between some types and cytological finds.

  12. [Species from genus Eimeria observed in domestic rabbit (Oryctolagus cuniculus) feces raised at the Municipality of Campos dos Goytacazes in the State of Rio de Janeiro, Brazil].

    Science.gov (United States)

    de Almeida, Adriana J; Mayen, Friederike L; de Oliveira, Francisco C R

    2006-01-01

    This study was designed to identify species of the genus Eimeria from eleven rabbit meat breeders from Campos dos Goytacazes. Fecal samples were collected from rabbits with different health conditions and consintence of feces. The following species of Eimeria were identified: E. perforans; E. magna; E. coecicola; E. irresidua; E. media; E. flavescens; E. nagpurensis; E. intestinalis, E. exigua and E. stiedae. Parasites from this genus were detected in 81.82% (9) of the rabbit meat production sites, regardless of the management and hygiene conditions. In all cases the infection was always caused from more than one species of Eimeria.

  13. High risk human papillomavirus genotyping in clinical samples: evaluation of different commercial tests.

    Science.gov (United States)

    Paolini, F; Rollo, F; Brandi, R; Benevolo, M; Mariani, L; Cercato, M C; Vocaturo, A; Venuti, A

    2011-01-01

    The aim of the present study is to compare the performance of several commercial human papillomavirus (HPV) tests in a cohort of 281 women. The hybrid capture II, the PreTect-HPV-Proofer, the linear array, and DR.HPVTMIVD were utilized to detect and type HPV in parallel with in-house PCR tests followed by direct automated sequencing or by sub-cloning and sequencing. The concordance levels along with other tests were evaluated with a Cohen's K value varying between 0.60 to 0.88, indicating good correlation with nearly perfect agreement between hybrid capture II, (HCII) and the linear array test. High sensitivity was recorded by the linear array and HCII with 100% (95% CI, 0.8021 to 1.0000) detection of cervical intraepithelial neoplasia (CIN) III by both methods. Conversely, the PreTect-HPV-Proofer showed high specificity with 12% (95% CI, 0.7966 to 0.9163) positivity on normal samples. The genotyping analysis showed that agreement among tests was only low to moderate with great differences between different HPV types. Multiple infections were detected with poor concordance and sub-cloning assays revealed the presence of a lower number of HPV in comparison to the other methods. In summary, the use of different HPV tests applied to the same group of cervical smears may possibly lead to incongruent results, suggesting the need to standardize type-specific sensitivity of genotyping methods and the need to evaluate their accuracy in detecting multiple HPV infections. This would be a prerequisite for the use of genotyping assays in cervical cancer screening programs.

  14. A multiclass method for the analysis of endocrine disrupting chemicals in human urine samples. Sample treatment by dispersive liquid-liquid microextraction.

    Science.gov (United States)

    Vela-Soria, F; Ballesteros, O; Zafra-Gómez, A; Ballesteros, L; Navalón, A

    2014-11-01

    The population is continuously exposed to endocrine disrupting chemicals (EDCs). This has influenced an increase in diseases and syndromes that are more frequent nowadays. Therefore, it is necessary to develop new analytical procedures to evaluate the exposure with the ultimate objective of establishing, in an accurate way, relationships between EDCs and harmful health effects. In the present work, a new method based on a sample treatment by dispersive liquid-liquid microextraction (DLLME) for the extraction of six parabens (methyl-, ethyl-, isopropyl-, propyl-, isobutyl and butylparaben), six benzophenones (benzophenone-1, benzophenone-2, benzophenone-3, benzophenone-6, benzophenone-8 and 4-hydroxybenzophenone) and two bisphenols (bisphenol A and bisphenol S) in human urine samples, followed by gas chromatography-tandem mass spectrometry (GC-MS/MS) analysis is proposed. An enzymatic treatment allows determining the total content of the target EDCs. The extraction parameters were accurately optimized using multivariate optimization strategies. Ethylparaben ring-(13)C6 and bisphenol A-d16 were used as surrogates. Found limits of quantification ranging from 0.2 to 0.5 ng mL(-1) and inter-day variability (evaluated as relative standard deviation) ranging from 2.0% to 14.9%. The method was validated using matrix-matched standard calibration followed by a recovery assay with spiked samples. Recovery rates ranged from 94% to 105%. A good linearity, for concentrations up to 300 ng mL(-1) for parabens and 40 ng mL(-1) for benzophenones and bisphenols, respectively, was obtained. The method was satisfactorily applied for the determination of target compounds in human urine samples from 20 randomly selected individuals.

  15. Chromatofocusing profile of purified human alpha-fetoprotein and albumin differs from those of crude samples: effect of protein concentration of the elution of the sample.

    Science.gov (United States)

    Leal, J A; Eddy, K B; Keel, B A

    1991-02-01

    Chromatofocusing was utilized to characterize charge microheterogeneity of purified human alpha-fetoprotein (AFP) and human serum albumin (HSA). Crude cord blood samples yielded three isoforms: AFP-IA, IB, and II, with pIs 4.57 (52%), 4.27 (43%), and less than 4.00 (5%), respectively. In contrast, 10 micrograms of purified AFP or 250,000 cpm of 125I-AFP eluted entirely as isoform AFP-II. 125I-AFP focused in the presence of crude cord blood, amniotic fluid, adult male serum, or 25 mg purified HSA resulted in elution profiles similar to those of crude cord blood. Pure AFP focused along with 0.1, 1.0, 5.0, or 10 mg HSA showed a gradual shift from AFP-II to AFP-I. With greater than or equal to 5 mg HSA, isoform I was further resolve into AFP-IA and IB. Similarly, 250,000 cpm of 125I-HSA, which also eluted entirely as isoform II, showed a gradual shift to isoform I when increasing concentrations of unlabeled HSA were added. The resolution of isoform HSA-I in HSA-IA, IB, and IC was again improved with greater than or equal to 5 mg unlabeled HSA. When carrier proteins of varying pI values were chromatofocused along with purified AFP, it was observed that only those proteins with pIs in the range of AFP caused significant alteration in the relative distribution of AFP. We conclude that sample protein concentration and composition must be carefully considered when chromatofocusing is being used for purified samples and when the elution profiles of samples from different origins and varying protein concentrations are being compared.

  16. 依赖粪便材料的大熊猫肠道耶尔森氏菌的检测%Feces-based Detection of Yersinia enterocolitica in Intestine of Ailuropoda melanoleuca by PCR

    Institute of Scientific and Technical Information of China (English)

    杨水云; 裴渭静; 孙飞龙; 陈曦; 吴明宇; 石晓强; 吴晓民; 任建设; 贾康胜

    2004-01-01

    It is inevitable to develop noninvasive sampling methods to do studies on giant panda even diagnose the diseases since which is so endangered that it's impossible to carry out invasive sampling. A non-invasive sampling method to detect the intestinal pathogen, Yersirda enterocolitica in feces of pandas based designing PCR primers was established in this study. The main procedures are based on bacteria enrichment and cell lysis before binding the pathogen DNA to silica powder at high concentration of Kalium iodide and neutral pH conditions. Before PCR cycles, the binded DNA is washed with 80% ethanol and eluted with diluted EDTA buffer. Restdts showed that the silica-based feces DNA-purification method could remove the inhibitors of PCR so applicable to detect the target pathogen.

  17. Could the domestic cat play a significant role in the transmission of Echinococcus multilocularis? A study based on qPCR analysis of cat feces in a rural area in France

    Directory of Open Access Journals (Sweden)

    Knapp Jenny

    2016-01-01

    Full Text Available Echinococcus multilocularis, a cestode parasite responsible for alveolar echinococcosis in humans, is often reported in Europe. It involves red foxes, domestic dogs, and domestic and wild cats as definitive hosts. The parasite infects small mammals and accidentally humans as intermediate hosts and develops in a similar way to a tumor, usually in the liver. Domestic animals are suspected of playing a role in parasite transmission, but this is rarely proven. Moreover, the role of domestic cats is thought to be small, because of experimental studies showing incomplete development of the parasite observed in their intestines. In the present study, we investigated copro-sampling performed in a rural and highly endemic area in Eastern France, on carnivore feces (n = 150. From these samples, the parasite was detected and identified by DNA analysis using quantitative PCR targeting part of a mitochondrial gene (Em-qPCR. Taeniid eggs were isolated from positive-Em-qPCR samples by flotation, and species identification was confirmed by sequencing on DNA extracts. From a total of 43 copro-samples from cats, four tested positive for E. multilocularis by the Em-qPCR. In two of these, we found parasite eggs that were identified as E. multilocularis. This finding was confirmed by sequencing, while one dog stool out of 61 collected was found to be positive, no egg was detectable. At the same time, 34% of fox stools tested positive for the parasite. The present study challenges the current idea that cats are only of minor significance in the E. multilocularis life cycle.

  18. pH adjustment of human blood plasma prior to bioanalytical sample preparation

    NARCIS (Netherlands)

    Hendriks, G.; Uges, D. R. A.; Franke, J. P.

    2008-01-01

    pH adjustment in bioanalytical sample preparation concerning ionisable compounds is one of the most common sample treatments. This is often done by mixing an aliquot of the sample with a proper buffer adjusted to the proposed pH. The pH of the resulting mixture however, does not necessarily have to

  19. Novel method for simultaneous aqueous in situ derivatization of THC and THC-COOH in human urine samples: validation and application to real samples.

    Science.gov (United States)

    Chericoni, S; Battistini, I; Dugheri, S; Pacenti, M; Giusiani, M

    2011-05-01

    The present work describes the validation of a novel aqueous in situ derivatization procedure with trimethyloxonium tetrafluoroborate (TMO) as methylating agent for the simultaneous, quantitative analysis of Δ(9)-tetrahydrocannabinol (THC) and 11-nor-Δ(9)-tetrahydrocannabinol carboxylic acid (THC-COOH) in human urine. The derivatizing agent is directly added to the urine sample and the methyl-derivatives are then recovered by liquid-liquid extraction procedure. Gas chromatography-mass spectrometry was used to detect the derivatives in selected ion monitoring mode. The limits of detection were 0.7 ng/mL for THC and 0.5 ng/mL for THC-COOH, whereas the limits of quantification were 1.9 and 0.9 ng/mL, respectively. The method has been applied to 60 real samples both positive and negative to immunochemical screening test resulting to be very useful and reliable in routine analysis of THC-COOH in human urine for toxicological and forensic purposes.

  20. Comparison of different sampling types across the rearing period in broiler flocks for isolation of Campylobacter spp.

    Science.gov (United States)

    Ingresa-Capaccioni, S; González-Bodí, S; Jiménez-Trigos, E; Marco-Jiménez, F; Catalá, P; Vega, S; Marin, C

    2015-04-01

    Campylobacter is the most common bacterial cause of human gastrointestinal disease in most developed countries. It is generally accepted that poultry products are a significant source of foodborne Campylobacter infections in humans. Assessing the effectiveness of any potential intervention at farm level requires monitoring of the Campylobacter status of broiler flocks, using appropriate sampling methods. The aim of this study was to assess the influence of the sample type across the rearing period for the detection of Campylobacter spp. at farm level. During this study, 21 commercial broiler farms were intensively sampled. Each farm was visited and sampled at different times during the rearing period (d 1, 7, 14, 21, 28, 35, and 42). On the first day of rearing, the status of the house and the day-old flock was evaluated, and environmental and cecal samples were collected. During rearing, 4 different sample types were collected: feces with sock swabs (sock swabs), feces directly from the litter (feces), cloacal swabs, and cecal content. All samples were analyzed according to ISO 10272-1:2006 (Annex E) and also by direct culture. The results of this study showed that Campylobacter spp. were detected in all of the sample types on d 14 of rearing. From this point on, the detection increased significantly, with a maximum detection rate by the end of rearing, regardless of the sample type. All samples that were negative upon direct culture were also negative after pre-enrichment. At the end of rearing, the percentage of samples positive for Campylobacter spp. was 71.4% for cecal samples, 61.9% for cloacal swabs, 45.2% for sock swabs, and 69.1% for fecal samples. C. jejuni was detected in all the sample types, with positive rates ranging from 67.1 to 76.0% for cecal samples and cloacal content, respectively. Cecal samples, cloacal swabs, and fecal samples cultured by direct plating onto modified charcoal cefoperazone deoxycholate agar (mCCDA) without pre-enrichment have

  1. Combining near infrared spectra of feces and geostatistics to generate forage nutritional quality maps across landscapes.

    Science.gov (United States)

    Pierre-Olivier, Jean; Bradley, Robert L; Tremblay, Jean-Pierre; Côté, Steeve D

    2015-09-01

    An important asset for the management of wild ungulates is recognizing the spatial distribution of forage quality across heterogeneous landscapes. To do so typically requires knowledge of which plant species are eaten, in what abundance they are eaten, and what their nutritional quality might be. Acquiring such data, however, may be difficult and time consuming. Here, we are proposing a rapid and cost-effective forage quality monitoring tool that combines near infrared (NIR) spectra of fecal samples and easily obtained data on plant community composition. Our approach rests on the premise that NIR spectra of fecal samples collected within low population density exclosures reflect the optimal forage quality of a given landscape. Forage quality can thus be based on the Mahalanobis distance of fecal spectral scans across the landscape relative to fecal spectral scans inside exclosures (referred to as DISTEX). The Gi* spatial autocorrelation statistic can then be applied among neighboring DISTEX values to detect and map "hot spots" and "cold spots" of nutritional quality over the landscape. We tested our approach in a heterogeneous boreal landscape on Anticosti Island (Québec, Canada), where white-tailed deer (Odocoileus virginianus) populations over the landscape have ranged from 20 to 50 individuals/km2 for at least 80 years, resulting in a loss of most palatable and nutritious plant species. Our results suggest that hot spots of forage quality occur when old-growth balsam fir stands comprise >39.8% of 300 ha neighborhoods, whereas cold spots occur in laggs (i.e., transition zones from forest to peatland). In terms of ground-level indicator plant species, the presence of Canada bunchberry (Cornus canadensis) was highly correlated with hot spots, whereas tamarack (Larix laricina) was highly correlated with cold spots. Mean DISTEX values were positively and significantly correlated with the neutral detergent fiber and acid detergent lignin contents of feces. While our

  2. An Investigation of Cellulose Digesting Bacteria in the Camel Feces Microbiome

    Science.gov (United States)

    Man, V.; Leung, F. C.

    2015-12-01

    Research Question: Is there a bacteria in camel feces that digests cellulose material and can be used for waste to energy projects? Fossil fuels are the current main resource of energy in the modern world. However, as the demand for fuel increases, biofuels have been proposed as an alternative energy source that is a more sustainable form of liquid fuel generation from living things or waste, commonly known as biofuels and ethanol. The Camelus dromedarius', also known as Arabian camel, diet consist of grass, grains, wheat and oats as well desert vegetation in their natural habitat. However, as the Arabian camel lacks the enzymes to degrade cellulose, it is hypothesized that cellulose digestion is performed by microbial symbionts in camel microbiota. Fecal samples were collected from the Camelus dromedarius in United Arab Emirates and diluted 10-7 times. The diluted sample was then streaked onto a Sodium Carboxymethyl Cellulose plate, and inoculated onto CMC and Azure-B plates. Afterwards, Congo Red was used for staining in order to identify clearance zones of single colonies that may potentially be used as a qualitative assays for cellulose digestion. Then the colonies undergo polymerase chain reaction amplification to produce amplified RNA fragments. The 16S ribosomal RNA gene is identified based on BLAST result using Sanger Sequencing. Amongst the three identified microbes: Bacillus, Staphylococcus and Escherichia coli, both Bacillus and Staphylococcus are cellulose-digesting microbes, and through the fermentation of lignocellulosic, biomasses can be converted into cellulosic ethanol (Biofuel). According to the Improvements in Life Cycle Energy Efficiency and Greenhouse Gas Emissions of Corn-Ethanol by Adam J. Liska, ""Ethanol reduces greenhouse gas emissions by 40-50% when compared directly to gasoline." The determination of bacterial communities that are capable of efficiently and effectively digesting cellulose materials requires that the bacteria be first

  3. Sample size calculations in human electrophysiology (EEG and ERP) studies: A systematic review and recommendations for increased rigor.

    Science.gov (United States)

    Larson, Michael J; Carbine, Kaylie A

    2017-01-01

    There is increasing focus across scientific fields on adequate sample sizes to ensure non-biased and reproducible effects. Very few studies, however, report sample size calculations or even the information needed to accurately calculate sample sizes for grants and future research. We systematically reviewed 100 randomly selected clinical human electrophysiology studies from six high impact journals that frequently publish electroencephalography (EEG) and event-related potential (ERP) research to determine the proportion of studies that reported sample size calculations, as well as the proportion of studies reporting the necessary components to complete such calculations. Studies were coded by the two authors blinded to the other's results. Inter-rater reliability was 100% for the sample size calculations and kappa above 0.82 for all other variables. Zero of the 100 studies (0%) reported sample size calculations. 77% utilized repeated-measures designs, yet zero studies (0%) reported the necessary variances and correlations among repeated measures to accurately calculate future sample sizes. Most studies (93%) reported study statistical values (e.g., F or t values). Only 40% reported effect sizes, 56% reported mean values, and 47% reported indices of variance (e.g., standard deviations/standard errors). Absence of such information hinders accurate determination of sample sizes for study design, grant applications, and meta-analyses of research and whether studies were adequately powered to detect effects of interest. Increased focus on sample size calculations, utilization of registered reports, and presenting information detailing sample size calculations and statistics for future researchers are needed and will increase sample size-related scientific rigor in human electrophysiology research.

  4. Avian Influenza A Virus Infections in Humans

    Science.gov (United States)

    ... their saliva, mucous and feces. Human infections with bird flu viruses can happen when enough virus gets into ... Virus (CVV) for a Highly Pathogenic Avian Influenza (Bird Flu) Virus ” for more information on this process. ...

  5. Comparing the proteome of snap frozen, RNAlater preserved, and formalin-fixed paraffin-embedded human tissue samples

    DEFF Research Database (Denmark)

    Bennike, Tue Bjerg; Kastaniegaard, Kenneth; Padurariu, Simona

    2016-01-01

    Large biobanks exist worldwide containing formalin-fixed, paraffin-embedded samples and samples stored in RNAlater. However, the impact of tissue preservation on the result of a quantative proteome analysis remains poorly described. Human colon mucosal biopsies were extracted from the sigmoideum...... and either immediately frozen, stabilized in RNAlater, or stabilized by formalin-fixation. In one set of biopsies, formalin stabilization was delayed for 30 min. The protein content of the samples was characterized by high throughput quantitative proteomics. We were able to identify a similar high number...

  6. Self-Sampling for Human Papillomavirus Testing among Non-Attenders Increases Attendance to the Norwegian Cervical Cancer Screening Programme

    DEFF Research Database (Denmark)

    Enerly, Espen; Bonde, Jesper; Schee, Kristina;

    2016-01-01

    Increasing attendance to screening offers the best potential for improving the effectiveness of well-established cervical cancer screening programs. Self-sampling at home for human papillomavirus (HPV) testing as an alternative to a clinical sampling can be a useful policy to increase attendance....... To determine whether self-sampling improves screening attendance for women who do not regularly attend the Norwegian Cervical Cancer Screening Programme (NCCSP), 800 women aged 25-69 years in the Oslo area who were due to receive a 2nd reminder to attend regular screening were randomly selected and invited...... alternative for increasing cervical cancer screening coverage in Norway....

  7. Genotypic analyses of shiga toxin-producing Escherichia coli O157 and non-O157 recovered from feces of domestic animals on rural farms in Mexico.

