Human Feces Research Articles

Comparison of the Serovars and Characteristics of Salmonella Isolated from Human Feces and Foods in the 1990s and 2010s in Tokyo.

Salmonella foodborne disease outbreaks have markedly decreased in recent years, and different Salmonella serovars have been isolated. To clarify the characteristics of Salmonella strains causing annual epidemics and to estimate the source, we conducted a serotyping test on 1,132 human-derived Salmonella isolates in the 1990s and 2010s, and 1,061 food-derived Salmonella isolates in the 2010s in Tokyo. The serovars commonly isolated from human feces in the 1990s and after 2012 were S. Enteritidis, S. Typhimurium, S. Infantis, S. Thompson, and S. Agona. The new main serovars isolated after 2012 were S. Schwarzengrund, S. Enterica serovar 4:i:-, and S. Chester. In contrast, the main serovars detected from foods after 2012 were S. Infantis, S. Schwarzengrund, S. Agona, S. Manhattan, S. Typhimurium, and S. enterica serovar UT: r:1,5. S. Schwarzengrund has recently been frequently isolated. These strains were mainly isolated from chicken meat and offal. It was suggested that the same serovars of human-derived isolates were also isolated from foods, especially chicken meat and offal, and that these were recently an important causative food of Salmonellosis.

Prevalence and Characterization of CRISPR Locus 2.1 Spacers in Escherichia coli Isolates Obtained from Feces of Animals and Humans.

The clustered regularly interspaced short palindromic repeat (CRISPR) has been studied as an immune system in prokaryotes for the survival of bacteriophages. The CRISPR system in prokaryotes records the invasion of bacteriophages or other genetic materials in CRISPR loci. Accordingly, CRISPR loci can reveal a history of infection records of bacteriophages and other genetic materials. Therefore, identification of the CRISPR array may help trace the events that bacteria have undergone. In this study, we characterized and identified the spacers of the CRISPR loci in Escherichia coli isolates obtained from the feces of animals and humans. Most CRISPR spacers were found to stem from phages. Although we did not find any patterns in CRISPR spacers according to sources, our results showed that phage-derived spacers mainly originated from the families Inoviridae, Myoviridae, Podoviridae, and Siphoviridae and the order Caudovirales, whereas plasmid-derived CRISPR spacers were mainly from the Enterobacteriaceae family. In addition, it is worth noting that the isolates from each animal and human source harbored source-specific spacers. Considering that some of these taxa are likely found in the gut of mammalian animals, CRISPR spacers identified in these E. coli isolates were likely derived from the bacteriophageome and microbiome in closed gut environments. Although the bacteriophageome database limits the characterization of CRISPR arrays, the present study showed that some spacers were specifically found in both animal and human sources. Thus, this finding may suggest the possible use of E. coli CRISPR spacers as a microbial source tracking tool. IMPORTANCE We characterized spacers of CRISPR locus 2.1 in E. coli isolates obtained from the feces of various sources. Phage-derived CRISPR spacers are mainly acquired from the order Caudovirales, and plasmid-derived CRISPR spacers are mostly from the Enterobacteriaceae family. This is thought to reflect the microbiome and phageome of the gut environment of the sources. Hence, spacers may help track the encounter of bacterial cells with bacterial cells, viruses, or other genetic materials. Interestingly, source-specific spacers are also observed. The identification of source-specific spacers is thought to help develop the methodology of microbial source tracking and understanding the interactions between viruses and bacteria. However, very few spacers have been uncovered to track where they originate. The accumulation of genome sequences can help identify the hosts of spacers and can be applied for microbial source tracking.

Characterization and epidemiology of antimicrobial resistance patterns of Salmonella spp. and Staphylococcus spp. in free-ranging rhesus macaque (Macaca mulatta) at high-risk interfaces with people and livestock in Bangladesh.