    Science.gov (United States)

    Amézquita-López, Bianca A; Quiñones, Beatriz; Cooley, Michael B; León-Félix, Josefina; Castro-del Campo, Nohelia; Mandrell, Robert E; Jiménez, Maribel; Chaidez, Cristóbal

    2012-01-01

    Shiga toxin-producing Escherichia coli (STEC) are zoonotic enteric pathogens associated with human gastroenteritis worldwide. Cattle and small ruminants are important animal reservoirs of STEC. The present study investigated animal reservoirs for STEC in small rural farms in the Culiacan Valley, an important agricultural region located in Northwest Mexico. A total of 240 fecal samples from domestic animals were collected from five sampling sites in the Culiacan Valley and were subjected to an enrichment protocol followed by either direct plating or immunomagnetic separation before plating on selective media. Serotype O157:H7 isolates with the virulence genes stx2, eae, and ehxA were identified in 40% (26/65) of the recovered isolates from cattle, sheep and chicken feces. Pulse-field gel electrophoresis (PFGE) analysis grouped most O157:H7 isolates into two clusters with 98.6% homology. The use of multiple-locus variable-number tandem repeat analysis (MLVA) differentiated isolates that were indistinguishable by PFGE. Analysis of the allelic diversity of MLVA loci suggested that the O157:H7 isolates from this region were highly related. In contrast to O157:H7 isolates, a greater genotypic diversity was observed in the non-O157 isolates, resulting in 23 PFGE types and 14 MLVA types. The relevant non-O157 serotypes O8:H19, O75:H8, O111:H8 and O146:H21 represented 35.4% (23/65) of the recovered isolates. In particular, 18.5% (12/65) of all the isolates were serotype O75:H8, which was the most variable serotype by both PFGE and MLVA. The non-O157 isolates were predominantly recovered from sheep and were identified to harbor either one or two stx genes. Most non-O157 isolates were ehxA-positive (86.5%, 32/37) but only 10.8% (4/37) harbored eae. These findings indicate that zoonotic STEC with genotypes associated with human illness are present in animals on small farms within rural communities in the Culiacan Valley and emphasize the need for the development of control

  8. A universal genome sequencing method for rotavirus A from human fecal samples which identifies segment reassortment and multi-genotype mixed infection

    National Research Council Canada - National Science Library

    Tran Thi Ngoc Dung; Pham Thanh Duy; October M Sessions; Uma K Sangumathi; Voong Vinh Phat; Pham Thi Thanh Tam; Nguyen Thi Nguyen To; Tran My Phuc; Tran Thi Hong Chau; Nguyen Ngoc Minh Chau; Ngoc Nguyen Minh; Guy E Thwaites; Maia A Rabaa; Stephen Baker

    2017-01-01

    Background Genomic characterization of rotavirus (RoV) has not been adopted at large-scale due to the complexity of obtaining sequences for all 11 segments, particularly when feces are used as starting material...

  9. The determination of budesonide and fluticasone in human sputum samples collected from COPD patients using LC-MS/MS

    NARCIS (Netherlands)

    Buscher, B.A.P.; Jägfeldt, H.; Sandman, H.; Brust-van Schaik, R.; Schaik, F. van; Brüll, L.P.

    2012-01-01

    A bioanalytical method for the quantitative determination of budesonide and fluticasone in human sputum was developed. Sputolysin ® Reagent was added to the sputum samples. After incubation (37°C; 60-70min under shaking) and automated solid phase extraction the extracts were analysed using LC-MS/MS.

  10. Selenium speciation in human urine samples by LC- and CE-ICP-MS-separation and identification of selenosugars

    DEFF Research Database (Denmark)

    Gammelgaard, Bente; Bendahl, L.

    2004-01-01

    Human urine samples were analysed by a reversed-phase chromatographic system and an ion-pair chromatographic system. The chromatographic system, was connected to the ICP-MS either by a microconcentric nebulizer (MCN) in combination with a cyclonic spraychamber or by a modified direct injection...

  11. Human Papillomavirus (HPV) Risk Factors, Vaccination Patterns, and Vaccine Perceptions among a Sample of Male College Students

    Science.gov (United States)

    Fontenot, Holly B.; Collins Fantasia, Heidi; Charyk, Anna; Sutherland, Melissa A.

    2014-01-01

    Objective: To examine human papillomavirus (HPV) vaccination rates, including initiation and completion of the vaccine series, and barriers to vaccination in a sample of male college students. Participants: Male students between the ages of 18 and 25 who reported being currently or previously sexually active (N = 735). Methods: A cross-sectional…

  12. Relationship of Bender Gestalt Developmental Scores and Human Drawing Developmental Scores in a Sample of Turkish Preschool Children

    Science.gov (United States)

    Ozer, Serap

    2009-01-01

    The Bender Gestalt test and Human Drawings are frequently utilized tests in assessing school readiness in children. This study was a pilot attempt to evaluate these two tests in a Turkish sample as they relate to first grade behaviour as measured by teacher ratings. One hundred and five children were evaluated at the end of kindergarten using the…

  13. Simultaneous kinetic spectrophotometric determination of norfloxacin and rifampicin in pharmaceutical formulation and human urine samples by use of chemometrics approaches

    Institute of Scientific and Technical Information of China (English)

    KOKOT; Serge

    2008-01-01

    A kinetic spectrophotometric method with aid of chemometrics is proposed for the simultaneous determination of norfloxacin and rifampicin in mixtures. The proposed method was applied for the simultaneous determination of these two compounds in pharmaceutical formulation and human urine samples,and the results obtained are similar to those obtained by high performance liquid chromatography.

  14. Hybrid Capture 2 and cobas human papillomavirus assays perform similarly on SurePath samples from women with abnormalities

    DEFF Research Database (Denmark)

    Fornari, D; Rebolj, M; Bjerregaard, B

    2016-01-01

    OBJECTIVE: In two laboratories (Departments of Pathology, Copenhagen University Hospitals of Herlev and Hvidovre), we compared cobas and Hybrid Capture 2 (HC2) human papillomavirus (HPV) assays using SurePath® samples from women with atypical squamous cells of undetermined significance (ASCUS) at...

  15. Relationship of Bender Gestalt Developmental Scores and Human Drawing Developmental Scores in a Sample of Turkish Preschool Children

    Science.gov (United States)

    Ozer, Serap

    2009-01-01

    The Bender Gestalt test and Human Drawings are frequently utilized tests in assessing school readiness in children. This study was a pilot attempt to evaluate these two tests in a Turkish sample as they relate to first grade behaviour as measured by teacher ratings. One hundred and five children were evaluated at the end of kindergarten using the…

  16. Role of the furrow of the proximal colon in the production of soft and hard feces in nutrias, Myocastor coypus.

    Science.gov (United States)

    Takahashi, T; Sakaguchi, E

    2000-11-01

    The bacterial level of soft feces is higher than that of hard feces in nutrias. This suggests the heterogeneity of bacterial density in the large intestine. To show the heterogeneity of bacteria in the contents of the large intestine in nutrias, we divided the contents of the large intestine into 12 regions, then measured the nitrogen (N), total amino acids (TAA) and diaminopimelic acid (DAP), a bacterial marker, of these regions. Levels of N, TAA and DAP varied along the cross section of the proximal colon. The greater curvature of the main lumen and furrow had higher N, TAA and DAP concentrations than the lesser curvature. We also examined the involvement of the furrow in producing two types of feces differing in bacterial nitrogen content by surgically preventing the flow of the furrow contents. We compared the concentrations of N, TAA and DAP between soft and hard feces among operated, sham-operated and intact animals. Surgical closure of the furrow abolished the difference in levels of N, TAA and DAP between soft and hard feces, suggesting that the furrow of the proximal colon is responsible for making the bacterial density higher in soft feces than in hard feces.

  17. Evaluation of the detection of Mycobacterium tuberculosis with metabolic activity in culture-negative human clinical samples.

    Science.gov (United States)

    Cubero, N; Esteban, J; Palenque, E; Rosell, A; Garcia, M J

    2013-03-01

    Mycobacterium tuberculosis is assumed to remain in a quiescent state during latent infection, being unable to grow in culture. The aim of this study was to evaluate the detection of viable but non-cultivable bacilli with metabolic activity in human clinical samples using a procedure that is independent of the immunological status of the patient. The study was performed on 66 human clinical samples, from patients subjected to routine diagnosis to rule out a mycobacterial infection. Specimens from pulmonary and extra-pulmonary origins were verified to contain human DNA before testing for M. tuberculosis DNA, rRNA and transient RNA by real-time quantitative PCR. Clinical records of 55 patients were also reviewed. We were able to detect viable but non-cultivable bacilli with a metabolic activity in both pulmonary and extra-pulmonary samples. Mycobacterium tuberculosis RNA was detected in the majority of culture-positive samples whereas it was detected in one-third of culture-negative samples, 20% of them showed metabolic activity. Amplifications of the ftsZ gene and particularly of the main promoter of the ribosomal operon rrnA, namely PCL1, seem to be good targets to detect active bacilli putatively involved in latent infection. Moreover, this last target would provide information on the basal metabolic activity of the bacilli detected. © 2012 The Authors. Clinical Microbiology and Infection © 2012 European Society of Clinical Microbiology and Infectious Diseases.

  18. Effect of Dietary Polydextrose on Feces Consistency and Macronutrient Digestibility in Healthy Dogs

    Directory of Open Access Journals (Sweden)

    K. Vasupen

    2011-01-01

    Full Text Available Problem statement: There is evidence that the addition of 3% STA-LITE® polydextrose to a dry food reduces the clinical signs of osteoarthritis in dogs. For the application of polydextrose as functional ingredient of dog foods, information as to its impact on the acceptance of food, feces consistency, fecal odor and digestibility of macronutrients and minerals is required. Approach: A feeding experiment with 4×4 Latin square design was carried out with 12 adult, healthy Golden Retrievers. The experimental diets consisted of a commercial dry food to which either 0, 2, 4 or 8% polydextrose was added. During the last 5 days of each two-week feeding period, feces of individual dogs were collected quantitatively. Feces consistency and fecal odor were scored. Fecal pH, fecal Lactobacilli and the apparent digestibility of macronutrients and minerals were determined. Results: The various inclusion levels of polydextrose did not affect food acceptance and intake and did not induce changes in body weight. Polydextrose intake did not influence fecal odor and the amount of feces production. Feces consistency was significantly affected by polydextrose; feces became softer with increasing dose. The apparent digestibilities of dietary dry matter, crude protein, crude fat and ash were not significantly affected by polydextrose intake. The apparent digestibilities of crude fiber and nitrogen-free extract were increased by the addition of polydextrose to the diet. Apparent digestibility of polydextrose was calculated to be 92% of intake. The high apparent digestibility of polydextrose points at fermentation of this functional ingredient in the hindgut of dogs, but it was not associated with a further lowering of fecal pH and increase in fecal Lactobacilli. Apparent calcium and phosphorus absorption were not significantly affected by polydextrose intake. Conclusion: In the clinical trial on canine osteoarthritis, a polydextrose dose of 3% in a dry dog food was

  19. A simple method of genomic DNA extraction from human samples for PCR-RFLP analysis.

    Science.gov (United States)

    Ghatak, Souvik; Muthukumaran, Rajendra Bose; Nachimuthu, Senthil Kumar

    2013-12-01

    Isolation of DNA from blood and buccal swabs in adequate quantities is an integral part of forensic research and analysis. The present study was performed to determine the quality and the quantity of DNA extracted from four commonly available samples and to estimate the time duration of the ensuing PCR amplification. Here, we demonstrate that hair and urine samples can also become an alternate source for reliably obtaining a small quantity of PCR-ready DNA. We developed a rapid, cost-effective, and noninvasive method of sample collection and simple DNA extraction from buccal swabs, urine, and hair using the phenol-chloroform method. Buccal samples were subjected to DNA extraction, immediately or after refrigeration (4-6°C) for 3 days. The purity and the concentration of the extracted DNA were determined spectrophotometerically, and the adequacy of DNA extracts for the PCR-based assay was assessed by amplifying a 1030-bp region of the mitochondrial D-loop. Although DNA from all the samples was suitable for PCR, the blood and hair samples provided a good quality DNA for restriction analysis of the PCR product compared with the buccal swab and urine samples. In the present study, hair samples proved to be a good source of genomic DNA for PCR-based methods. Hence, DNA of hair samples can also be used for the genomic disorder analysis in addition to the forensic analysis as a result of the ease of sample collection in a noninvasive manner, lower sample volume requirements, and good storage capability.

  20. Extraction and sensitive detection of toxins A and B from the human pathogen Clostridium difficile in 40 seconds using microwave-accelerated metal-enhanced fluorescence.

    Directory of Open Access Journals (Sweden)

    Lovleen Tina Joshi

    Full Text Available Clostridium difficile is the primary cause of antibiotic associated diarrhea in humans and is a significant cause of morbidity and mortality. Thus the rapid and accurate identification of this pathogen in clinical samples, such as feces, is a key step in reducing the devastating impact of this disease. The bacterium produces two toxins, A and B, which are thought to be responsible for the majority of the pathology associated with the disease, although the relative contribution of each is currently a subject of debate. For this reason we have developed a rapid detection assay based on microwave-accelerated metal-enhanced fluorescence which is capable of detecting the presence of 10 bacteria in unprocessed human feces within 40 seconds. These promising results suggest that this prototype biosensor has the potential to be developed into a rapid, point of care, real time diagnostic assay for C. difficile.

  1. Use of real-time PCR to detect canine parvovirus in feces of free-ranging wolves

    Science.gov (United States)

    Mech, L. David; Almberg, Emily S.; Smith, Douglas; Goyal, Sagar; Singer, Randall S.

    2012-01-01

    Using real-time PCR, we tested 15 wolf (Canis lupus) feces from the Superior National Forest (SNF), Minnesota, USA, and 191 from Yellowstone National Park (YNP), USA, collected during summer and 13 during winter for canine parvovirus (CPV)-2 DNA. We also tested 20 dog feces for CPV-2 DNA. The PCR assay was 100% sensitive and specific with a minimum detection threshold of 104 50% tissue culture infective dose. Virus was detected in two winter specimens but none of the summer specimens. We suggest applying the technique more broadly especially with winter feces.

  2. Detection of Alphacoronavirus vRNA in the Feces of Brazilian Free-Tailed Bats (Tadarida brasiliensis from a Colony in Florida, USA

    Directory of Open Access Journals (Sweden)

    Tania S. Bonny

    2017-02-01

    Full Text Available Bats are natural reservoirs of coronaviruses and other viruses with zoonotic potential. Florida has indigenous non-migratory populations of Brazilian free-tailed bats (Tadarida brasiliensis that mostly roost in colonies in artificial structures. Unlike their counterparts in Brazil and Mexico, the viruses harbored by the Florida bats have been underexplored. We report the detection of an alphacoronavirus RNA-dependent RNA polymerase (RdRp gene sequence in the feces of two of 19 different T. brasiliensis that were capture/release bats that had been evaluated for overall health. The RdRp sequence is similar but not identical to previously detected sequences in the feces of two different species of bats (T. brasiliensis and Molossus molossus in Brazil. In common with the experience of others doing similar work, attempts to isolate the virus in cell cultures were unsuccessful. We surmise that this and highly related alphacoronavirus are carried by Brazilian free-tailed bats living in a wide eco-spatial region. As various coronaviruses (CoVs that affect humans emerged from bats, our study raises the question whether CoVs such as the one detected in our work are yet-to-be-detected pathogens of humans and animals other than bats.

  3. The evaluation of human hand odor volatiles on various textiles: a comparison between contact and noncontact sampling methods.

    Science.gov (United States)

    Prada, Paola A; Curran, Allison M; Furton, Kenneth G

    2011-07-01

    The focus of this study is to compare contact and noncontact human scent collection procedures across an array of textiles (cotton, rayon, polyester, and wool) to determine an optimized collection method for human scent evidence. Six subjects were sampled in triplicate for each textile and collection mode, and the samples were then analyzed through headspace solid-phase micro-extraction in combination with gas chromatography/mass spectrometry (SPME-GC/MS). Contact sampling with cotton material has been shown to be the collection method that yielded the greatest number of volatile compounds and the highest scent mass amounts. Through Spearman rank correlations, it was shown that an individual's scent profile is more reproducible within samples collected on the same textile type than between different materials. Furthermore, contact sampling with cotton fabric demonstrated the greatest reproducibility producing the lowest amount of type I and type II errors with 90.85% of the samples distinguished at the 0.9 match/no match threshold.

  4. Occurrence of aflatoxin M1 in human milk samples in Vojvodina, Serbia: Estimation of average daily intake by babies.

    Science.gov (United States)

    Radonić, Jelena R; Kocić Tanackov, Sunčica D; Mihajlović, Ivana J; Grujić, Zorica S; Vojinović Miloradov, Mirjana B; Škrinjar, Marija M; Turk Sekulić, Maja M

    2017-01-02

    The objectives of the study were to determine the aflatoxin M1 content in human milk samples in Vojvodina, Serbia, and to assess the risk of infants' exposure to aflatoxins food contamination. The growth of Aspergillus flavus and production of aflatoxin B1 in corn samples resulted in higher concentrations of AFM1 in milk and dairy products in 2013, indicating higher concentrations of AFM1 in human milk samples in 2013 and 2014 in Serbia. A total number of 60 samples of human milk (colostrum and breast milk collected 4-8 months after delivery) were analyzed for the presence of AFM1 using the Enzyme Linked Immunosorbent Assay method. The estimated daily intake of AFM1 through breastfeeding was calculated for the colostrum samples using an average intake of 60 mL/kg body weight (b.w.)/day on the third day of lactation. All breast milk collected 4-8 months after delivery and 36.4% of colostrum samples were contaminated with AFM1. The greatest percentage of contaminated colostrum (85%) and all samples of breast milk collected 4-8 months after delivery had AFM1 concentration above maximum allowable concentration according to the Regulation on health safety of dietetic products. The mean daily intake of AFM1 in colostrum was 2.65 ng/kg bw/day. Results of our study indicate the high risk of infants' exposure, who are at the early stage of development and vulnerable to toxic contaminants.

  5. Androgenic biomarker prof|ling in human matrices and cell culture samples using high throughput, electrospray tandem mass spectrometry.

    Science.gov (United States)

    Wilton, John H; Titus, Mark A; Efstathiou, Eleni; Fetterly, Gerald J; Mohler, James L

    2014-05-01

    BACKGROUND. A high throughput, high pressure liquid chromatographic (HPLC) method with triple quadrupole mass spectral detection (LC/MS/MS) was validated for the measurement of 5 endogenous androgens in human plasma and serum and applied to various in vivo and in vitro study samples to pursue a better understanding of the interrelationship of the androgen axis, intracrine metabolism, and castration-recurrent prostate cancer (CaP). A Shimadzu HPLC system interfaced with a Sciex QTRAP 5500 mass spectrometer with electrospray ionization was used with in line column-switching. Samples were liquid/liquid extracted and chromatographed on a Luna C18(2) column at 60°C with a biphasic gradient using a 15-min run time. The method was validated for five androgens in human plasma and serum, and applied to four sets of samples. Plasma (n=188) and bone marrow aspirate (n=129) samples from patients with CaP, who received abiraterone acetate plus prednisone for up to 945 days(135 weeks), had undetectable androgens after 8 weeks of treatment. Plasma dehydroepiandrosterone(DHEA) concentrations were higher in African Americans than Caucasian Americans with newly diagnosed CaP. Analysis of prostate tumor tissue homogenates demonstrated reproducible testosterone (T) and dihydrotestosterone (DHT) concentrations with a minimal sample size of 1.0–2.0 mg of tissue. Finally, cell pellet and media samples from the LNCaP C4-2 cell line showed conversion of T to DHT. The proposed LC/MS/MS method was validated for quantitation of five endogenous androgens in human plasma and serum, and effectively profiles androgens in clinical specimens and cell culture samples.

  6. Women's experience with home-based self-sampling for human papillomavirus testing.

    Science.gov (United States)

    Sultana, Farhana; Mullins, Robyn; English, Dallas R; Simpson, Julie A; Drennan, Kelly T; Heley, Stella; Wrede, C David; Brotherton, Julia M L; Saville, Marion; Gertig, Dorota M

    2015-11-04

    Increasing cervical screening coverage by reaching inadequately screened groups is essential for improving the effectiveness of cervical screening programs. Offering HPV self-sampling to women who are never or under-screened can improve screening participation, however participation varies widely between settings. Information on women's experience with self-sampling and preferences for future self-sampling screening is essential for programs to optimize participation. The survey was conducted as part of a larger trial ("iPap") investigating the effect of HPV self-sampling on participation of never and under-screened women in Victoria, Australia. Questionnaires were mailed to a) most women who participated in the self-sampling to document their experience with and preference for self-sampling in future, and b) a sample of the women who did not participate asking reasons for non-participation and suggestions for enabling participation. Reasons for not having a previous Pap test were also explored. About half the women who collected a self sample for the iPap trial returned the subsequent questionnaire (746/1521). Common reasons for not having cervical screening were that having Pap test performed by a doctor was embarrassing (18 %), not having the time (14 %), or that a Pap test was painful and uncomfortable (11 %). Most (94 %) found the home-based self-sampling less embarrassing, less uncomfortable (90 %) and more convenient (98%) compared with their last Pap test experience (if they had one); however, many were unsure about the test accuracy (57 %). Women who self-sampled thought the instructions were clear (98 %), it was easy to use the swab (95 %), and were generally confident that they did the test correctly (81 %). Most preferred to take the self-sample at home in the future (88 %) because it was simple and did not require a doctor's appointment. Few women (126/1946, 7 %) who did not return a self-sample in the iPap trial returned the questionnaire. Their main

  7. Translational Targeted Proteomics Profiling of Mitochondrial Energy Metabolic Pathways in Mouse and Human Samples.

    Science.gov (United States)

    Wolters, Justina C; Ciapaite, Jolita; van Eunen, Karen; Niezen-Koning, Klary E; Matton, Alix; Porte, Robert J; Horvatovich, Peter; Bakker, Barbara M; Bischoff, Rainer; Permentier, Hjalmar P

    2016-09-01

    Absolute measurements of protein abundance are important in the understanding of biological processes and the precise computational modeling of biological pathways. We developed targeted LC-MS/MS assays in the selected reaction monitoring (SRM) mode to quantify over 50 mitochondrial proteins in a single run. The targeted proteins cover the tricarboxylic acid cycle, fatty acid β-oxidation, oxidative phosphorylation, and the detoxification of reactive oxygen species. Assays used isotopically labeled concatemers as internal standards designed to target murine mitochondrial proteins and their human orthologues. Most assays were also suitable to quantify the corresponding protein orthologues in rats. After exclusion of peptides that did not pass the selection criteria, we arrived at SRM assays for 55 mouse, 52 human, and 51 rat proteins. These assays were optimized in isolated mitochondrial fractions from mouse and rat liver and cultured human fibroblasts and in total liver extracts from mouse, rat, and human. The developed proteomics approach is suitable for the quantification of proteins in the mitochondrial energy metabolic pathways in mice, rats, and humans as a basis for translational research. Initial data show that the assays have great potential for elucidating the adaptive response of human patients to mutations in mitochondrial proteins in a clinical setting.

  8. A PCR-DGGE method for detection and identification of Campylobacter, Helicobacter, Arcobacter and related Epsilobacteria and its application to saliva samples from humans and domestic pets

    DEFF Research Database (Denmark)

    Petersen, Randi Føns; Harrington, C. S.; Kortegaard, H. E.

    2007-01-01

    distinguished. This approach detected Epsilobacteria in all saliva samples collected from humans, cats and dogs, and identified Campylobacter concisus and/or Campylobacter gracilis in the human samples. The pet animal samples were taken from individuals with oral/dental diseases; PCR-DGGE identified up to four...

  9. Biomarkers for Detection and Monitoring of B16 Melanoma in Mouse Urine and Feces

    Directory of Open Access Journals (Sweden)

    Aviv Sever

    2015-01-01

    Full Text Available Melanoma is the most malignant type of skin cancer. Early detection of melanoma is thus critical for patient prognosis and survival. At present, examination by a skilled dermatologist followed by biopsy of suspicious lesions is the diagnostic gold standard. The aim of the present study was to examine an alternative and noninvasive method for the diagnosis of melanoma at an early stage. We identified and compared the volatile organic compounds (VOCs in mouse urine and feces, before and after a subcutaneous injection of B16 melanoma cells. We identified a total of 16 VOCs in urine and 13 VOCs in feces that could serve as potential biomarkers. Statistical analysis significantly discriminated between the cancer and control groups. These results should be validated in a larger-scale animal study, after which a study could be designed in patients to develop a melanoma biomarker.

  10. Comparative study of cultivation of feces in vermiculite or charcoal to obtain larvae of Strongyloides venezuelensis

    Directory of Open Access Journals (Sweden)

    Steveen Rios Ribeiro

    2014-10-01

    Full Text Available Introduction We compared feces culturing in charcoal or vermiculite to obtain Strongyloides venezuelensis larvae. Methods Feces (5g from infected rats was mixed with vermiculite (10g or coal (10g in plastic cups and incubated at 28°C for 48h. Larvae were recovered using Baermann-Moraes method. Results Significantly higher number of positive larval cultures were recovered from vermiculite than from charcoal (15/17 and 4/17, respectively; p < 0.001; 990.6 ± 307.5 and 215 ± 78.1 larvae, p = 0.027. Conclusions Vermiculite yields more larvae and provides cleaner pellets, improving larvae identification and facilitating their use for other purposes.

  11. Intestinal ciliate composition found in the feces of racing horses from Izmir, Turkey.

    Science.gov (United States)

    Gürelli, Gözde; Göçmen, Bayram

    2012-08-01

    Species composition and distribution of intestinal ciliates were investigated in the feces from 15 racing horses living near Izmir, Turkey. Thirty-seven species belonging to 21 genera were identified. Although no new species were observed, this is the first report on intestinal ciliates in racing horses living in Turkey. The mean number of ciliates was 26.4 ± 13.9 × 10(4) cells ml(-1) of feces and the mean number of ciliate species per host was 18.8 ± 7.1. No ciliates were observed in one horse. Bundleia and Polymorphella were found to be the two dominant genera, occurring in high proportions. In contrast, Didesmis and Prorodonopsis were only observed at a low frequency. Bundleia nana, Blepharoconus hemiciliatus, Paraisotrichopsis composita, Prorodonopsis coli and Spirodinium equi were newly recorded from Turkey. Copyright © 2012 Elsevier GmbH. All rights reserved.

  12. Early and Non-Invasive Detection of Chronic Wasting Disease Prions in Elk Feces by Real-Time Quaking Induced Conversion

    Science.gov (United States)

    Cheng, Yo Ching; Hannaoui, Samia; John, Theodore R.; Dudas, Sandor; Czub, Stefanie; Gilch, Sabine

    2016-01-01

    Chronic wasting disease (CWD) is a fatal prion disease of wild and captive cervids in North America. Prions are infectious agents composed of a misfolded version of a host-encoded protein, termed PrPSc. Infected cervids excrete and secrete prions, contributing to lateral transmission. Geographical distribution is expanding and case numbers in wild cervids are increasing. Recently, the first European cases of CWD have been reported in a wild reindeer and two moose from Norway. Therefore, methods to detect the infection early in the incubation time using easily available samples are desirable to facilitate effective disease management. We have adapted the real-time quaking induced conversion (RT-QuIC) assay, a sensitive in vitro prion amplification method, for pre-clinical detection of prion seeding activity in elk feces. Testing fecal samples from orally inoculated elk taken at various time points post infection revealed early shedding and detectable prion seeding activity throughout the disease course. Early shedding was also found in two elk encoding a PrP genotype associated with reduced susceptibility for CWD. In summary, we suggest that detection of CWD prions in feces by RT-QuIC may become a useful tool to support CWD surveillance in wild and captive cervids. The finding of early shedding independent of the elk’s prion protein genotype raises the question whether prolonged survival is beneficial, considering accumulation of environmental prions and its contribution to CWD transmission upon extended duration of shedding. PMID:27829062

  13. Susceptibility to antifungal agents of Candida spp. from blood and feces collected in Novi Sad in 3-year period (2008-2010

    Directory of Open Access Journals (Sweden)

    Jelesić Zora Z.

    2011-01-01

    Full Text Available Candidemia is an important emerging nosocomial infection in patients with risk factors. Candida species from nonsterile sites can give insight into the characteristics of strains that may cause invasive disease. The aim of this study was to evaluate antifungal susceptibility of Candida blood and fecal isolates in Novi Sad, Vojvodina. During a 3-year period (2008 to 2010, 424 isolates of Candida spp. were collected, 30 bloodstream isolates and 394 strains from fecal samples. In vitro susceptibility of these isolates to five antifungal agents was established using commercial ATB FUNGUS 3 (Bio-Mérieux. Predominant species was Candida albicans (6 isolates from blood and 269 from feces. Resistance to one or more antifungal agents was less common in Candida albicans (3.63% than in other species (24.83%. Resistance to itraconazole was the most commonly found in both groups of isolates, 9.64% strains from feces and 20% from blood samples. Twelve isolates were multiply resistant, usually to fluconazole, itraconazole, and voriconazole. Resistance to amphotericine B was extremely rare. Although resistance to antimycotics of Candida spp. is rare at present, continued surveillance of antifungal susceptibility is necessary in order to monitor trends, and to choose the right empiric therapy.

  14. Slide Coagglutination for Salmonella typhi Antigens in Broths Inoculated with Feces from Typhoid Fever Patients

    Science.gov (United States)

    1981-12-01

    SLIDE COAGGLUTINATION FOR SALMONELLA TYPHI ANTIGENS IN BROTHS INOCULATED WITH FECES FROM TYPHOID FEVER PATIENTS R. C. Rockhill, L. W. Rumans and M...permission of the Editor, Southeast Asian Journal of Tropical Medicine and Public Health SLIDE COAGGLUTINATION FOR SALMONELLA TYPHI ANTIGENS IN...525 Vol. 12 No. 4 December 1981 1 1P .. .. . --U- 1- "J SLIDE COAGGLUTINATION O Salmonella typhi ANTIGFNS the Infectious Disease Hospital and cultured

  15. Characterization of a New Calicivirus Isolated from Feces of a Dog,

    Science.gov (United States)

    Canine calicivirus (CaCV), isolated from feces of a dog with diarrhea, was readily propagated in cultures of canine cells and in a dolphin cell line...although some of the properties differed slightly from those of previously described caliciviruses ; evidence was also obtasined for caliciviral RNA...species in infected cells. Based on tests with antisera to numerous caliciviruses ; evidence was also obtained for caliciviral RNA species in infected

  16. Fragmentation of contaminant and endogenous DNA in ancient samples determined by shotgun sequencing; prospects for human palaeogenomics.

    Directory of Open Access Journals (Sweden)

    Marc García-Garcerà

    Full Text Available BACKGROUND: Despite the successful retrieval of genomes from past remains, the prospects for human palaeogenomics remain unclear because of the difficulty of distinguishing contaminant from endogenous DNA sequences. Previous sequence data generated on high-throughput sequencing platforms indicate that fragmentation of ancient DNA sequences is a characteristic trait primarily arising due to depurination processes that create abasic sites leading to DNA breaks. METHODOLOGY/PRINCIPALS FINDINGS: To investigate whether this pattern is present in ancient remains from a temperate environment, we have 454-FLX pyrosequenced different samples dated between 5,500 and 49,000 years ago: a bone from an extinct goat (Myotragus balearicus that was treated with a depurinating agent (bleach, an Iberian lynx bone not subjected to any treatment, a human Neolithic sample from Barcelona (Spain, and a Neandertal sample from the El Sidrón site (Asturias, Spain. The efficiency of retrieval of endogenous sequences is below 1% in all cases. We have used the non-human samples to identify human sequences (0.35 and 1.4%, respectively, that we positively know are contaminants. CONCLUSIONS: We observed that bleach treatment appears to create a depurination-associated fragmentation pattern in resulting contaminant sequences that is indistinguishable from previously described endogenous sequences. Furthermore, the nucleotide composition pattern observed in 5' and 3' ends of contaminant sequences is much more complex than the flat pattern previously described in some Neandertal contaminants. Although much research on samples with known contaminant histories is needed, our results suggest that endogenous and contaminant sequences cannot be distinguished by the fragmentation pattern alone.

  17. Concentration of organochlorines in human brain, liver, and adipose tissue autopsy samples from Greenland

    DEFF Research Database (Denmark)

    Dewailly, Éric; Mulvad, Gert; Pedersen, Henning S.

    1999-01-01

    report results of organochlorine determination in liver, brain, omental fat, and subcutaneous abdominal fat samples collected from deceased Greenlanders between 1992 and 1994. Eleven chlorinated pesticides and 14 polychlorinated biphenyl congeners were measured in tissue lipid extracts by high...

  18. Assessing Acceptability of Self-Sampling Kits, Prevalence, and Risk Factors for Human Papillomavirus Infection in American Indian Women.

    Science.gov (United States)

    Winer, Rachel L; Gonzales, Angela A; Noonan, Carolyn J; Cherne, Stephen L; Buchwald, Dedra S

    2016-10-01

    We evaluated the feasibility and acceptability of self-sampling for human papillomavirus (HPV) testing and calculated the prevalence of and risk factors for high-risk (hr) HPV infections in a community-based sample of American Indian women. To this end, we recruited 329 Hopi women aged 21-65 years to self-collect vaginal samples for hrHPV testing. Samples were tested by polymerase chain reaction for 14 hrHPV genotypes. We used Chi square tests to identify correlates of preference for clinician Pap testing versus HPV self-sampling, and age-adjusted Poisson regression to evaluate correlates of hrHPV prevalence. We found that satisfaction with HPV self-sampling was high, with 96 % of women reporting that the sample was easy to collect and 87 % reporting no discomfort. The majority (62 %) indicated that they preferred HPV self-sampling to receiving a Pap test from a clinician. Preference for Pap testing over HPV self-sampling was positively associated with adherence to Pap screening and employment outside the home. All samples evaluated were satisfactory for HPV testing, and 22 % were positive for hrHPV. HrHPV prevalence peaked in the late 20 s and declined with increasing age. HrHPV positivity was inversely associated with having children living the household. In conclusion, HPV self-sampling is feasible and acceptable to Hopi women, and could be effective in increasing rates of cervical cancer screening in Hopi communities. HrHPV prevalence was similar to estimates in the general United States population.

  19. Molecular Diagnosis of Strongyloides Stercoralis Infection by PCR Detection of Specific DNA in Human Stool Samples

    Directory of Open Access Journals (Sweden)

    Eb Kia

    2011-06-01

    Full Text Available Background: Strongyloidiasis is mostly an asymptomatic infection and diagnosis of latent infec­tions is difficult due to limitations of current parasitological and serological methods. This study was conducted to set up a PCR-based method for molecular diagnosis of Strongyloides stercor­alis infection by detection of copro-DNA in stool samples.Methods: A total of 782 fresh stool samples were collected and examined by agar plate culture. Among those sixteen stool samples, which confirmed to be infected with S. stercoralis were exam­ined as positive control to set up each single and nested PCR, using two primer sets design­ing to amplify partial ribosomal DNA of S. stercoralis genome. Since, single PCR method yielded higher efficacy in detecting positive samples, in the second step, 30 stool samples, which found negative for S. stercoralis by agar plate culture of single stool sample, were examined by sin­gle PCR. Data analysis was performed using McNemar's χ2 test, with consideration of a P-value of <0.05 as indication of significant difference.Results: In amplification of DNA extracted from stool samples, single PCR detected S. stercor­alis DNA target in all 16 positive samples, while nested PCR amplified DNA in only 75% of sam­ples. In the second step, single PCR amplified S. stercoralis extracted DNA in 5 out of 30 sam­ples which were negative by coproculture.Conclusion: Single PCR method amplifying a short (100bp target represented more efficacies for detection of S. stercoralis in faecal examination compared to agar plate culture and nested PCR, which amplified longer target.

  20. Insights into the processes behind the contamination of degraded human teeth and bone samples with exogenous sources of DNA

    DEFF Research Database (Denmark)

    Gilbert, M. T. P.; Hansen, Anders J.; Willerslev, E.

    2006-01-01

    A principal problem facing human DNA studies that use old and degraded remains is contamination from other sources of human DNA. In this study we have attempted to contaminate deliberately bones and teeth sampled from a medieval collection excavated in Trondheim, Norway, in order to investigate...... and are difficult to decontaminate using the tested protocol. We believe that this is largely due to the porous nature of bone and teeth facilitating the deep penetration of the contaminant DNA. To simulate a more realistic handling situation, 27 further teeth were directly handled and washed, then decontaminated...

  1. Determination of trace elements in human liver biopsy samples by ICP-MS and TXRF: hepatic steatosis and nickel accumulation.

    Science.gov (United States)

    Varga, Imre; Szebeni, Agnes; Szoboszlai, Norbert; Kovács, Béla

    2005-10-01

    Human liver biopsy samples, collected from 52 individuals, were analysed by inductively coupled plasma-mass spectrometry (ICP-MS) and total reflection X-ray fluorescence (TXRF) spectrometry in a retrospective study (i.e. patient selection and liver biopsy were not for the purpose of element analysis). The freeze-dried samples (typically 0.5-2 mg dry weight) were digested in a laboratory microwave digestion system and solutions with a final volume of 1 mL were prepared. The concentrations of Cr, Mn, Fe, Ni, Cu, Zn, Rb, and Pb were determined by use of a Thermo Elemental X7 ICP-MS spectrometer. TXRF measurements were performed with an Atomika Extra IIA spectrometer. Yttrium was employed as an internal standard, prepared by dissolution of 5N-purity yttria (Y(2)O(3)) in our laboratory. The accuracy was tested by analysis of NIST 1577a Bovine Liver certified reference material. The concentrations of Fe, Cu, Zn, and Rb determined in human liver biopsy samples were in good agreement with data published by other authors. The distribution of nickel in the samples was surprisingly uneven-nickel concentrations ranged from 0.7 to 12 microg g(-1) (dry weight) in 38 samples and in several samples were extremely high, 36-693 microg g(-1). Analysis of replicate procedural blanks and control measurements were performed to prevent misinterpretation of the data. For patients with steatosis (n=14) Ni concentrations were consistently high except for two who had levels close to those measured for the normal group. As far as we are aware no previous literature data are available on the association of steatosis with high concentration of nickel in human liver biopsies taken from living patients.

  2. Chromosome aberrations induced in human lymphocytes by U-235 fission neutrons: I. Irradiation of human blood samples in the "dry cell" of the TRIGA Mark II nuclear reactor.

    Science.gov (United States)

    Fajgelj, A; Lakoski, A; Horvat, D; Remec, I; Skrk, J; Stegnar, P

    1991-11-01

    A set-up for irradiation of biological samples in the TRIGA Mark II research reactor in Ljubljana is described. Threshold activation detectors were used for characterisation of the neutron flux, and the accompanying gamma dose was measured by TLDs. Human peripheral blood samples were irradiated "in vitro" and biological effects evaluated according to the unstable chromosomal aberrations induced. Biological effects of two types of cultivation of irradiated blood samples, the first immediately after irradiation and the second after 96 h storage, were studied. A significant difference in the incidence of chromosomal aberrations between these two types of samples was obtained, while our dose-response curve fitting coefficients alpha 1 = (7.71 +/- 0.09) x 10(-2) Gy-1 (immediate cultivation) and alpha 2 = (11.03 +/- 0.08) x 10(-2) Gy-1 (96 h delayed cultivation) are in both cases lower than could be found in the literature.

  3. Demonstration of carboxylesterase in cytology samples of human nasal respiratory epithelium

    Energy Technology Data Exchange (ETDEWEB)

    Rodgers, D.A.; Nikula, K.J.; Avila, K. [and others

    1995-12-01

    The epithelial lining of the nasal airways is a target for responses induced by a variety of toxicant exposures. The high metabolic capacity of this tissue has been suggested to play a role in both protection of the airways through detoxication of certain toxicants, as well as in activation of other compounds to more toxic metabolites. Specifically, nasal carboxylesterase (CE) has been shown to mediate the toxicity of inhaled esters and acrylates by converting them to more toxic acid and alcohol metabolites which can be cytotoxic and/or carcinogenic to the nasal mucosa. Due to difficulties in extrapolating rodent models to human, new paradigms using human cells and tissues are essential to understanding and evaluating the metabolic processes in human nasal epithelium.