Antimicrobial resistance (AMR) is a growing global health threat for humans and animals. Environmental contamination of antimicrobials from human and domestic animal feces has been linked to AMR in wildlife populations, including rhesus macaques. This study aimed to describe the eco-epidemiology of AMR within Salmonella and Staphylococcus species isolated from rhesus macaques. We followed macaque groups for 4 h per day (2 days) to observe the direct and indirect contact rate and type between macaques and people and livestock. We collected 399 freshly defecated, non-invasive fecal samples from macaques at seven sites in Bangladesh in January-June 2017. Bacterial isolation and identification were conducted using culture, biochemical characteristics, and polymerase chain reaction (PCR). An antimicrobial susceptibility test (AST) for 12 antimicrobials for each organism was conducted using the Kirby-Bauer disc diffusion method. The overall prevalence of Salmonella spp. and Staphylococcus spp. in rhesus macaques was 5% (n = 18; 95% CI: 3-7%) and 16% (n = 64; 95% CI: 13-20%), respectively. All the isolated Salmonella spp. and most of the Staphylococcus spp. (95%; 61/64; 95% CI: 86.9-99%) were resistant to at least one antimicrobial. The odds of a fecal sample having antimicrobial-resistant Salmonella spp (OR = 6.6; CI: 0.9-45.8, P = 0.05) and Staphylococcus spp. (OR = 5.6; CI: 1.2-26, P = 0.02) were significantly higher in samples collected at peri-urban sites than those collected at rural and urban sites. Salmonella spp. were most frequently resistant to tetracycline (89%), azithromycin (83%), sulfamethoxazole-trimethoprim (50%), and nalidixic acid (44%). Staphylococcus spp. were found to be highly resistant to ampicillin (93%), methicillin (31%), clindamycin (26%), and rifampicin (18%). Both bacterial species produced colonies with multidrug resistance to up to seven antimicrobials. Direct and indirect contact rates (within 20 m for at least 15 min) and resource sharing between macaques and people were higher in urban sites, while macaque-livestock contact rates were higher in rural sites. The study shows that resistant microorganisms are circulating in rhesus macaque, and direct and indirect contact with humans and livestock might expand the resistant organisms.

In Vitro Screen of Lactobacilli Strains for Gastrointestinal and Vaginal Benefits

Traditional probiotics comprise mainly lactic acid bacteria that are safe for human use, tolerate acid and bile, and adhere to the epithelial lining and mucosal surfaces. In this study, one hundred commercial and non-commercial strains that were isolated from human feces or vaginal samples were tested with regards to overall growth in culture media, tolerance to acid and bile, hydrogen peroxide (H2O2) production, and adhesion to vaginal epithelial cells (VECs) and to blood group antigens. As a result, various of the tested lactobacilli strains were determined to be suitable for gastrointestinal or vaginal applications. Commercial strains grew better than the newly isolated strains, but tolerance to acid was a common property among all tested strains. Tolerance to bile varied considerably between the strains. Resistance to bile and acid correlated well, as did VEC adhesion and H2O2 production, but H2O2 production was not associated with resistance to bile or acid. Except for L. iners strains, vaginal isolates had better overall VEC adhesion and higher H2O2 production. Species- and strain-specific differences were evident for all parameters. Rank-ordered clustering with nine clusters was used to identify strains that were suitable for gastrointestinal or vaginal health, demonstrating that the categorization of strains for targeted health indications is possible based on the parameters that were measured in this study.

Akkermansia biwaensis sp. nov., an anaerobic mucin-degrading bacterium isolated from human faeces.

A bacterial strain, WON2089T, was isolated from the faeces of healthy Japanese adults and is able to use mucin as the sole carbon and nitrogen source. Sequencing of its 16S rRNA gene showed that WON2089T has 98.0 and 94.4% similarity to Akkermansia muciniphila MucT and Akkermansia glycaniphila PytT, respectively, while phylogenetic tree analysis confirmed that it belongs to the genus Akkermansia. The whole genome of WON2089T was sequenced, which showed that it shares 84.5 % average nucleotide identity (ANI) and 24.9 % digital DNA-DNA hybridization (dDDH) with its closest relative, A. muciniphila MucT. Cells of WON2089T are non-motile, anaerobic and oval-shaped (0.4-0.5×0.5-1.0 µm). The strain is Gram-stain-negative and grows in the temperature range of 25-45 °C (optimum, 30-37 °C) and pH range of pH 5.5-9.5 (optimum, pH 6.5-8.0). WON2089T can utilize d-glucose, d-mannitol, lactose and d-mannose, as assessed by API20A strips. The major cellular fatty acids are C15 : 0 anteiso, C15 : 0 3OH and C18 : 1 ω9c (55.5, 7.5 and 5.8 % of total fatty acids, respectively). Based on 16S rRNA sequencing, ANI, dDDH and acid formation from d-mannitol, WON2089T is distinct from previously reported species of the genus Akkermansia. Based on phenotypic, phylogenetic and genetic characteristics, WON2089T represents a novel species of the genus Akkermansia and the name Akkermansia biwaensis sp. nov. is proposed. The type strain is WON2089T (= NBRC 115679T= DSM 114407T).