  4. Genotypic characterization of Staphylococcus aureus obtained from humans and bovine mastitis samples in India

    Directory of Open Access Journals (Sweden)

    K Prashanth

    2011-01-01

    Full Text Available Aim and Background: Staphylococcus aureus is a major human pathogen that also causes important infections in cattle and sheep. The present study aimed to test genetic diversity among strains of S. aureus isolated from cattle (n=34 and humans (n=22 by DNA typing. Materials and Methods: Fluorescent amplified fragment length polymorphism (FAFLP is the genotyping tool used in the study. The presence of the mecA and Panton-Valentine leukocidin (PVL genes among these strain groups was also checked. Results: A dendrogram deduced from FAFLP showed that all the strains clustered into 10 groups (A-J with a relative genetic divergence of less than 8%. Sixty-seven percent of the isolates from bovine sources clustered together in two clades (A and H, while another major cluster with 13 isolates (59% (Cluster G had all strains from a human host. The remaining strains from both the hosts clustered independently into smaller clusters with the exception of two strains of human origin, which clustered along with a bovine cluster. Thirteen strains belonging to cluster G were highly clonal. About 77% of strains obtained from human infections were methicillin-resistant S. aureus (MRSA, whereas only 29% of strains from bovine origin were MRSA. Only three strains from human origin showed PVL positive, while no strain from cattle had PVL genes. The complete absence of PVL genes in all the bovine strains in the study appears to be significant. Conclusions: FAFLP can be successfully applied to assess the genetic relationship of S. aureus isolates from different hosts. The study also provided the valuable epidemiological data on S. aureus from bovine sources in India, which is lacking.

  5. A new liquid chromatography-tandem mass spectrometry method for determination of parabens in human placental tissue samples.

    Science.gov (United States)

    Jiménez-Díaz, I; Vela-Soria, F; Zafra-Gómez, A; Navalón, A; Ballesteros, O; Navea, N; Fernández, M F; Olea, N; Vílchez, J L

    2011-05-15

    Endocrine disruptors are a group of organic compounds widely used, which are ubiquitous in the environment and in biological samples. The main effect of these compounds is associated with their ability to mimic or block the action of natural hormones in living organisms, including humans. Parabens (esters of p-hydroxybenzoic acid) belong to this group of compounds. In this work, we propose a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to asses the presence of parabens most commonly used in industrial applications (methyl-, ethyl-, propyl- and butyl-paraben) in samples of human placental tissue. The method involves the extraction of the analytes from the samples using ethyl acetate, followed by a clean-up step using centrifugation prior to their quantification by LC-MS/MS using an atmospheric pressure chemical ionization (APCI) interface in the negative mode. Deuterated bisphenol A (BPA-d(16)) was used as surrogate. Found detection limits (LOD) ranged from 0.03 to 0.06 ng g(-1) and quantification limits (LOQ) from 0.1 to 0.2 ng g(-1), while inter- and intra-day variability was under 13.8%. The method was validated using standard addition calibration and a spike recovery assay. Recovery rates for spiked samples ranged from 82% to 108%. This method was satisfactorily applied for the determination of parabens in 50 placental tissue samples collected from women who live in the province of Granada (Spain).

  6. Optimizing human semen cryopreservation by reducing test vial volume and repetitive test vial sampling

    DEFF Research Database (Denmark)

    Jensen, Christian F S; Ohl, Dana A; Parker, Walter R

    2015-01-01

    OBJECTIVE: To investigate optimal test vial (TV) volume, utility and reliability of TVs, intermediate temperature exposure (-88°C to -93°C) before cryostorage, cryostorage in nitrogen vapor (VN2) and liquid nitrogen (LN2), and long-term stability of VN2 cryostorage of human semen. DESIGN: Prospec......OBJECTIVE: To investigate optimal test vial (TV) volume, utility and reliability of TVs, intermediate temperature exposure (-88°C to -93°C) before cryostorage, cryostorage in nitrogen vapor (VN2) and liquid nitrogen (LN2), and long-term stability of VN2 cryostorage of human semen. DESIGN...

  7. [Evolution of intrabacterial glucides during differentiation of two types of feces in the colon of the domestic rabbit].

    Science.gov (United States)

    Bonnafous, R; Raynaud, P

    1978-02-27

    In the Rabbit, during excretion of hard feces, a correlation exists between the stock of bacterial carbohydrates at the caecal level and the intensity of the bacterial lysis observed in the proximal colon.

  8. Direct determination of lead in human urine and serum samples by electrothermal atomic absorption spectrometry and permanent modifiers

    OpenAIRE

    Andrada,Daniel; Pinto,Frederico G.; Magalhães, Cristina Gonçalves; Nunes,Berta R.; Franco,Milton B.; Silva,José Bento Borba da

    2006-01-01

    The object of the present study was the development of alternative methods for the direct determination of lead in undigested samples of human urine and serum by electrothermal atomic absorption spectrometry (ET AAS). Thus, some substances have been investigated to act as chemical modifiers. Volumes of 20 µL of diluted samples, 1 + 1, v/v for urine and 1 + 4, v/v for serum, with HNO3 1% v/v and 0.02% v/v of cetil trimethyl ammonium chloride (CTAC) were prepared directly in the autosampler cup...

  9. Determination of total selenium and Se-77 in isotopically enriched human samples by ICP-dynamic reaction cell-MS

    DEFF Research Database (Denmark)

    Sloth, Jens Jørgen; Larsen, Erik Huusfeldt; Bügel, Susanne H.;

    2003-01-01

    This paper describes an analytical method for the simultaneous quantitative determination of total selenium (Se) and Se-77 in isotopically enriched human plasma, urine and faeces by inductively coupled plasma- dynamic reaction cell- mass spectrometry ( ICP- DRC- MS). The samples originated from...... and the digested faecal samples were diluted using an aqueous diluent containing 0.5% Triton X-100, 2% nitric acid and 3% methanol. Selenium was detected as Se-76, Se-77 and Se-80 by ICP- DRC- MS. Selenium originating from the natural isotope abundance yeast and other selenium sources from the diet was determined...

  10. Detection of Clostridium tetani in human clinical samples using tetX specific primers targeting the neurotoxin.

    Science.gov (United States)

    Ganesh, Madhu; Sheikh, Nasira K; Shah, Pooja; Mehetre, Gajanan; Dharne, Mahesh S; Nagoba, Basavraj S

    2016-01-01

    Tetanus resulting from ear injury remains an important health problem, particularly in the developing world. We report the successful detection of Clostridium tetani using tetX specific primers targeting the Cl. tetani neurotoxin. The sample was obtained from an ear discharge of a case of otogenic tetanus in a 2-year-old male child. Based on the culture results of the ear discharge, Gram staining and virulence testing by genotyping, a diagnosis of tetanus was confirmed. This is the first report from India on the successful detection of Cl. tetani in a human clinical sample using tetX specific primers targeting the Cl. tetani neurotoxin.

  11. Goat milk as a non-invasive sample for confirmation of Mycobacterium avium subspecies paratuberculosis by IS900 PCR

    Directory of Open Access Journals (Sweden)

    Bharathy Sukumar

    2014-09-01

    Full Text Available Mycobacterium avium subsp. paratuberculosis (MAP causes Johne's disease (JD in cattle, sheep, goats and other ruminants, and Crohn’s disease in humans. MAPs are shed to external environment through feces and milk. The present study was aimed to evaluate the utility of milk as a non-invasive sample in stage II MAP infections in goats using IS900 polymerase chain reaction (PCR and sequencing analysis. A total of 32 milk samples from lactating does were collected. Within these 32 milk samples, 15 were collected from pre-confirmed JD positive goats. By IS900 PCR, all the 15 (100% known JD positive goat milk samples revealed the presence of MAP. However, no unknown goat was identified as MAP positive. The results of this study established the usefulness of milk as a non-invasive sample in screening and confirmation of stage II MAP infection in goats.

  12. Bridging the gap between sample collection and laboratory analysis: using dried blood spots to identify human exposure to chemical agents

    Science.gov (United States)

    Hamelin, Elizabeth I.; Blake, Thomas A.; Perez, Jonas W.; Crow, Brian S.; Shaner, Rebecca L.; Coleman, Rebecca M.; Johnson, Rudolph C.

    2016-05-01

    Public health response to large scale chemical emergencies presents logistical challenges for sample collection, transport, and analysis. Diagnostic methods used to identify and determine exposure to chemical warfare agents, toxins, and poisons traditionally involve blood collection by phlebotomists, cold transport of biomedical samples, and costly sample preparation techniques. Use of dried blood spots, which consist of dried blood on an FDA-approved substrate, can increase analyte stability, decrease infection hazard for those handling samples, greatly reduce the cost of shipping/storing samples by removing the need for refrigeration and cold chain transportation, and be self-prepared by potentially exposed individuals using a simple finger prick and blood spot compatible paper. Our laboratory has developed clinical assays to detect human exposures to nerve agents through the analysis of specific protein adducts and metabolites, for which a simple extraction from a dried blood spot is sufficient for removing matrix interferents and attaining sensitivities on par with traditional sampling methods. The use of dried blood spots can bridge the gap between the laboratory and the field allowing for large scale sample collection with minimal impact on hospital resources while maintaining sensitivity, specificity, traceability, and quality requirements for both clinical and forensic applications.

  13. Urine versus brushed samples in human papillomavirus screening: study in both genders.

    NARCIS (Netherlands)

    Hauwers, K.W.M. d'; Depuydt, C.; Bogers, J.P.; Stalpaert, M.; Vereecken, A.; Wyndaele, J.J.; Tjalma, W.

    2007-01-01

    AIM: To investigate whether urine is a good medium for screening and whether there is a correlation between the amount of extracted DNA and human papillomavirus (HPV)-positivity. METHODS: In the present study, 30 first-voided urine (FVU) specimens and 20 urethroglandular swabs using cervex-brushes f

  14. Mycotoxin detection in human samples from patients exposed to environmental molds.

    Science.gov (United States)

    Hooper, Dennis G; Bolton, Vincent E; Guilford, Frederick T; Straus, David C

    2009-04-01

    The goal of this study was to determine if selected mycotoxins (trichothecenes, aflatoxins, and ochratoxins) could be extracted and identified in human tissue and body fluids from patients exposed to toxin producing molds in their environment. Human urine and methanol extracted tissues and sputum were examined. Trichothecenes were tested using competitive ELISA techniques. Aflatoxins B1, B2, G1, and G2, and ochratoxin A were tested by using immunoaffinity columns and fluorometry. Test sensitivity and specificity were determined. Levels of detection for the various mycotoxins varied from 0.2 ppb for trichothecenes, 1.0 ppb for aflatoxins, and 2.0 ppb for ochratoxins. Trichothecene levels varied in urine, sputum, and tissue biopsies (lung, liver, brain) from undetectable ( 10.0 ppb. Negative control patients had no detectable mycotoxins in their tissues or fluids. These data show that mycotoxins can be detected in body fluids and human tissue from patients exposed to mycotoxin producing molds in the environment, and demonstrate which human tissues or fluids are the most likely to yield positive results.

  15. Isolation of DNA from bacterial samples of the human gastrointestinal tract

    NARCIS (Netherlands)

    Zoetendal, E.G.; Heilig, G.H.J.; Klaassens, E.S.; Booijink, C.C.G.M.; Kleerebezem, M.; Smidt, H.; Vos, de W.M.

    2006-01-01

    The human gastrointestinal (GI) tract contains a complex microbial community that develops in time and space. The most widely used approaches to study microbial diversity and activity are all based on the analysis of nucleic acids, DNA, rRNA and mRNA. Here, we present a DNA isolation protocol that i

  16. A novel minimally-invasive method to sample human endothelial cells for molecular profiling.

    Directory of Open Access Journals (Sweden)

    Stephen W Waldo

    Full Text Available The endothelium is a key mediator of vascular homeostasis and cardiovascular health. Molecular research on the human endothelium may provide insight into the mechanisms underlying cardiovascular disease. Prior methodology used to isolate human endothelial cells has suffered from poor yields and contamination with other cell types. We thus sought to develop a minimally invasive technique to obtain endothelial cells derived from human subjects with higher yields and purity.Nine healthy volunteers underwent endothelial cell harvesting from antecubital veins using guidewires. Fluorescence-activated cell sorting (FACS was subsequently used to purify endothelial cells from contaminating cells using endothelial surface markers (CD34/CD105/CD146 with the concomitant absence of leukocyte and platelet specific markers (CD11b/CD45. Endothelial lineage in the purified cell population was confirmed by expression of endothelial specific genes and microRNA using quantitative polymerase chain reaction (PCR.A median of 4,212 (IQR: 2161-6583 endothelial cells were isolated from each subject. Quantitative PCR demonstrated higher expression of von Willebrand Factor (vWF, P<0.001, nitric oxide synthase 3 (NOS3, P<0.001 and vascular cell adhesion molecule 1 (VCAM-1, P<0.003 in the endothelial population compared to similarly isolated leukocytes. Similarly, the level of endothelial specific microRNA-126 was higher in the purified endothelial cells (P<0.001.This state-of-the-art technique isolates human endothelial cells for molecular analysis in higher purity and greater numbers than previously possible. This approach will expedite research on the molecular mechanisms of human cardiovascular disease, elucidating its pathophysiology and potential therapeutic targets.

  17. A Novel Minimally-Invasive Method to Sample Human Endothelial Cells for Molecular Profiling

    Science.gov (United States)

    Waldo, Stephen W.; Brenner, Daniel A.; McCabe, James M.; Dela Cruz, Mark; Long, Brian; Narla, Venkata A.; Park, Joseph; Kulkarni, Ameya; Sinclair, Elizabeth; Chan, Stephen Y.; Schick, Suzaynn F.; Malik, Namita; Ganz, Peter; Hsue, Priscilla Y.

    2015-01-01

    Objective The endothelium is a key mediator of vascular homeostasis and cardiovascular health. Molecular research on the human endothelium may provide insight into the mechanisms underlying cardiovascular disease. Prior methodology used to isolate human endothelial cells has suffered from poor yields and contamination with other cell types. We thus sought to develop a minimally invasive technique to obtain endothelial cells derived from human subjects with higher yields and purity. Methods Nine healthy volunteers underwent endothelial cell harvesting from antecubital veins using guidewires. Fluorescence-activated cell sorting (FACS) was subsequently used to purify endothelial cells from contaminating cells using endothelial surface markers (CD34 / CD105 / CD146) with the concomitant absence of leukocyte and platelet specific markers (CD11b / CD45). Endothelial lineage in the purified cell population was confirmed by expression of endothelial specific genes and microRNA using quantitative polymerase chain reaction (PCR). Results A median of 4,212 (IQR: 2161 – 6583) endothelial cells were isolated from each subject. Quantitative PCR demonstrated higher expression of von Willebrand Factor (vWF, P<0.001), nitric oxide synthase 3 (NOS3, P<0.001) and vascular cell adhesion molecule 1 (VCAM-1, P<0.003) in the endothelial population compared to similarly isolated leukocytes. Similarly, the level of endothelial specific microRNA-126 was higher in the purified endothelial cells (P<0.001). Conclusion This state-of-the-art technique isolates human endothelial cells for molecular analysis in higher purity and greater numbers than previously possible. This approach will expedite research on the molecular mechanisms of human cardiovascular disease, elucidating its pathophysiology and potential therapeutic targets. PMID:25679506

  18. Geometric morphometric analysis of craniofacial variation, ontogeny and modularity in a cross-sectional sample of modern humans

    NARCIS (Netherlands)

    Wellens, H.L.L.; Kuijpers-Jagtman, A.M.; Halazonetis, D.J.

    2013-01-01

    This investigation aimed to quantify craniofacial variation in a sample of modern humans. In all, 187 consecutive orthodontic patients were collected, of which 79 were male (mean age 13.3, SD 3.7, range 7.5-40.8) and 99 were female (mean age 12.3, SD 1.9, range 8.7-19.1). The male and female subgrou

  19. Comparison of human glomerulus proteomic profiles obtained from low quantities of samples by different mass spectrometry with the comprehensive database

    Directory of Open Access Journals (Sweden)

    Liu Zan

    2011-08-01

    Full Text Available Abstract Background We have previously constructed an in-depth human glomerulus proteome database from a large amount of sample for understanding renal disease pathogenesis and aiding the biomarker exploration. However, it is usually a challenge for clinical research to get enough tissues for large-scale proteomic characterization. Therefore, in this study, we focused on high-confidence proteomics analysis on small amounts of human glomeruli comparable to those obtained from biopsies using different mass spectrometers and compared these results to the comprehensive database. Results One microgram of human glomerular protein digest was analyzed each on five LC- combined mass spectrometers (LIT-TOF, LTQ-Orbitrap, Q-TOF, LIT and MALDI-TOF/TOF yielding 139, 185, 94, 255 and 108 proteins respectively identified with strict criteria to ensure high confidence (> 99% and low false discovery rate (FDR ( Conclusion This study showed representative human glomerulus proteomic profiles obtained from biopsies through analysis of comparable amounts of samples by different mass spectrometry. Our results implicated that high abundant proteins are more likely to be reproducibly identified in multiple mass spectrometers runs and different mass spectrometers. Furthermore, many podocyte essential proteins such as nephrin, podocin, podocalyxin and synaptopodin were also identified from the small samples in this study. Bioinformatic enrichment analysis results extended our understanding of the major glomerular proteins about their subcellular distributions and functions. The present study indicated that the proteins localized in certain cellular compartments, such as actin cytoskeleton, mitochondrial matrix, cell surface, basolateral plasma membrane, contractile fiber, proteinaceous extracellular matrix and adherens junction, represent high abundant glomerular proteins and these subcellular structures are also highly significantly over-represented in the glomerulus

  20. An investigation into the concurrent collection of human scent and epithelial skin cells using a non-contact sampling device.

    Science.gov (United States)

    Caraballo, Norma Iris; Mendel, Julian; Holness, Howard; La Salvia, Joel; Moroose, Tina; Eckenrode, Brian; Stockham, Rex; Furton, Kenneth; Mills, DeEtta

    2016-09-01

    In criminal investigations, the collection of human scent often employs a non-contact, dynamic airflow device, known as the Scent Transfer Unit 100 (STU-100), to transfer volatile organic compounds (VOCs) from an object/person onto a collection material that is subsequently presented to human scent discriminating canines. Human scent is theorized to be linked to epithelial skin cells that are shed at a relatively constant rate allowing both scent and cellular material to be deposited into the environment and/or onto objects. Simultaneous collection of cellular material, with adequate levels of nuclear deoxyribonucleic acid (nDNA), and human scent using a non-invasive methodology would facilitate criminal investigations. This study evaluated the STU-100 for the concurrent collection of human scent and epithelial skin cells from a porous (paper) and non-porous (stainless steel bar) object that was held for a specified period of time in the dominant hand of twenty subjects (10 females and 10 males). Human scent analysis was performed using headspace static solid-phase microextraction with gas chromatography-mass spectrometry (HS-SPME/GC-MS). A polycarbonate filter was used to trap epithelial skin cells which, upon extraction, were subsequently analyzed, inter-laboratory, using the quantitative polymerase chain reaction (qPCR). The STU-100 proved to be inadequate for collecting the minimum number of epithelial skin cells required to obtain nuclear DNA concentrations above the limit of detection for the qPCR kit. With regard to its use for human scent collection, a reduction in the number and mass of compounds was observed when compared to samples that were directly collected. However, when the indirect collection of human scent from the two different objects was compared, a greater number and mass of compounds was observed from the non-porous object than from the porous object. This outcome suggests that the matrix composition of the scent source could affect the

  1. Liquid chromatographic determination of quinolones in water and human urine samples after microextraction by packed sorbent.