Archaeological Evidence for the Dietary Practices and Lifestyle of 18th Century Lisbon, Portugal—Combined Steroidal Biomarker and Microparticle Analysis of the Carbonized Faecal Remains

The study of the urban context in the contemporary center of Portugal’s capital city uncovered traces of daily lives that were abruptly interrupted and utterly transformed by the Great Lisbon Earthquake on the morning of 1 November 1755. Charred organic residue was recovered from a cylindrical vessel excavated from the storage area of the town house at the Rossio square. The archaeological sample was studied through a multi-analytical approach based on microstructural, elemental and biomolecular characterization by attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FT-IR), variable pressure scanning electron microscopy coupled to energy dispersive X-ray spectroscopy (VP-SEM-EDS), and gas chromatography coupled with mass spectrometry (GC-MS). The residue was identified as human faeces collected in the ceramic vessel for disposal, and further analysis provided additional information about diet and the living conditions in the 18th century.

Occurrence and risk assessment of antibiotics in feces of elderly individuals in Shenzhen.

The occurrence of antibiotics in the feces of elderly individuals in Shenzhen, China, was investigated by monitoring 78 compounds to understand the adverse effects and its association with antibiotic residues in animal products collected from local markets. In total, 18 compounds belonging to 5 classes of antibiotics were identified in 74 of 140 fecal samples. Furthermore, 17.9% of the fecal samples contained at least two antibiotics, and 14.3% of the samples showed antibiotic concentrations higher than 100μg/kg. Cephalothin exhibited the highest detection frequency (22.1%), followed by azithromycin (15.7%) and tilmicosin (12.9%). Oxytetracycline, norfloxacin, and azithromycin showed extremely high concentrations (> 1000μg/kg). Eight antibiotics were detected in the animal products, with detection frequencies ranging from 4.8 to 40.0%. Five antibiotics exhibited similar detection frequencies and strong correlations between the human fecal and animal product samples. Health risk assessment based on hazard quotients showed that ciprofloxacin in animal products and human feces posed a medium and high risk, respectively. The hazard quotients of oxytetracycline, norfloxacin, and azithromycin in the feces were greater than 1, indicating a high health risk. These findings suggest that the elderly individuals were frequently exposed to antibiotics via the food chain and faced health risks posed by these antibiotics.

Virus Emissions from Toilet Flushing: Comparing Urine-Diverting to Mix Flush Toilets.

High levels of viruses can be found in human excrement from infected individuals, a fraction of which can be emitted from toilet flushing. Unlike the common mix flush toilet (MFT), the urine-diverting toilet (UDT) separates urine from the toilet water. Specific focus on urine-associated viruses is needed because the UDT can emit different levels of urine-associated and fecal-borne viruses and urine has different properties compared to feces that can affect emission levels (e.g., protein content). In this work, we quantified emission levels of surrogate bacteriophages for urine-associated and fecal-borne viruses, MS2 and T3, from flushing a UDT and an MFT, with and without protein in the water. Emission levels of viruses in the water of the UDT were lower than that of the MFT by up to 1.2-log10 and 1.3-log10 for T3 and MS2, respectively. If urine is completely diverted in the UDT, virus emissions can be reduced by up to 4-log10. Based on these results and typical levels in urine and feces, we estimate that up to 107 and 108 gene copies of human viruses per flush can be released from the UDT and MFT, respectively. Lower emissions observed with the UDT suggest reduced exposure to viruses from flushing the UDT.

Interventions to improve sanitation for preventing diarrhoea.

Interventions to improve sanitation for preventing diarrhoea.

In <i>Vitro </i>Evaluation of Putative Probiotic Candidates Isolated from Various Origins