    Science.gov (United States)

    Rani, Susheela; Kumar, Ashwini; Malik, Ashok Kumar; Singh, Baldev

    2012-01-01

    A method for the simultaneous determination of quinolones in water and urine samples by microextraction in a sorbent-packed syringe (MEPS) with LC is described. MEPS is a new miniaturized SPE technique that can be used with chromatographic instruments without any modifications. In MEPS, approximately 1 mg of the solid packing material is inserted into a syringe (100-250 microL) as a plug. Sample preparation takes place on the packed bed. The new method is promising, easy to use, economical, and rapid. The determination of quinolones in groundwater and urine was performed using MEPS as a sample preparation method with LC-UV determination. Four quinolone antibiotics--enrofloxacin, enoxacin, danofloxacin, and nalidixic acid--in groundwater and urine samples were used as analytes. The extraction recovery was found to be between 64.9 and 98.9%. The results showed high correlation coefficients (R2 > 0.992) for all of the analytes within the calibration range. The LOQ was between 0.091 and 0.315 ng/mL.

  2. Development and validation of a point-of-care test for detecting hantavirus antibodies in human and rodent samples.

    Science.gov (United States)

    Koishi, Andrea Cristine; Aoki, Mateus Nóbrega; Jorge, Taissa Ricciardi; Suzukawa, Andréia Akemi; Zanluca, Camila; Levis, Silvana; Duarte Dos Santos, Claudia Nunes

    2016-07-01

    Hantaviruses are etiologic agents of a zoonotic disease transmitted mainly from wild rodents to humans, causing Hemorrhagic Fever with Renal Syndrome in Eurasia and the Hantavirus Cardiopulmonary Syndrome in the Americas (HCPS), reaching a lethality rate of 40% in Brazil. Hantavirus diagnostic and seroprevalence are often based on the presence of IgM and IgG antibodies against the virus. Here we propose a rapid test assay able to identify hantavirus antibodies with sensibility and specificity similar to ELISA assays. We analyzed five groups of samples, including healthy human population and small mammals of endemic areas, suspected cases of HCPS, patients with non-related infections and a serum panel from a different geographical region. The test presented good rates of sensibility (87-100%) and specificity (97-100%) for all groups, being a promising tool suitable for both rodent and human hantavirus epidemiological surveys.

  3. Quantification of Human and Animal Viruses to Differentiate the Origin of the Fecal Contamination Present in Environmental Samples

    Directory of Open Access Journals (Sweden)

    Sílvia Bofill-Mas

    2013-01-01

    Full Text Available Many different viruses are excreted by humans and animals and are frequently detected in fecal contaminated waters causing public health concerns. Classical bacterial indicator such as E. coli and enterococci could fail to predict the risk for waterborne pathogens such as viruses. Moreover, the presence and levels of bacterial indicators do not always correlate with the presence and concentration of viruses, especially when these indicators are present in low concentrations. Our research group has proposed new viral indicators and methodologies for determining the presence of fecal pollution in environmental samples as well as for tracing the origin of this fecal contamination (microbial source tracking. In this paper, we examine to what extent have these indicators been applied by the scientific community. Recently, quantitative assays for quantification of poultry and ovine viruses have also been described. Overall, quantification by qPCR of human adenoviruses and human polyomavirus JC, porcine adenoviruses, bovine polyomaviruses, chicken/turkey parvoviruses, and ovine polyomaviruses is suggested as a toolbox for the identification of human, porcine, bovine, poultry, and ovine fecal pollution in environmental samples.

  4. Possible additional exposure to dioxin and dioxin-like compounds from waste incineration. Biomonitoring using human milk and animal samples

    Energy Technology Data Exchange (ETDEWEB)

    Sampaio, C.; M. Fatima Reis; J. Pereira Miguel [Inst. of Preventive Medicine, Univ. of Lisbon (Portugal); Murk, A. [Wageningen Univ., Dept. of Toxicology (Netherlands)

    2004-09-15

    In the ambit of an Environmental Health Survey Program relative to a MSW facility, which has been operating near to Lisbon since 1999 a biomonitoring study using human breast milk has been performed. Specific aims of this study were: (1) determine whether living in the vicinity of the incinerator increases dioxin maternal body burden and accordingly perinatal (intra-uterus and lactacional) exposure; (2) to investigate the possibility of increased human exposure to dioxins and dioxin-like compounds via locally produced food items from animal origin. Therefore, levels of dioxins and dioxin-like compounds have been determined in human milk samples collected in the vicinity of the incinerator and in a control area, for comparison. From the same areas, cow and sheep milk and eggs from free-range chickens have also been collected to get an indication of possible local additional exposure to air-borne dioxins via the food chain. Analyses of TCDD-equivalents (TEQs) were mainly performed with a reporter gene assay for dioxin-like activity, the DR-CALUX bioassay (Dioxin Responsive Chemical Activated LUciferase gene eXpression).To determine congeners profile, some human milk samples have also been analysed for PCDD/Fs and relevant dioxin-like PCBs, by using high-resolution gas chromatography and high-resolution mass spectrometry (HRGC/HRMS). Both the Ethics Committees of the Faculty of Medicine, University of Lisbon, and of the Maternity Dr. Alfredo da Costa have approved the study protocol.

  5. Gene expression data from acetaminophen-induced toxicity in human hepatic in vitro systems and clinical liver samples

    Directory of Open Access Journals (Sweden)

    Robim M. Rodrigues

    2016-06-01

    Full Text Available This data set is composed of transcriptomics analyses of (i liver samples from patients suffering from acetaminophen-induced acute liver failure (ALF and (ii hepatic cell systems exposed to acetaminophen and their respective controls. The in vitro systems include widely employed cell lines i.e. HepaRG and HepG2 cells as well as a novel stem cell-derived model i.e. human skin-precursors-derived hepatocyte-like cells (hSKP-HPC. Data from primary human hepatocytes was also added to the data set “Open TG-GATEs: a large-scale toxicogenomics database” (Igarashi et al., 2015 [1]. Changes in gene expression due to acetaminophen intoxication as well as comparative information between human in vivo and in vitro samples are provided. The microarray data have been deposited in NCBI׳s Gene Expression Omnibus and are accessible through GEO Series accession number GEO: GSE74000. The provided data is used to evaluate the predictive capacity of each hepatic in vitro system and can be directly compared with large-scale publically available toxicogenomics databases. Further interpretation and discussion of these data feature in the corresponding research article “Toxicogenomics-based prediction of acetaminophen-induced liver injury using human hepatic cell systems” (Rodrigues et al., 2016 [2].

  6. Occurrence of artificial sweeteners in human liver and paired blood and urine samples from adults in Tianjin, China and their implications for human exposure.

    Science.gov (United States)

    Zhang, Tao; Gan, Zhiwei; Gao, Chuanzi; Ma, Ling; Li, Yanxi; Li, Xiao; Sun, Hongwen

    2016-09-14

    In this study, acesulfame (ACE), saccharin (SAC) and cyclamate (CYC) were found in all paired urine and blood samples collected from healthy adults, with mean values of 4070, 918 and 628 ng mL(-1), respectively, in urine and 9.03, 20.4 and 0.72 ng mL(-1), respectively, in blood. SAC (mean: 84.4 ng g(-1)) and CYC (4.29 ng g(-1)) were detectable in all liver samples collected from liver cancer patients, while ACE was less frequently detected. Aspartame (ASP) was not found in any analyzed human sample, which can be explained by the fact that this chemical metabolized rapidly in the human body. Among all adults, significantly positive correlations between SAC and CYC levels were observed (p < 0.001), regardless of human matrices. Nevertheless, no significant correlations between concentrations of SAC (or CYC) and ACE were found in any of the human matrices. Our results suggest that human exposure to SAC and CYC is related, whereas ACE originates from a discrete source. Females (or young adults) were exposed to higher levels of SAC and CYC than males (or elderly). The mean renal clearance of SAC was 730 mL per day per kg in adults, which was significantly (p < 0.001) lower than those for CYC (10 800 mL per day per kg) and ACE (10 300 mL per day per kg). The average total daily intake of SAC and ACE was 9.27 and 33.8 μg per kg bw per day, respectively.

  7. Detection of head-to-tail DNA sequences of human bocavirus in clinical samples.

    Directory of Open Access Journals (Sweden)

    Jessica Lüsebrink

    Full Text Available Parvoviruses are single stranded DNA viruses that replicate in a so called "rolling-hairpin" mechanism, a variant of the rolling circle replication known for bacteriophages like φX174. The replication intermediates of parvoviruses thus are concatemers of head-to-head or tail-to-tail structure. Surprisingly, in case of the novel human bocavirus, neither head-to-head nor tail-to-tail DNA sequences were detected in clinical isolates; in contrast head-to-tail DNA sequences were identified by PCR and sequencing. Thereby, the head-to-tail sequences were linked by a novel sequence of 54 bp of which 20 bp also occur as conserved structures of the palindromic ends of parvovirus MVC which in turn is a close relative to human bocavirus.

  8. Homo floresiensis contextualized: a geometric morphometric comparative analysis of fossil and pathological human samples.

    Directory of Open Access Journals (Sweden)

    Karen L Baab

    Full Text Available The origin of hominins found on the remote Indonesian island of Flores remains highly contentious. These specimens may represent a new hominin species, Homo floresiensis, descended from a local population of Homo erectus or from an earlier (pre-H. erectus migration of a small-bodied and small-brained hominin out of Africa. Alternatively, some workers suggest that some or all of the specimens recovered from Liang Bua are pathological members of a small-bodied modern human population. Pathological conditions proposed to explain their documented anatomical features include microcephaly, myxoedematous endemic hypothyroidism ("cretinism" and Laron syndrome (primary growth hormone insensitivity. This study evaluates evolutionary and pathological hypotheses through comparative analysis of cranial morphology. Geometric morphometric analyses of landmark data show that the sole Flores cranium (LB1 is clearly distinct from healthy modern humans and from those exhibiting hypothyroidism and Laron syndrome. Modern human microcephalic specimens converge, to some extent, on crania of extinct species of Homo. However in the features that distinguish these two groups, LB1 consistently groups with fossil hominins and is most similar to H. erectus. Our study provides further support for recognizing the Flores hominins as a distinct species, H. floresiensis, whose affinities lie with archaic Homo.

  9. Homo floresiensis contextualized: a geometric morphometric comparative analysis of fossil and pathological human samples.

    Science.gov (United States)

    Baab, Karen L; McNulty, Kieran P; Harvati, Katerina

    2013-01-01

    The origin of hominins found on the remote Indonesian island of Flores remains highly contentious. These specimens may represent a new hominin species, Homo floresiensis, descended from a local population of Homo erectus or from an earlier (pre-H. erectus) migration of a small-bodied and small-brained hominin out of Africa. Alternatively, some workers suggest that some or all of the specimens recovered from Liang Bua are pathological members of a small-bodied modern human population. Pathological conditions proposed to explain their documented anatomical features include microcephaly, myxoedematous endemic hypothyroidism ("cretinism") and Laron syndrome (primary growth hormone insensitivity). This study evaluates evolutionary and pathological hypotheses through comparative analysis of cranial morphology. Geometric morphometric analyses of landmark data show that the sole Flores cranium (LB1) is clearly distinct from healthy modern humans and from those exhibiting hypothyroidism and Laron syndrome. Modern human microcephalic specimens converge, to some extent, on crania of extinct species of Homo. However in the features that distinguish these two groups, LB1 consistently groups with fossil hominins and is most similar to H. erectus. Our study provides further support for recognizing the Flores hominins as a distinct species, H. floresiensis, whose affinities lie with archaic Homo.

  10. Forensic Identification of Human Blood: comparison of two one-step presumptive tests for blood screening of crime scene samples.

    Directory of Open Access Journals (Sweden)

    Ana Flávia Belchior Andrade

    2014-08-01

    Full Text Available Blood is the most common body fluid found at crime scenes. One-step presumptive tests have been designed as a rapid immunological test for the qualitative detection of human hemoglobin in stool samples (faecal occult blood their usefulness for forensic purposes has been demonstrated before. In this study we compare Hexagon OBTI kit and FOB One-step Bioeasy kit sensitivity in the analysis of diluted blood samples. With Hexagon OBTI, positive test results are achieved in whole blood dilutions up to 1:1.000. Sensitivity decreased with aged samples, if samples were not stored under low temperatures regardless of which presumptive test is used. Whole blood tests must take into consideration that “hook” effect may interfere. Comparing both tests, OBTI Hexagon Kit is more sensible to detect diluted blood, showing a wider detection window in all conditions. This is interesting when analyzing forensic samples as forensic analysts usually do not know about the history of the analyzed sample before its collection.

  11. Human herpes virus-8 DNA in bronchoalveolar lavage samples from patients with AIDS-associated pulmonary Kaposi's sarcoma

    DEFF Research Database (Denmark)

    Benfield, T L; Dodt, K K; Lundgren, Jens Dilling

    1997-01-01

    Kaposi's sarcoma (KS) is the most frequent AIDS-associated neoplasm, and often disseminates to visceral organs, including the lungs. An ante-mortem diagnosis of pulmonary KS is difficult. Recently, DNA sequences resembling a new human herpes virus (HHV-8), have been identified in various forms...... of KS. We hypothesized that these sequences are present in samples obtained by bronchoalveolar lavage (BAL) in patients with pulmonary KS. Utilizing a nested polymerase chain reaction (PCR), 7/12 BAL cell samples from HIV-infected patients with endobronchial KS were positive for HHV-8 DNA. In contrast......, only 2/39 samples from HIV-infected patients without evidence of KS were positive (p = 0.007). Detection of HHV-8 in BAL cells of patients with pulmonary KS was highly specific (95%), with a sensitivity of 58% and a positive predictive value of 78%. In conclusion, HHV-8 is associated with pulmonary KS...

  12. Concentration of organochlorines in human brain, liver, and adipose tissue autopsy samples from Greenland

    DEFF Research Database (Denmark)

    Dewailly, Éric; Mulvad, Gert; Pedersen, Henning S.

    1999-01-01

    Organochlorines are persistent lipophilic compounds that accumulate in Inuit people living in circumpolar countries. Organochlorines accumulate as a result of the Inuits' large consumption of sea mammal fat; however, available data are limited to blood lipids, milk fat, and adipose tissue. We...... report results of organochlorine determination in liver, brain, omental fat, and subcutaneous abdominal fat samples collected from deceased Greenlanders between 1992 and 1994. Eleven chlorinated pesticides and 14 polychlorinated biphenyl congeners were measured in tissue lipid extracts by high......-resolution gas chromatography with electron capture detection. Mean concentrations of polychlorinated biphenyls, 2, 2'-bis(4-chlorophenyl)-1,1-dichloroethylene, ss-hexachlorocyclohexane, hexachlorobenzene, mirex, trans-nonachlor, and oxychlordane in adipose tissue samples from Greenlanders were 3-34-fold higher...

  13. Optimal sample storage and extraction procotols for reliable multilocus genotyping of the human parasite Schistosoma mansoni.

    Science.gov (United States)

    Van den Broeck, F; Geldof, S; Polman, K; Volckaert, F A M; Huyse, T

    2011-08-01

    Genotyping individual larval stages and eggs of natural parasite populations is complicated by the difficulty of obtaining reliable genotypes from low quantity DNA template. A suitable storage and extraction protocol, together with a thorough quantification of genotyping errors are therefore crucial for molecular epidemiological studies. Here we test the robustness, handling time, ease of use, cost effectiveness and success rate of various fixation (Whatman FTA(®) Classic and Elute Cards, 70% EtOH and RNAlater(®)) and subsequent DNA extraction methods (commercial kits and proteinase K protocol). None of these methods require a cooling chain and are therefore suitable for field collection. Based on a multiplex microsatellite PCR with nine loci the success and reliability of each technique is evaluated by the proportion of samples with at least eight scored loci and the proportion of genotyping errors. If only the former is taken into account, FTA(®) Elute is recommended (83% success; 44% genotyping error; 0.2 €/sample; 1h 20 m handling time). However, when also considering the genotyping errors, handling time and ease of use, we opt for 70% EtOH with the 96-well plate technology followed by a simple proteinase K extraction (73% success; 0% genotyping error; 0.2 €/sample; 15m handling time). For eggs we suggest (1) to pool all eggs per person in 1.5 ml tubes filled with 70% EtOH for transport and (2) to identify each egg to species level prior to genotyping. To this end we extended the Rapid diagnostic PCR developed by Webster et al. (2010) with a S. mansoni-specific primer to discriminate between S. mansoni, S. haematobium and S. bovis in a single PCR reaction. The success rate of genotyping eggs was 75% (0% genotyping error). This is the first study to incorporate genotyping errors through re-amplification for the evaluation of schistosome sampling protocols and the identification of error-prone loci.

  14. Faster fermentation of cooked carrot cell clusters compared to cell wall fragments in vitro by porcine feces.

    Science.gov (United States)

    Day, Li; Gomez, Justine; Øiseth, Sofia K; Gidley, Michael J; Williams, Barbara A

    2012-03-28

    Plant cell walls are the major structural component of fruits and vegetables, which break down to cell wall particles during ingestion (oral mastication) or food processing. The major health-promoting effect of cell walls occurs when they reach the colon and are fermented by the gut microbiota. In this study, the fermentation kinetics of carrot cell wall particle dispersions with different particle size and microstructure were investigated in vitro using porcine feces. The cumulative gas production and short-chain fatty acids (SCFAs) produced were measured at time intervals up to 48 h. The results show that larger cell clusters with an average particle size (d(0.5)) of 298 and 137 μm were more rapidly fermented and produced more SCFAs and gas than smaller single cells (75 μm) or cell fragments (50 μm), particularly between 8 and 20 h. Confocal microscopy suggests that the junctions between cells provides an environment that promotes bacterial growth, outweighing the greater specific surface area of smaller particles as a driver for more rapid fermentation. The study demonstrates that it may be possible, by controlling the size of cell wall particles, to design plant-based foods for fiber delivery and promotion of colon fermentation to maximize the potential for human health.

  15. Antibacterial metabolites secreted under glucose-limited environment of the mimicked proximal colon model by lactobacilli abundant in infant feces.

    Science.gov (United States)

    Kanjan, Pochanart; Hongpattarakere, Tipparat

    2016-09-01

    The most abundance of anti-Salmonella lactic acid bacteria (LAB) was found in feces of naturally born, exclusively breastfed Thai infants. Six strains of Lactobacillus plantarum and one strain of Lactobacillus paracasei were selected and identified. In the co-cultivation assay, L. plantarum subsp. plantarum I62 showed the strongest and broadest antibacterial activity against Escherichia coli, Shigella sonnei, Salmonella Paratyphi A, and Salmonella Typhimurium SA 2093 under the mimicked proximal colon condition, in which glucose and other nutrients were limited. According to GC-MS analysis, the major antibacterial contribution of organic acids secreted by L. plantarum I62 grown in the presence of glucose was dramatically reduced from 95.8 to 41.9 % under glucose-limited niche. The production of low-pK a acids, such as lactic, 1,2-benzenedicarboxylic, and 3-phenyllactic acids, was remarkably dropped. Surprisingly, higher-pK a acids such as 5-chlorobenzimidazole-2-carboxylic, pyroglutamic, palmitic, and oleic acids were enhanced. Moreover, cyclic dipeptides, ketones, alkanes, alcohols, and miscellaneous compounds, which were pH-independent antibacterial metabolites, became dominant. The electron microscopy strongly supported the synergistic attacks of the multiple antibacterial components targeting outer and cytoplasmic membranes leading to severe leakage and cell disruption of Salmonella Typhimurium. This strain poses to be a potential probiotic candidate for effectively controlling and treating human foodborne bacterial infection.

  16. Nitrogen isotopic patterns of vegetation as affected by breeding activity of Black-tailed Gull (Larus crassiostris): A coupled analysis of feces, inorganic soil nitrogen and flora

    Energy Technology Data Exchange (ETDEWEB)

    Mizota, C., E-mail: mizota@iwate-u.ac.jp [Iwate University, Ueda 3-18-8, Morioka, Iwate 020-8550 (Japan)

    2009-11-15

    Two currently breeding colonies (Matsushima Bay and Rishiri island; northern Japan) of predominant Black-tailed Gull (Larus crassiostris) were studied for N isotopic patterns of flora, which is affected by increased supply of inorganic soil N derived from the microbial transformation of feces. Coupled samples of feces, topsoil and flora were collected in early to mid July (2008), when input of fecal N onto soils was at its maximum. As bird migration and breeding continued, native Japanese red-pine (Pinus densiflora), junipers (Juniperus chinensis and Juniperus rigita; Matsushima Bay colony) and Sasa senanensis (Rishiri colony) declined, while ornithocoprophilus exotic plants succeeded. Among tree species on the islands, P. densiflora with ectomycorrizal colonization appears highly susceptible to elevated concentrations of NH{sub 4}-N in the topsoil. A mechanism for best explaining the plant succession associated with the breeding activity of Black-tailed Gull was evidenced by two parameters: first, concomitant elevation of N content in the flora and second, inorganic soil N content, along with changes in N isotopic composition ({delta}{sup 15}N). Earlier isotopic data on the foliar N affected by breeding activity were compiled and reviewed. Emphasis was put on isotopic information for inorganic N in soils that controls plant succession.

  17. Validation and quality control of ELISAs for the use with human saliva samples.

    Science.gov (United States)

    Jaedicke, Katrin M; Taylor, John J; Preshaw, Philip M

    2012-03-30

    Enzyme-linked immunosorbent assays (ELISAs) have proven to be a powerful tool for fast and reliable sample analysis, in both clinical diagnostics and in research. Most assays are now available for use with a range of different analytical fluids, including serum, plasma or urine. In recent years, saliva has drawn attention as a potentially valuable diagnostic fluid; however few ELISAs have been validated for use with saliva, or their validation is often incomplete. Saliva has a number of different physical characteristics than, for example, cell culture medium or serum and assuming an ELISA which works well with serum samples will also do so with saliva potentially could lead to erroneous data and conclusions. In this report, we provide a detailed protocol to validate any ELISA for use with saliva samples and show the results of validation procedures for 13 ELISAs for using saliva. Our findings suggest that the majority of ELISAs work reliably with saliva, even if the assay was not specifically designed for this biological fluid. However, we also report a few cases where recovery or intra-and inter-assay variations were unexpectedly high, emphasising the importance of performing a validation procedure for each assay before using it with saliva to ensure accurate and reliable data.

  18. Screening Procedure From Cattle Feces and the Prevalence of Escherichia coli O157 in Taiwan‘s Dairy Cattle

    Institute of Scientific and Technical Information of China (English)

    Chin-ChengChou; Tzu-MingPan; 等

    2001-01-01

    Objective:The aims of this study were to establish a screening procedure for the identification of Escherichia coliO157(E.coli O157) from bovine feces and to apply the procedure to the detection of bacteria in dairy herds in taiwan to locate the contaminated fieled.Methods:The Estabilished procedure for screeing E.coil O157 from bovine fecs in compsed of four steps:enrichment,selective culturing,phenotyping and gentyping.Modified trypticase soy broth (mTSB) containing 20 mg/L of novobioicin was used for the enrichment step.Using sorbitol MacConkey agar containing 0.05mg/L of cefixime and 2.5mg/L of potassium tellurite did the selective culturing step.The phenotyping step included the species confirmation of E.coli,the serotyping of O157 and H7 and the ability of verocytotoxin production that were tested by different commercial kits.The genotyping step for the confirmatioon of O157 antigen and verocytotoxin producing ability were performed by polymerase chain reaction.Results:mTSB had a better enrichment effect for E.Coli O157 than gram-negative broth had in the procedure,with mTSB medium,the effect was about 100-fold for selection.The detection limit of the screening procedure was 0.56±0.13CFU/g,Using the screening procedure described above,E.coli O157 was found in 3 of 1223(0.25%) fresh bovine fecal specimens,and 2 of 21(9.52%)dariy herds in Tainwan.Two of the current three screened strains were verocytotoxin II producting E.coli O157:H7 with the titer of 1:32;the other one was E.Coli O157:NM without verocytotoxin-producing ability.Conclusion:A procedure for the screening of E.coli O157 from bovine feces had been established successfully,Fiedl sampling results showed that dairy herds in Taiwan had the potertial risk of spreading E.coli O157.Bacterial Isolation of toxin-producing strains of E.Coli O157 indicated that public health concerns are necrssary.\\

  19. Laser-induced breakdown spectroscopy analysis of human deciduous teeth samples.

    Science.gov (United States)

    Khalid, Arooj; Bashir, Shazia; Akram, Mahreen; Hayat, Asma

    2015-12-01

    Laser-induced breakdown spectroscopy (LIBS) analysis of human deciduous teeth has been performed by employing Nd:YAG laser (1064 nm, 10 ns) for the evaluation of plasma parameters as well as elemental analysis. The plasma parameters, i.e., electron temperature and electron number density of laser-induced teeth plasma at various fluencies, have been evaluated. Both parameters show an increasing trend up to a certain value of laser fluence, i.e., 2.6 J/cm(2). With further increase in laser fluence up to a value of 3.9 J/cm(2), a decreasing trend is observed which is due to shielding effect. With further increase in laser fluence up to a maximum value of 10.5 J/cm(2), the insignificant changes in plasma parameters are observed which are attributed to saturation phenomenon governed by self-regulating regime. Emission spectroscopy results exhibit that laser fluence is the controlling factor for both plasma parameters. The elemental analysis was also performed at constant laser fluence of 2.6 J/cm(2) by evaluating the variation in detected elemental concentration of Ca, Fe, Sr, Zn, and Pb in three different parts of human teeth, i.e., enamel, dentine, and cementum. The lower concentration of Ca as compared to the standard values of CaCO3 (self-fabricated pellet) reveals that enamel is the most deciduous part of the human teeth. However, at the same time, it is also observed that the highest concentration of micro minerals is also found in enamel, then in dentine, and lowest in cementum. Carious or unhealthy tooth is identified by enhanced concentration of micro minerals (Pb, Sr, Zn, and Fe). The highest concentration of micro minerals as compared to other parts of teeth (dentine and root cementum) and lower concentration of Ca as compared to standard CaCO3 pellet in enamel confirm that enamel is the most deciduous part of the teeth.

  20. Heteropolymeric triplex-based genomic assay to detect pathogens or single-nucleotide polymorphisms in human genomic samples.

    Directory of Open Access Journals (Sweden)

    Jasmine I Daksis

    Full Text Available Human genomic samples are complex and are considered difficult to assay directly without denaturation or PCR amplification. We report the use of a base-specific heteropolymeric triplex, formed by native duplex genomic target and an oligonucleotide third strand probe, to assay for low copy pathogen genomes present in a sample also containing human genomic duplex DNA, or to assay human genomic duplex DNA for Single Nucleotide Polymorphisms (SNP, without PCR amplification. Wild-type and mutant probes are used to identify triplexes containing FVL G1691A, MTHFR C677T and CFTR mutations. The specific triplex structure forms rapidly at room temperature in solution and may be detected without a separation step. YOYO-1, a fluorescent bis-intercalator, promotes and signals the formation of the specific triplex. Genomic duplexes may be assayed homogeneously with single base pair resolution. The specific triple-stranded structures of the assay may approximate homologous recombination intermediates, which various models suggest may form in either the major or minor groove of the duplex. The bases of the stable duplex target are rendered specifically reactive to the bases of the probe because of the activity of intercalated YOYO-1, which is known to decondense duplex locally 1.3 fold. This may approximate the local decondensation effected by recombination proteins such as RecA in vivo. Our assay, while involving triplex formation, is sui generis, as it is not homopurine sequence-dependent, as are "canonical triplexes". Rather, the base pair-specific heteropolymeric triplex of the assay is conformation-dependent. The highly sensitive diagnostic assay we present allows for the direct detection of base sequence in genomic duplex samples, including those containing human genomic duplex DNA, thereby bypassing the inherent problems and cost associated with conventional PCR based diagnostic assays.

  1. A novel integrated strategy for detection of human bocavirus based on a heminested PCR assay combined with boiling lysis method of samples in human specimens.

    Science.gov (United States)

    Chen, Long; Yao, Qing; Ma, Jing; Li, Jianning; Zhang, Qian; Yang, Yi; Li, Fang; Sun, Yuning

    2014-07-01

    Human bocavirus (HBoV) has been shown to be associated with acute respiratory tract infection in children. The aim of the work was to develop a novel integrated strategy for human bocavirus detection: heminested PCR assay combined with boiling lysis method of samples. The detection limit of the heminested PCR assay was 1.2 copies of a recombinant DNA plasmid, and no cross-reaction with other respiratory viruses or bacteria was observed. By using the integrated strategy, a total of 202 secretions of the lower respiratory tract of children with acute respiratory diseases were collected and tested. The samples were treated and lysed in boiling lysis buffer rather than extracting viral DNA from secretions, then these sample lysates could be templates and tested by heminested PCR assay, and the amplification of HBoV DNA was detected by using agarose gel electrophoresis. The results showed that, only 7 samples were found to be positive by conventional single-round PCR; importantly, the other new 41 samples were positive by heminested PCR assay. Additionally, the genomic viral DNA was extracted from all positive and some negative specimens, amplified, and sequenced. The results were perfectly consistent with those of the integrated strategy. Taken together, these results suggest that the novel integrated strategy (heminested PCR assay combined with boiling lysis method of samples) is a convenient, sensitive, cost-effective and reliable detective method for HBoV detection and will have broad application prospects in clinical diagnosis. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Distribution of human waste samples in relation to sizing waste processing in space

    Science.gov (United States)

    Parker, Dick; Gallagher, S. K.

    1992-01-01

    Human waste processing for closed ecological life support systems (CELSS) in space requires that there be an accurate knowledge of the quantity of wastes produced. Because initial CELSS will be handling relatively few individuals, it is important to know the variation that exists in the production of wastes rather than relying upon mean values that could result in undersizing equipment for a specific crew. On the other hand, because of the costs of orbiting equipment, it is important to design the equipment with a minimum of excess capacity because of the weight that extra capacity represents. A considerable quantity of information that had been independently gathered on waste production was examined in order to obtain estimates of equipment sizing requirements for handling waste loads from crews of 2 to 20 individuals. The recommended design for a crew of 8 should hold 34.5 liters per day (4315 ml/person/day) for urine and stool water and a little more than 1.25 kg per day (154 g/person/day) of human waste solids and sanitary supplies.

  3. Partial Sequence Analysis of the Genome of Human Herpesvirus 7 YY5 Isolated from Saliva Samples

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective To isolate and identify Nanjing local strains of Human Herpesvirus 7 (HH-V-7), and to analyze their partia l genome characteristic. Methods The saliva specimens were collected from 2 healthy adults and 5 children with kidney disease. After treatment with antibiotics and filtering. they were inoculated on to the phytohemagglutin stimulated umbilical cord blood mononuclear cells ( CBMCs). When the infected cells presented the typical ballooning and polykaryotic cytopathic effects (CPE), we identified them by transvnission electron microscopy and polymerase chain reaction.PCR product was also sequenced. Results Four strains were isolated from the seven saliva specimens. The 186-base-pair fragment of the isolated strain YY5 PCR products was sequenced, which encoded part of the HHV-7 U10 gene. The DNA sequence revealed an identity of 57. 5% and 36.0%, respectively with HHV-6 and human cytomegalovirus ( HCMV). At the amino acid level, the similarity of 51.6% was found between HHV-7 and HHV-6, and that of 25.8% between HHV-7and HCMV. Conclusion The isolated viruses were HHV-7, and 186 bp fragments revealed an identity with HHV-7 RK and Jl of 100%.

  4. Acetaminophen and Meloxicam Inhibit Platelet Aggregation and Coagulation in Blood Samples from Humans

    Science.gov (United States)

    2014-01-01

    used for over 50 years to relieve pains associated with arthritis , headache, mus- cular aches, and to reduce fever in adults and children. As in most...participant was sampled once with a total of 100-ml blood volume. Exclusion criteria included pregnancy , on- going therapeutic anticoagulation, and use... rheumatic disease. Clin Ther 2000; 22:400–410; S0149-2918(00)89009-8 [pii] 10.1016/ S0149-2918(00)89009-8. 5 Hawkey C, Kahan A, Steinbruck K, Alegre C

  5. Metagenomic Analysis of Antibiotic Resistance Genes in Dairy Cow Feces following Therapeutic Administration of Third Generation Cephalosporin.

    Directory of Open Access Journals (Sweden)

    Lindsey Chambers

    Full Text Available Although dairy manure is widely applied to land, it is relatively understudied compared to other livestock as a potential source of antibiotic resistance genes (ARGs to the environment and ultimately to human pathogens. Ceftiofur, the most widely used antibiotic used in U.S. dairy cows, is a 3rd generation cephalosporin, a critically important class of antibiotics to human health. The objective of this study was to evaluate the effect of typical ceftiofur antibiotic treatment on the prevalence of ARGs in the fecal microbiome of dairy cows using a metagenomics approach. β-lactam ARGs were found to be elevated in feces from Holstein cows administered ceftiofur (n = 3 relative to control cows (n = 3. However, total numbers of ARGs across all classes were not measurably affected by ceftiofur treatment, likely because of dominance of unaffected tetracycline ARGs in the metagenomics libraries. Functional analysis via MG-RAST further revealed that ceftiofur treatment resulted in increases in gene sequences associated with "phages, prophages, transposable elements, and plasmids", suggesting that this treatment also enriched the ability to horizontally transfer ARGs. Additional functional shifts were noted with ceftiofur treatment (e.g., increase in genes associated with stress, chemotaxis, and resistance to toxic compounds; decrease in genes associated with metabolism of aromatic compounds and cell division and cell cycle, along with measureable taxonomic shifts (increase in Bacterioidia and decrease in Actinobacteria. This study demonstrates that ceftiofur has a broad, measureable and immediate effect on the cow fecal metagenome. Given the importance of 3rd generation cephalospirins to human medicine, their continued use in dairy cattle should be carefully considered and waste treatment strategies to slow ARG dissemination from dairy cattle manure should be explored.

  6. Mercury in human hair and blood samples from people living in Wanshan mercury mine area, Guizhou, China: an XAS study.

    Science.gov (United States)

    Li, Yu-Feng; Chen, Chunying; Li, Bai; Li, Wei; Qu, Liya; Dong, Zeqin; Nomura, Masaharu; Gao, Yuxi; Zhao, Jinxuan; Hu, Wei; Zhao, Yuliang; Chai, Zhifang

    2008-03-01

    Human hair and blood samples from persons living in the town of Wanshan, a mercury mine area in Guizhou Province of China, were collected and the quantitative speciation and structural information of Hg and S in hair samples and of Hg in erythrocyte and serum samples were studied using X-ray absorption spectroscopy. Least-squares fitting of the X-ray absorption near-edge spectra found that inorganic mercury is the major mercury species in hair samples (91.74%), while inorganic and methyl mercury are both about 50% of total mercury in RBC and serum samples, which is in agreement with the data obtained by acidic extraction, fractionation of Hg(2+) and CH(3)Hg(+) and quantification by ICP-MS. Curve-fitting analysis revealed that the Hg-S bond length and coordination number in hair were 0.248+/-0.002 nm and 3.10, respectively, while the S-Hg bond length and coordination number in hair were 0.236+/-0.002 nm and 4.05. The Hg-S bond length and coordination number in RBC were 0.251+/-0.003 nm and 4.09, respectively, while they were 0.228+/-0.002 nm and 4.08 in serum, respectively. The techniques for speciation, structural and binding information described in this study will find the potential application in similar studies of other elements.

  7. Epo receptors are not detectable in primary human tumor tissue samples.

    Directory of Open Access Journals (Sweden)

    Steve Elliott

    Full Text Available Erythropoietin (Epo is a cytokine that binds and activates an Epo receptor (EpoR expressed on the surface of erythroid progenitor cells to promote erythropoiesis. While early studies suggested EpoR transcripts were expressed exclusively in the erythroid compartment, low-level EpoR transcripts were detected in nonhematopoietic tissues and tumor cell lines using sensitive RT-PCR methods. However due to the widespread use of nonspecific anti-EpoR antibodies there are conflicting data on EpoR protein expression. In tumor cell lines and normal human tissues examined with a specific and sensitive monoclonal antibody to human EpoR (A82, little/no EpoR protein was detected and it was not functional. In contrast, EpoR protein was reportedly detectable in a breast tumor cell line (MCF-7 and breast cancer tissues with an anti-EpoR polyclonal antibody (M-20, and functional responses to rHuEpo were reported with MCF-7 cells. In another study, a functional response was reported with the lung tumor cell line (NCI-H838 at physiological levels of rHuEpo. However, the specificity of M-20 is in question and the absence of appropriate negative controls raise questions about possible false-positive effects. Here we show that with A82, no EpoR protein was detectable in normal human and matching cancer tissues from breast, lung, colon, ovary and skin with little/no EpoR in MCF-7 and most other breast and lung tumor cell lines. We show further that M-20 provides false positive staining with tissues and it binds to a non-EpoR protein that migrates at the same size as EpoR with MCF-7 lysates. EpoR protein was detectable with NCI-H838 cells, but no rHuEpo-induced phosphorylation of AKT, STAT3, pS6RP or STAT5 was observed suggesting the EpoR was not functional. Taken together these results raise questions about the hypothesis that most tumors express high levels of functional EpoR protein.

  8. HPLC determination of ciprofloxacin, cloxacillin, and ibuprofen drugs in human urine samples.

    Science.gov (United States)

    Espinosa-Mansilla, Anunciación; Muñoz de la Peña, Arsenio; González Gómez, David; Cañada-Cañada, Florentina

    2006-08-01

    This paper reports, for the first time, a liquid chromatographic method for the simultaneous determination of three frequently co-administered active principles, two antibiotics, ciprofloxacin (CIPRO) and cloxacillin (CLOXA) belonging to the fluoroquinolones and beta-lactam families, respectively, and ibuprofen (IBU), a non-steroidal anti-inflammatory drug. The chromatographic separation was performed on a C-18 analytical column, using isocratic elution with methanol-acetonitrile-pH 3 formate buffer (CT = 0.1 M) (15:12:73, v/v/v) for 3 min and, after that, a linear gradient with methanol-acetonitrile (88:12, v/v) for 8 min. Several absorption spectra were obtained for each peak using a DAD detector. Chromatograms at the maximum absorption wavelength for each analyte, 220 nm for both IBU and CLOXA, and 280 nm for CIPRO were selected as the most suitable. The proposed chromatographic method requires about 15 min per sample. The presence of a urine background was tested and no interference was found. The method was satisfactorily applied to the determination of CIPRO, CLOXA, and IBU, in fortified urine, and in urine samples from a patient undergoing treatment with these three active principles, among others. Limits of quantification in urine were 1.00, 1.70, and 2.87 microg/mL for CIPRO, CLOXA, and IBU, respectively.

  9. Gay and Bisexual Men's Perceptions of the Donation and Use of Human Biological Samples for Research: A Qualitative Study.

    Directory of Open Access Journals (Sweden)

    Chris Patterson

    Full Text Available Human biological samples (biosamples are increasingly important in diagnosing, treating and measuring the prevalence of illnesses. For the gay and bisexual population, biosample research is particularly important for measuring the prevalence of human immunodeficiency virus (HIV. By determining people's understandings of, and attitudes towards, the donation and use of biosamples, researchers can design studies to maximise acceptability and participation. In this study we examine gay and bisexual men's attitudes towards donating biosamples for HIV research. Semi-structured telephone interviews were conducted with 46 gay and bisexual men aged between 18 and 63 recruited in commercial gay scene venues in two Scottish cities. Interview transcripts were analysed thematically using the framework approach. Most men interviewed seemed to have given little prior consideration to the issues. Participants were largely supportive of donating tissue for medical research purposes, and often favourable towards samples being stored, reused and shared. Support was often conditional, with common concerns related to: informed consent; the protection of anonymity and confidentiality; the right to withdraw from research; and ownership of samples. Many participants were in favour of the storage and reuse of samples, but expressed concerns related to data security and potential misuse of samples, particularly by commercial organisations. The sensitivity of tissue collection varied between tissue types and collection contexts. Blood, urine, semen and bowel tissue were commonly identified as sensitive, and donating saliva and as unlikely to cause discomfort. To our knowledge, this is the first in-depth study of gay and bisexual men's attitudes towards donating biosamples for HIV research. While most men in this study were supportive of donating tissue for research, some clear areas of concern were identified. We suggest that these minority concerns should be accounted

  10. A new treatment by dispersive liquid-liquid microextraction for the determination of parabens in human serum samples.

    Science.gov (United States)

    Vela-Soria, F; Ballesteros, O; Rodríguez, I; Zafra-Gómez, A; Ballesteros, L; Cela, R; Navalón, A

    2013-09-01

    Alkyl esters of p-hydroxybenzoic acid (parabens) are a family of compounds that have been in use since the 1920s as preservatives in cosmetic formulations, with one of the lowest rates of skin problems reported in dermatological patients. However, in the last few years, many scientific publications have demonstrated that parabens are weak endocrine disruptors, meaning that they can interfere with the function of endogenous hormones, increasing the risk of breast cancer. In the present work, a new sample treatment method is introduced based on dispersive liquid-liquid microextraction for the extraction of the most commonly used parabens (methyl-, ethyl-, propyl-, and butylparaben) from human serum samples followed by separation and quantification using ultrahigh performance liquid chromatography-tandem mass spectrometry. The method involves an enzymatic treatment to quantify the total content of parabens. The extraction parameters (solvent and disperser solvent, extractant and dispersant volume, pH of the sample, salt addition, and extraction time) were accurately optimized using multivariate optimization strategies. Ethylparaben ring (13)C6-labeled was used as surrogate. Limits of quantification ranging from 0.2 to 0.7 ng mL(-1) and an interday variability (evaluated as relative standard deviations) from 3.8 to 11.9 % were obtained. The method was validated using matrix-matched calibration standard and a spike recovery assay. Recovery rates for spiked samples ranged from 96 to 106 %, and a good linearity up to concentrations of 100 ng mL(-1) was obtained. The method was satisfactorily applied for the determination of target compounds in human serum samples.

  11. Molecular identification of nocardia isolates from clinical samples and an overview of human nocardiosis in Brazil.

    Directory of Open Access Journals (Sweden)

    Paulo Victor Pereira Baio

    Full Text Available BACKGROUND: Nocardia sp. causes a variety of clinical presentations. The incidence of nocardiosis varies geographically according to several factors, such as the prevalence of HIV infections, transplants, neoplastic and rheumatic diseases, as well as climate, socio-economic conditions and laboratory procedures for Nocardia detection and identification. In Brazil the paucity of clinical reports of Nocardia infections suggests that this genus may be underestimated as a cause of human diseases and/or either neglected or misidentified in laboratory specimens. Accurate identification of Nocardia species has become increasingly important for clinical and epidemiological investigations. In this study, seven clinical Nocardia isolates were identified by multilocus sequence analysis (MLSA and their antimicrobial susceptibility was also determined. Most Nocardia isolates were associated to pulmonary disease. METHODOLOGY/PRINCIPAL FINDINGS: The majority of Brazilian human isolates in cases reported in literature were identified as Nocardia sp. Molecular characterization was used for species identification of Nocardia nova, Nocardia cyriacigeorgica, Nocardia asiatica and Nocardia exalbida/gamkensis. Data indicated that molecular analysis provided a different Nocardia speciation than the initial biochemical identification for most Brazilian isolates. All Nocardia isolates showed susceptibility to trimethoprim-sulfamethoxazole, the antimicrobial of choice in the treatment nocardiosis. N. nova isolated from different clinical specimens from one patient showed identical antimicrobial susceptibility patterns and two distinct clones. CONCLUSIONS/SIGNIFICANCE: Although Brazil is the world's fifth-largest country in terms of land mass and population, pulmonary, extrapulmonary and systemic forms of nocardiosis were reported in only 6 of the 26 Brazilian states from 1970 to 2013. A least 33.8% of these 46 cases of nocardiosis proved fatal. Interestingly, coinfection

  12. Assessing employability capacities and career adaptability in a sample of human resource professionals

    Directory of Open Access Journals (Sweden)

    Melinde Coetzee

    2015-03-01

    Full Text Available Orientation: Employers have come to recognise graduates’ employability capacities and their ability to adapt to new work demands as important human capital resources for sustaining a competitive business advantage.Research purpose: The study sought (1 to ascertain whether a significant relationship exists between a set of graduate employability capacities and a set of career adaptability capacities and (2 to identify the variables that contributed the most to this relationship.Motivation for the study: Global competitive markets and technological advances are increasingly driving the demand for graduate knowledge and skills in a wide variety of jobs. Contemporary career theory further emphasises career adaptability across the lifespan as a critical skill for career management agency. Despite the apparent importance attached to employees’ employability and career adaptability, there seems to be a general lack of research investigating the association between these constructs.Research approach, design and method: A cross-sectional, quantitative research design approach was followed. Descriptive statistics, Pearson product-moment correlations and canonical correlation analysis were performed to achieve the objective of the study. The participants (N = 196 were employed in professional positions in the human resource field and were predominantly early career black people and women.Main findings: The results indicated positive multivariate relationships between the variables and showed that lifelong learning capacities and problem solving, decision-making and interactive skills contributed the most to explaining the participants’ career confidence, career curiosity and career control.Practical/managerial implications: The study suggests that developing professional graduates’ employability capacities may strengthen their career adaptability. These capacities were shown to explain graduates’ active engagement in career management strategies

  13. Sampling rate dependence of correlation at long time lags in BOLD fMRI measurements on humans and gel phantoms.

    Science.gov (United States)

    Mikkelsen, Kaare B; Lund, Torben E

    2013-01-01

    The aim of this study is to investigate the effects of sampling rate on Hurst exponents derived from Blood Oxygenation Level Dependent functional Magnetic Resonance Imaging (BOLD fMRI) resting state time series. fMRI measurements were performed on 2 human subjects and a selection of gel phantoms. From these, Hurst exponents were calculated. It was found that low sampling rates induced non-trivial exponents at sharp material transitions, and that Hurst exponents of human measurements had a strong TR-dependence. The findings are compared to theoretical considerations regarding the fractional Gaussian noise model and resampling, and it is found that the implications are problematic. This result should have a direct influence on the way future studies of low-frequency variation in BOLD fMRI data are conducted, especially if the fractional Gaussian noise model is considered. We recommend either using a different model (examples of such are referenced in the conclusion), or standardizing experimental procedures along an optimal sampling rate.

  14. Rapid milk group classification by 1H NMR analysis of Le and H epitopes in human milk oligosaccharide donor samples.

    Science.gov (United States)

    van Leeuwen, Sander S; Schoemaker, Ruud J W; Gerwig, Gerrit J; van Leusen-van Kan, Ellen J M; Dijkhuizen, Lubbert; Kamerling, Johannis P

    2014-08-01

    Human milk oligosaccharides (HMOs) are a major constituent of human breast milk and play an important role in reducing the risk of infections in infants. The structures of these HMOs show similarities with blood group antigens in protein glycosylation, in particular in relation to fucosylation in Lewis blood group-type epitopes, matching the maternal pattern. Previously, based on the Secretor and Lewis blood group system, four milk groups have been defined, i.e. Lewis-positive Secretors, Lewis-positive non-Secretors, Lewis-negative Secretors and Lewis-negative non-Secretors. Here, a rapid one-dimensional (1)H nuclear magnetic resonance (NMR) analysis method is presented that identifies the presence/absence of (α1-2)-, (α1-3)- and (α1-4)-linked fucose residues in HMO samples, affording the essential information to attribute different HMO samples to a specific milk group. The developed method is based on the NMR structural-reporter-group concept earlier established for glycoprotein glycans. Further evaluation of the data obtained from the analysis of 36 HMO samples shows that within each of the four milk groups the relative levels of the different fucosylation epitopes can greatly vary. The data also allow a separation of the Lewis-positive Secretor milk group into two sub-groups.

  15. Genotypic Characterization of Toxoplasma gondii Strains Associated with Human Toxoplasmosis in Spain: Direct Analysis from Clinical Samples

    Science.gov (United States)

    Fuentes, Isabel; Rubio, Jose M.; Ramírez, Carmen; Alvar, Jorge

    2001-01-01

    Genetic analysis of the SAG2 locus was performed to determine the prevalence of the different genotypes of Toxoplasma gondii (strain types I, II, and III) associated with human toxoplasmosis in Spain. This determination was made directly from primary clinical samples, obviating the previous process of isolation in mice or cell culture. A total of 34 isolates of T. gondii, collected from immunocompromised patients and congenital infection cases, were analyzed. Restriction fragment length polymorphism in PCR-amplified SAG2 products was used to group strains into one of the three genotypes of T. gondii. Complete characterization of the SAG2 gene was successful in 76.5% of the cases, demonstrating the feasibility of direct genotype analysis from clinical samples of different origins. Strains of T. gondii type II were the most prevalent in immunocompromised patients, with 52% of cases, while strains of type I were present in 75% of the congenital infection cases. These data differ from previous reports that show type II strains to be mostly associated with all kinds of human toxoplasmosis. These differences might be an effect of selection in the process of culture and isolation of the samples performed by other researchers prior to strain characterization. PMID:11283088

  16. Electrochemical genosensor array for the simultaneous detection of multiple high-risk human papillomavirus sequences in clinical samples

    Energy Technology Data Exchange (ETDEWEB)

    Civit, Laia [Nanobiotechnology and Bioanalysis Group, Departament d' Enginyeria Quimica, Universitat Rovira i Virgili, 43007 Tarragona (Spain); Fragoso, Alex, E-mail: alex.fragoso@urv.cat [Nanobiotechnology and Bioanalysis Group, Departament d' Enginyeria Quimica, Universitat Rovira i Virgili, 43007 Tarragona (Spain); Hoelters, Sebastian; Duerst, Matthias [Department for Gynecology, Jena University Hospital, Friedrich-Schiller-University Jena, D-07743 Jena (Germany); O' Sullivan, Ciara K., E-mail: ciara.osullivan@urv.cat [Nanobiotechnology and Bioanalysis Group, Departament d' Enginyeria Quimica, Universitat Rovira i Virgili, 43007 Tarragona (Spain); Institucio Catalana de Recerca i Estudis Avancats, Passeig Lluis Companys 23, 08010 Barcelona (Spain)

    2012-02-17

    Highlights: Black-Right-Pointing-Pointer High-risk human papillomavirus is detected in virtually all-invasive cervical cancers. Black-Right-Pointing-Pointer Electrochemical genosensor for simultaneous detection of multiple high-risk HPV applied to cervical scrape samples. Black-Right-Pointing-Pointer Excellent correlation with HPV genotyping carried out within a hospital laboratory. - Abstract: An electrochemical genosensor array for the simultaneous detection of three high-risk human papillomavirus (HPV) DNA sequences, HPV16, 18 and 45, exhibiting high sensitivity and selectivity is presented. The electrodes of a 4 Multiplication-Sign 4 array were modified via co-immobilization of a 1:100 (mol/mol) mixture of a thiolated probe and an oligoethyleneglycol-terminated bipodal thiol. Detection of synthetic and PCR products was carried out in a sandwich type format, with the target hybridized between a surface immobilized probe and a horseradish peroxidase-labelled secondary reporter probe. The detection limits obtained in the detection of each individual target were in the pM range, allowing the application of this sensor for the detection of samples obtained from PCR amplification of cervical scrape samples. The results obtained exhibited an excellent correlation with the HPV genotyping carried out within a hospital laboratory. Multiplexing and cross-reactivity studies demonstrated high selectivity over potential interfering sequences, facilitating application of the developed platform for the high-throughput screening of multiple high-risk DNA sequences.

  17. Development and evaluation of a novel multicopy-element-targeting triplex PCR for detection of Mycobacterium avium subsp. paratuberculosis in feces.

    Science.gov (United States)

    Sevilla, Iker A; Garrido, Joseba M; Molina, Elena; Geijo, María V; Elguezabal, Natalia; Vázquez, Patricia; Juste, Ramón A

    2014-06-01

    The enteropathy called paratuberculosis (PTB), which mainly affects ruminants and has a worldwide distribution, is caused by Mycobacterium avium subsp. paratuberculosis. This disease significantly reduces the cost-effectiveness of ruminant farms, and therefore, reliable and rapid detection methods are needed to control the spread of the bacterium in livestock and in the environment. The aim of this study was to identify a specific and sensitive combination of DNA extraction and amplification to detect M. avium subsp. paratuberculosis in feces. Negative bovine fecal samples were inoculated with increasing concentrations of two different bacterial strains (field and reference) to compare the performance of four extraction and five amplification protocols. The best results were obtained using the JohnePrep and MagMax extraction kits combined with an in-house triplex real-time PCR designed to detect IS900, ISMap02 (an insertion sequence of M. avium subsp. paratuberculosis present in 6 copies per genome), and an internal amplification control DNA simultaneously. These combinations detected 10 M. avium subsp. paratuberculosis cells/g of spiked feces. The triplex PCR detected 1 fg of genomic DNA extracted from the reference strain K10. The performance of the robotized version of the MagMax extraction kit combined with the IS900 and ISMap02 PCR was further evaluated using 615 archival fecal samples from the first sampling of nine Friesian cattle herds included in a PTB control program and followed up for at least 4 years. The analysis of the results obtained in this survey demonstrated that the diagnostic method was highly specific and sensitive for the detection of M. avium subsp. paratuberculosis in fecal samples from cattle and a very valuable tool to be used in PTB control programs. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  18. PCR Inhibition of a Quantitative PCR for Detection of Mycobacterium avium Subspecies Paratuberculosis DNA in Feces: Diagnostic Implications and Potential Solutions.

    Science.gov (United States)

    Acharya, Kamal R; Dhand, Navneet K; Whittington, Richard J; Plain, Karren M

    2017-01-01

    Molecular tests such as polymerase chain reaction (PCR) are increasingly being applied for the diagnosis of Johne's disease, a chronic intestinal infection of ruminants caused by Mycobacterium avium subspecies paratuberculosis (MAP). Feces, as the primary test sample, presents challenges in terms of effective DNA isolation, with potential for PCR inhibition and ultimately for reduced analytical and diagnostic sensitivity. However, limited evidence is available regarding the magnitude and diagnostic implications of PCR inhibition for the detection of MAP in feces. This study aimed to investigate the presence and diagnostic implications of PCR inhibition in a quantitative PCR assay for MAP (High-throughput Johne's test) to investigate the characteristics of samples prone to inhibition and to identify measures that can be taken to overcome this. In a study of fecal samples derived from a high prevalence, endemically infected cattle herd, 19.94% of fecal DNA extracts showed some evidence of inhibition. Relief of inhibition by a five-fold dilution of the DNA extract led to an average increase in quantification of DNA by 3.3-fold that consequently increased test sensitivity of the qPCR from 55 to 80% compared to fecal culture. DNA extracts with higher DNA and protein content had 19.33 and 10.94 times higher odds of showing inhibition, respectively. The results suggest that the current test protocol is sensitive for herd level diagnosis of Johne's disease but that test sensitivity and individual level diagnosis could be enhanced by relief of PCR inhibition, achieved by five-fold dilution of the DNA extract. Furthermore, qualitative and quantitative parameters derived from absorbance measures of DNA extracts could be useful for prediction of inhibitory fecal samples.

  19. PCR Inhibition of a Quantitative PCR for Detection of Mycobacterium avium Subspecies Paratuberculosis DNA in Feces: Diagnostic Implications and Potential Solutions

    Science.gov (United States)

    Acharya, Kamal R.; Dhand, Navneet K.; Whittington, Richard J.; Plain, Karren M.

    2017-01-01

    Molecular tests such as polymerase chain reaction (PCR) are increasingly being applied for the diagnosis of Johne’s disease, a chronic intestinal infection of ruminants caused by Mycobacterium avium subspecies paratuberculosis (MAP). Feces, as the primary test sample, presents challenges in terms of effective DNA isolation, with potential for PCR inhibition and ultimately for reduced analytical and diagnostic sensitivity. However, limited evidence is available regarding the magnitude and diagnostic implications of PCR inhibition for the detection of MAP in feces. This study aimed to investigate the presence and diagnostic implications of PCR inhibition in a quantitative PCR assay for MAP (High-throughput Johne’s test) to investigate the characteristics of samples prone to inhibition and to identify measures that can be taken to overcome this. In a study of fecal samples derived from a high prevalence, endemically infected cattle herd, 19.94% of fecal DNA extracts showed some evidence of inhibition. Relief of inhibition by a five-fold dilution of the DNA extract led to an average increase in quantification of DNA by 3.3-fold that consequently increased test sensitivity of the qPCR from 55 to 80% compared to fecal culture. DNA extracts with higher DNA and protein content had 19.33 and 10.94 times higher odds of showing inhibition, respectively. The results suggest that the current test protocol is sensitive for herd level diagnosis of Johne’s disease but that test sensitivity and individual level diagnosis could be enhanced by relief of PCR inhibition, achieved by five-fold dilution of the DNA extract. Furthermore, qualitative and quantitative parameters derived from absorbance measures of DNA extracts could be useful for prediction of inhibitory fecal samples. PMID:28210245

  20. Psychrophilic dry anaerobic digestion of dairy cow feces: Long-term operation

    Energy Technology Data Exchange (ETDEWEB)

    Massé, Daniel I., E-mail: Daniel.masse@agr.gc.ca; Cata Saady, Noori M.

    2015-02-15

    Highlights: • Psychrophilic dry anaerobic digestion (PDAD) of cow feces (CF) is feasible. • PDAD of CF is as efficient as mesophilic and thermophilic AD at TCL 21 days. • CF (13–16% TS at OLR 5.0 g TCOD{sub fed} kg{sup −1} inoculum d{sup −1}) yielded 222 ± 27 {sub N}L CH{sub 4} kg{sup −1} VS fed. - Abstract: This paper reports experimental results which demonstrate psychrophilic dry anaerobic digestion of cow feces during long-term operation in sequence batch reactor. Cow feces (13–16% total solids) has been anaerobically digested in 12 successive cycles (252 days) at 21 days treatment cycle length (TCL) and temperature of 20 °C using psychrotrophic anaerobic mixed culture. An average specific methane yield (SMY) of 184.9 ± 24.0, 189.9 ± 27.3, and 222 ± 27.7 {sub N}L CH{sub 4} kg{sup −1} of VS fed has been achieved at an organic loading rate of 3.0, 4.0, and 5.0 g TCOD kg{sup −1} inoculum d{sup −1} and TCL of 21 days, respectively. The corresponding substrate to inoculum ratio (SIR) was 0.39 ± 0.06, 0.48 ± .02, 0.53 ± 0.05, respectively. Average methane production rate of 10 ± 1.4 {sub N}L CH{sub 4} kg{sup −1} VS fed d{sup −1} has been obtained. The low concentration of volatile fatty acids indicated that hydrolysis was the reaction limiting step.

  1. Metabolic and pharmacokinetic studies of scutellarin in rat plasma, urine, and feces

    Institute of Scientific and Technical Information of China (English)

    Jian-feng XING; Hai-sheng YOU; Ya-lin DONG; Jun LU; Si-ying CHEN; Hui-fang ZHU; Qian DONG; Mao-yi WANG; Wei-hua DONG

    2011-01-01

    Aim: To study the metabolic and pharmacokinetic profile of scutellarin, an active component from the medical plant Erigeron brevis-capus (Vant) Hand-Mazz, and to investigate the mechanisms underlying the low bioavailability of scutellarin though oral or intravenous administration in rats.Methods: HPLC method was developed for simultaneous detection of scutellarin and scutellarein (the aglycone of scutellarin) in rat plasma, urine and feces. The in vitro metabolic stability study was carried out in rat liver microsomes from different genders. Results. After a single oral dose of scutellarin (400 mg/kg), the plasma concentrations of scutellarin and scutellarein in female rats were significantly higher than in male ones. Between the female and male rats, significant differences in AUC, t and C for scutel-larin were found. The pharmacokinetic parameters of scutellarin in the urine also showed significant gender differences. After a single oral dose of scutellarin (400 mg/kg), the total percentage excretion of scutellarein in male and female rats was 16.5% and 8.61%, respectively. The total percentage excretion of scutellarin and scutellarein in the feces was higher with oral administration than with intravenous administration. The in vitro t and CL value for scuteliarin in male rats was significantly higher than that in female rats.Conclusion: The results suggest that a large amount of ingested scutellarin was metabolized into scutellarein in the gastrointestinal tract and then excreted with the feces, leading to the extremely low oral bioavailability of scutellarin. The gender differences of pharma-cokinetic parameters of scutellarin and scutallarein are due to the higher CL and lower absorption in male rats.

  2. Significance of Coronavirus Mutants in Feces and Diseased Tissues of Cats Suffering from Feline Infectious Peritonitis

    Directory of Open Access Journals (Sweden)

    Niels C. Pedersen

    2009-08-01

    Full Text Available The internal FECV→FIPV mutation theory and three of its correlates were tested in four sibs/half-sib kittens, a healthy contact cat, and in four unrelated cats that died of FIP at geographically disparate regions. Coronavirus from feces and extraintestinal FIP lesions from the same cat were always >99% related in accessory and structural gene sequences. SNPs and deletions causing a truncation of the 3c gene product were found in almost all isolates from the diseased tissues of the eight cats suffering from FIP, whereas most, but not all fecal isolates from these same cats had intact 3c genes. Other accessory and structural genes appeared normal in both fecal and lesional viruses. Deliterious mutations in the 3c gene were unique to each cat, indicating that they did not originate in one cat and were subsequently passed horizontally to the others. Compartmentalization of the parental and mutant forms was not absolute; virus of lesional type was sometimes found in feces of affected cats and virus identical to fecal type was occasionally identified in diseased tissues. Although 3c gene mutants in this study were not horizontally transmitted, the parental fecal virus was readily transmitted by contact from a cat that died of FIP to its housemate. There was a high rate of mutability in all structural and accessory genes both within and between cats, leading to minor genetic variants. More than one variant could be identified in both diseased tissues and feces of the same cat. Laboratory cats inoculated with a mixture of two closely related variants from the same FIP cat developed disease from one or the other variant, but not both. Significant genetic drift existed between isolates from geographically distinct regions of the Western US.

  3. Sesavirus: prototype of a new parvovirus genus in feces of a sea lion.

    Science.gov (United States)

    Phan, Tung Gia; Gulland, Frances; Simeone, Claire; Deng, Xutao; Delwart, Eric

    2015-02-01

    We describe the nearly complete genome of a highly divergent parvovirus, we tentatively name Sesavirus, from the feces of a California sea lion pup (Zalophus californianus) suffering from malnutrition and pneumonia. The 5,049-base-long genome contained two major ORFs encoding a 553-aa nonstructural protein and a 965-aa structural protein which shared closest amino acid identities of 25 and 28 %, respectively, with members of the copiparvovirus genus known to infect pigs and cows. Given the low degree of similarity, Sesavirus might be considered as prototype for a new genus with a proposed name of Marinoparvovirus in the subfamily Parvovirinae.

  4. Methodology for the analysis of fenbendazole and its metabolites in plasma, urine, feces, and tissue homogenates.

    Science.gov (United States)

    Barker, S A; Hsieh, L C; Short, C R

    1986-05-15

    New methodology for the extraction and analysis of the anthelmintic fenbendazole and its metabolites from plasma, urine, liver homogenates, and feces from several animal species is presented. Quantitation of fenbendazole and its metabolites was conducted by high-pressure liquid chromatography using ultraviolet detection at 290 nm. The combined extraction and analysis procedures give excellent recoveries in all of the different biological matrices examined. High specificity, low limits of detection, and excellent linearity, accuracy, and inter- and intrasample variability were also obtained. The study of fenbendazole pharmacokinetics in vitro and in vivo should be greatly enhanced through the utilization of these methods.

  5. Enumeration and Isolation of cpe-Positive Clostridium perfringens Spores from Feces

    OpenAIRE

    Heikinheimo, A.; Lindström, M.; Korkeala, H

    2004-01-01

    A hydrophobic grid membrane filter-colony hybridization (HGMF-CH) method for the enumeration and isolation of cpe gene-carrying (cpe-positive) Clostridium perfringens spores from feces was developed. A 425-bp DNA probe specific for the cpe gene was sensitive and specific when tested with bacterial DNA and pure cultures. The enumeration of cpe-positive C. perfringens by the HGMF-CH method proved to be as sensitive as nested PCR combined with the most-probable number technique when tested with ...

  6. Quantification of bioavailable chlortetracycline in pig feces using a bacterial whole-cell biosensor

    DEFF Research Database (Denmark)

    Hansen, L. H.; Aarestrup, Frank Møller; Sørensen, S. J.

    2002-01-01

    and maintenance of fecal coliform bacteria resistant to tetracycline. Initially, large quantities of water-extractable CTC were excreted from the pigs and measurable amounts were detected even at 30 days after treatment cessation. This led to a sharp rise in the number of tetracycline resistant coliform bacteria...... in the feces, to within the same order of magnitude as the total coliform count. The high level of tetracycline resistance was maintained in spite of the declining concentration of tetracycline. (C) 2002 Elsevier Science B.V. All rights reserved....

  7. Detection and sequencing of West Nile virus RNA from human urine and serum samples during the 2014 seasonal period.

    Science.gov (United States)

    Nagy, Anna; Bán, Enikő; Nagy, Orsolya; Ferenczi, Emőke; Farkas, Ágnes; Bányai, Krisztián; Farkas, Szilvia; Takács, Mária

    2016-07-01

    West Nile virus, a widely distributed mosquito-borne flavivirus, is responsible for numerous animal and human infections in Europe, Africa and the Americas. In Hungary, the average number of human infections falls between 10 and 20 cases each year. The severity of clinically manifesting infections varies widely from the milder form of West Nile fever to West Nile neuroinvasive disease (WNND). In routine laboratory diagnosis of human West Nile virus infections, serological methods are mainly applied due to the limited duration of viremia. However, recent studies suggest that detection of West Nile virus RNA in urine samples may be useful as a molecular diagnostic test for these infections. The Hungarian National Reference Laboratory for Viral Zoonoses serologically confirmed eleven acute human infections during the 2014 seasonal period. In three patients with neurological symptoms, viral RNA was detected from both urine and serum specimens, albeit for a longer period and in higher copy numbers with urine. Phylogenetic analysis of the NS3 genomic region of three strains and the complete genome of one selected strain demonstrated that all three patients had lineage-2 West Nile virus infections. Our findings reaffirm the utility of viral RNA detection in urine as a molecular diagnostic procedure for diagnosis of West Nile virus infections.

  8. Using CF11 cellulose columns to inexpensively and effectively remove human DNA from Plasmodium falciparum-infected whole blood samples

    Directory of Open Access Journals (Sweden)

    Venkatesan Meera

    2012-02-01

    Full Text Available Abstract Background Genome and transcriptome studies of Plasmodium nucleic acids obtained from parasitized whole blood are greatly improved by depletion of human DNA or enrichment of parasite DNA prior to next-generation sequencing and microarray hybridization. The most effective method currently used is a two-step procedure to deplete leukocytes: centrifugation using density gradient media followed by filtration through expensive, commercially available columns. This method is not easily implemented in field studies that collect hundreds of samples and simultaneously process samples for multiple laboratory analyses. Inexpensive syringes, hand-packed with CF11 cellulose powder, were recently shown to improve ex vivo cultivation of Plasmodium vivax obtained from parasitized whole blood. This study was undertaken to determine whether CF11 columns could be adapted to isolate Plasmodium falciparum DNA from parasitized whole blood and achieve current quantity and purity requirements for Illumina sequencing. Methods The CF11 procedure was compared with the current two-step standard of leukocyte depletion using parasitized red blood cells cultured in vitro and parasitized blood obtained ex vivo from Cambodian patients with malaria. Procedural variations in centrifugation and column size were tested, along with a range of blood volumes and parasite densities. Results CF11 filtration reliably produces 500 nanograms of DNA with less than 50% human DNA contamination, which is comparable to that obtained by the two-step method and falls within the current quality control requirements for Illumina sequencing. In addition, a centrifuge-free version of the CF11 filtration method to isolate P. falciparum DNA at remote and minimally equipped field sites in malaria-endemic areas was validated. Conclusions CF11 filtration is a cost-effective, scalable, one-step approach to remove human DNA from P. falciparum-infected whole blood samples.

  9. Human Biomonitoring of food contaminants in Spanish children: Design, sampling and lessons learned.

    Science.gov (United States)

    Pérez, Rosa; Domenech, Eva; Coscollà, Clara; Yusà, Vicent

    2017-08-12

    Human Biomonitoring (HBM) studies are highly useful for evaluating population exposure to environmental contaminants and are being carried out in increasing numbers all over the world. The use of HBM in the field of food safety, in a risk assessment context, presents a growing interest as more health-based guidance values (HBGV) in biological matrices are derived, and can be used in a complementary way to the external exposure approaches such as total diet studies or surveillance programmes. The aims of the present work are: i) to describe the methodological framework of the BIOVAL study, a cross-sectional HBM program carried out by the Health Department of the Regional Government of Valencia (Spain), that is linked to the food safety official control, and is focused on children from 6 to 11 years of age ii) to explain and discuss the pre-analytical results iii) to report and discuss on lessons learned from its design and implementation. The study population included 666 children from whom urine and hair were taken in order to analyse different biomarkers of exposure to food pollutants. Copyright © 2017 Elsevier GmbH. All rights reserved.

  10. Citrobacter europaeus sp. nov., isolated from water and human faecal samples.

    Science.gov (United States)

    Ribeiro, Teresa G; Clermont, Dominique; Branquinho, Raquel; Machado, Elisabete; Peixe, Luísa; Brisse, Sylvain

    2017-01-01

    Strains 97/79T and A121, recovered respectively from human faeces and well water, were compared to currently known species of the genus Citrobacter using genotypic and phenotypic approaches. Multilocus sequence analysis based on housekeeping genes fusA, leuS, pyrG, rpoB and recN, showed that the two strains formed a distinct phylogenetic lineage within the genus Citrobacter. Average nucleotide identity (ANI) between strains 97/79T and A121 was 99.2 %, whereas ANI values of strain 97/79T with the type strains of closely related species of the genus Citrobacter, C. werkmanii, C. braakii, C. freundii, C. youngae and C. pasteurii, were all below 93.0 %. The ability to metabolize different compounds also discriminated strains 97/79T and A121 from other species of the genus Citrobacter. Based on these results, strains 97/79T and A121 represent a novel species of the genus Citrobacter, for which the name Citrobacter europaeus sp. nov. is proposed, with strain 97/79T (=CIP 106467T=DSM 103031T) as the type strain. The DNA G+C content of strain 97/79T is 52.0 %.

  11. Stable isotope dilution analysis of salicylic acid and hydroquinone in human skin samples by gas chromatography with mass spectrometric detection.

    Science.gov (United States)

    Judefeind, Anja; van Rensburg, Peet Jansen; Langelaar, Stephan; du Plessis, Jeanetta

    2007-06-01

    A sensitive and accurate gas chromatographic-mass spectrometric (GC-MS) method has been developed for the quantitative determination of salicylic acid (SA) and hydroquinone (HQ) from human skin samples and cosmetic emulsions. Deuterium labeled SA-d(6) and HQ-d(6) were used as internal standards (IS). The samples were extracted with methanol, dried under nitrogen and derivatized with N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA)+1% trimethylchlorosilane (TMCS). Quantification was performed in SIM mode with a limit of quantification (LOQ) of 50 ng ml(-1) for SA and 10 ng ml(-1) for HQ. The inter-day variation (R.S.D.) was less than 5% and the accuracy was better than 13.3% for both compounds. The recoveries from the different matrices ranged between 93.1 and 103.3% for SA, and 97.3 and 100.8% for HQ.

  12. Evaluation of middlebrook 7H11 associated with human or sheep blood for the detection of mycobacterium tuberculosis in sputum samples

    OpenAIRE

    Agapito, Juan; Escuela de Tecnología Médica, Facultad de Medicina, Universidad Peruana Cayetano Heredia. Lima, Perú Instituto de Medicina Tropical Alexander von Humboldt, Universidad Peruana Cayetano Heredia. Lima, Perú. microbiólogo.; Cuadros, Luis; Escuela de Tecnología Médica, facultad de Medicina, universidad Peruana Cayetano Heredia. Lima, Perú. Tecnólogo médico.; Tarrillo, Sergio; Escuela de Tecnología Médica, facultad de Medicina, universidad Peruana Cayetano Heredia. Lima, Perú. Tecnólogo médico.; Soto, Alonso; Asociación Latinoamericana de Biotecnología. Lima, Perú. Hospital Nacional Hipólito unanue. Lima, Perú. Facultad de Medicina, universidad Ricardo Palma. Lima, Perú. Médico Internista.

    2009-01-01

    Objective. To evaluate the diagnostic yield of the media Middlebrook 7H11 combined with human or ovine blood in comparison with the Ogawa solid media for the diagnosis of pulmonary tuberculosis. Material and methods. We evaluated sputum samples of patients with clinical suspicion of pulmonary tuberculosis. The samples were seeded in Middlebrook 7H11 agar associated with human or ovine blood and in Ogawa media. Results. A total of 130 samples were collected. The positivity for M.tuberculos...

  13. Concentrations of persistent organic pollutants (POPs) in human blood samples from Mexico City, Mexico.

    Science.gov (United States)

    Orta-García, Sandra; Pérez-Vázquez, Francisco; González-Vega, Carolina; Varela-Silva, José Antonio; Hernández-González, Lidia; Pérez-Maldonado, Iván

    2014-02-15

    Studies in Mexico have demonstrated exposure to persistent organic pollutants (POPs) in people living in different sites through the country. However, studies evaluating exposure to POPs in people living in Mexico City (one of most contaminated places in the world) are scarce. Therefore, the aim of this study was to assess the levels of polybrominated diphenyl ethers (PBDEs), polychlorinated biphenyls (PCBs), dichlorodiphenyltrichloroethane (DDT) and its metabolite dichlorodiphenyldichloroethylene (DDE) in the blood as exposure biomarkers in people living in Mexico City. A total of 123 participants (blood donors aged 20-60 years) were recruited during 2010 in Mexico City. Quantitative analyses of blood samples were performed using gas chromatography coupled with mass spectrometry. Levels of the assessed compounds ranged from non-detectable (

  14. Analysis of chlorpheniramine in human urine samples using dispersive liquid-liquid microextraction combined with high-performance liquid chromatography

    Directory of Open Access Journals (Sweden)

    Mehdi Maham

    2014-09-01

    Full Text Available A simple and environmentally friendly microextraction technique was used for determination of chlorpheniramine (CPM, an antihistamine drug, in human urine samples using dispersive liquid-liquid microextraction (DLLME followed by high performance liquid chromatography with diode array detection (HPLC-DAD. In this extraction technique, an appropriate mixture of acetonitrile (disperser solvent and carbon tetrachloride (extraction solvent was rapidly injected into the urine sample containing the target analyte. Tiny droplets of extractant were formed and dispersed into the sample solution and then sedimented at the bottom of the conical test tube by centrifugation. Under optimal conditions, the calibration curve was linear in the range of 0.055-5.5 µg mL-1, with a detection limit of 16.5 ng mL-1. This proposed method was successfully applied to the analysis of real urine samples. Low consumption of toxic organic solvents, simplicity of operation, low cost and acceptable figures of merit are the main advantages of the proposed technique.

  15. Brominated flame retardants in food and environmental samples from a production area in China: concentrations and human exposure assessment.

    Science.gov (United States)

    Li, Peng; Wu, Hui; Li, Qiuxu; Jin, Jun; Wang, Ying

    2015-11-01

    Human exposure to brominated flame retardants (BFRs: decabromodiphenyl ether (BDE209), decabromodiphenyl ethane (DBDPE), hexabromobenzene (HBB), pentabromoethylbenzene (PBEB), pentabromotoluene (PBT), 1,2,3,4,5-pentabromobenzene (PBBz), and 2,3,5,6-tetrabromo-p-xylene (TBX)) in a brominated flame retardant production area (Weifang, Shandong Province, China) was estimated. Thirty food samples, 14 air samples, and 13 indoor dust samples were analyzed. BDE209 and DBDPE were the dominant BFRs in all samples. Higher alternative brominated flame retardant (including DBDPE, HBB, PBEB, PBT, PBBz, and TBX) concentrations were found in vegetables than in fish and meat; thus, plant-original foods might be important alternative BFR sources in the study area. The BDE209 and alternative BFR concentrations in air were 1.5×10(4) to 2.2×10(5) and 620 to 3.6×10(4) pg/m3, respectively. Mean total BFR exposures through the diet, inhalation, and indoor dust ingestion were 570, 3000, and 69 ng/d, respectively (16, 82, and 2% of total intake, respectively). Inhalation was the dominant BFR source except for DBDPE, for which diet dominated. BDE209 contributed 85% of the total BFR intake in the study area.

  16. Modulatory effects of propolis samples from Latin America (Brazil, Cuba and Mexico) on cytokine production by human monocytes.

    Science.gov (United States)

    Conti, Bruno J; Santiago, Karina B; Búfalo, Michelle C; Herrera, Yahima F; Alday, Efrain; Velazquez, Carlos; Hernandez, Javier; Sforcin, José M

    2015-10-01

    Propolis has been used in folk medicine in different regions of the world including Latin America. Propolis is a resinous mixture of substances collected by honey bees from several botanical sources, and its composition contains a rich chemical variety, depending on the geographical area and plant sources. Our aim was to compare the modulatory effect of propolis samples from three different countries of Latin America (Brazil, Cuba and Mexico) on pro- and anti-inflammatory cytokine production (tumor necrosis factor (TNF)-α and interleukin (IL)-10, respectively) by human monocytes. Cells were incubated with propolis for 18 h at 37°C. Cell viability was assessed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide method, and cytokine production was determined by ELISA. All samples did not affect monocyte viability. Brazilian propolis stimulated both TNF-α and IL-10 production by monocytes. Cuban propolis stimulated TNF-α and inhibited IL-10 production, while Mexican sample exerted the opposite effect, inhibiting TNF-α and stimulating IL-10 production. The major compounds found in Brazilian, Cuban and Mexican propolis samples were artepillin C, isoflavonoids and pinocembrin, respectively. Brazilian, Cuban and Mexican propolis contained different components that may exert pro- and anti-inflammatory activity depending on concentration, what may provide a novel approach to the development of immunomodulatory drugs containing propolis. © 2015 Royal Pharmaceutical Society.

  17. Study of sample preparation for quantitative analysis of amino acids in human sweat by liquid chromatography-tandem mass spectrometry.

    Science.gov (United States)

    Delgado-Povedano, M M; Calderón-Santiago, M; Priego-Capote, F; Luque de Castro, M D

    2016-01-01

    The determination of physiological levels of amino acids is important to aid in the diagnosis and treatment of several diseases and nutritional status of individuals. Amino acids are frequently determined in biofluids such as blood (serum or plasma) and urine; however, there are less common biofluids with different concentration profiles of amino acids that could be of interest. One of these biofluids is sweat that can be obtained in a non-invasive manner and is characterized by low complex composition. The analysis of amino acids in human sweat requires the development of sample preparation strategies according to the sample matrix and small collected volume. The influence of sample preparation on the quantitative analysis of amino acids in sweat by LC-MS/MS has been assessed through a comparison between two strategies: dilution of sweat and centrifugal microsolid-phase extraction (c-μSPE). In both cases, several dilution factors were assayed for in-depth knowledge of the matrix effects, and the use of c-μSPE provided the best results in terms of accuracy. The behavior of the target analytes was a function of the dilution factor, thus providing a pattern for sample preparation that depended on the amino acid to be determined. The concentration of amino acids in sweat ranges between 6.20 ng mL(-1) (for homocysteine) and 259.77 µg mL(-1) (for serine) with precision, expressed as relative standard deviation, within 1.1-21.4%.

  18. A comparative study of some physico-chemical properties of human serum albumin samples from different