Present study describes isolation of potential probiotic lactic acid bacteria from chicken crop, human feces, buttermilk and chilly. The isolated <i>Lactobacillus</i> strains survive, tolerate and grow in MRS medium spiked with bile, salt and having acidic pH. The <i>Lactobacillus</i> isolates possess several probiotic properties, viz. (i) ability to bind gastrointestinal mucosa, up to ≥80% cells adhered mucin, (ii) 50% cells retained viability during oro-gastro-intestinal transit, (iii) all the isolates exhibited broad anti-microbial spectrum against food spoilage and gastro-intestinal pathogens,<i> Limosilactobacillus fermentum</i> SBM showed maximum inhibition, (iv) ability to produce enzymatic activities like L-asparaginase, β-galactosidase and bile salt hydrolase (BSH) activities, <i>Limosilactobacillus fermentum</i> SBM showed maximum L-asparaginase activity (2.567 U/ml), and<i> Lactiplantibacillus pentosus</i> GCHI showed maximum β-galactosidase activity (296±0.1 Miller’s Unit), (v) <i>Lactiplantibacillus pentosus</i> GCHI aggregated up to ≥92% after 24 h, and (vi) the <i>Lactobacillus</i> isolates were susceptible towards nucleic acid synthesis inhibitors and cell wall synthesis inhibitor antibiotics. These <i>Lactobacillus</i> strains do not possess haemolytic, mucin degrading and DNase activities indicating their safety. Further characterization of these strains indicated potential probiotic properties and their suitability in food formulations as probiotics. The study presents an interesting illustration of mining of potential probiotic strains from nature exhibiting health benefits for human being and animals.

Bacteroides faecium sp. nov. isolated from human faeces.

A novel bacterial strain, CBA7301T, was isolated from human faeces and was characterised using a polyphasic taxonomic approach. A phylogenetic analysis based on 16S rRNA gene sequences revealed that CBA7301T represented a member of the genus Bacteroides, in the family Bacteroidaceae. The similarity between the 16S rRNA gene sequence of CBA7301T and that of its most closely related species, Bacteroides faecichinchillae JCM 17102T, was 96.2 %, and the average nucleotide identity between these two strains was 77.9 %. The genome size was 6 782 182 bp, and the DNA G+C content was 42.5 mol%. Cells of CBA7301T were Gram-stain-negative, strictly anaerobic and rod-shaped. The optimal growth of this organism occurred at 30-35 °C, pH 7.0 and 0.5 % (w/v) NaCl. The respiratory quinone was menaquinone 10. The predominant polar lipids were phosphatidylethanolamine, phospholipids and aminophospholipids. The major cellular fatty acid was anteiso-C15 : 0. According to the results of the polyphasic taxonomic analysis, CBA7301T represents a novel species of the genus Bacteroides, which we named Bacteroides faecium sp. nov. The type strain is CBA7301T (=KCCM 43355T=ATCC TSD-227T).

Formulation of a Simulated Wastewater Influent Composition for Use in the Research of Technologies for Managing Wastewaters Generated during Manned Long-Term Space Exploration and Other Similar Situations—Literature-Based Composition Development

The prospect of humans inhabiting planetary bodies is gaining interest among research and development communities, with the moon being considered as a transitory base camp and Mars the next planet humans will inhabit. NASA's Mission to Mars program is set to have humans inhabiting Mars within on-planet space camps by the Year 2030, which has tremendously increased research and development for space exploration-including research oriented toward human life support in long-term planetary lodging camps. The sustenance of human life on Mars will not be trivial due to the unavailability of an appropriate atmosphere and usable water. This situation requires a self-sustaining human life support system that can provide the basic needs such are breathable air, potable water, food, and energy. The feasibility of sending a payload with resources adequate to support long-term human inhabitation is not reasonable, which means every resource within a Mars space camp is valuable, including human-produced wastes. A biorefinery system that treats wastewater and can also produce valuable products such as oxygen, food, and energy offers a form of circular utilization of valuable resources. To conduct research for such systems requires a wastewater influent that is representative of the wastewater to be generated by the space crew within this isolated, confined environment, which is different from what is generated on Earth due to limited variability in diet, human activity, and lifestyle in this confined area. Collection of actual wastewater influent from an isolated environment supporting humans is challenging. Additionally, to ensure a safe working environment in the laboratory and avoid the imposed threat of handling actual human feces, the proposed synthetic, non-human feces containing wastewater influent formulation offers an easy-to-produce and safer-to-handle option. This paper reviews several synthetic wastewater compositions that have been formulated for space exploration purposes. None of the formulations were found to be realistic nor adequate for a space-camp-type scenario. Thus, the formulation of a synthetic wastewater for simulating a wastewater influent from a human space-based camp is proposed in this paper. In addition, the physical, chemical, and biodegradation characteristics of the final formulation designed are presented to illustrate the value of the proposed influent formulation.

Enterococcus faecium Bacteriophage vB_EfaH_163, a New Member of the Herelleviridae Family, Reduces the Mortality Associated with an E. faecium vanR Clinical Isolate in a Galleria mellonella Animal Model.

The rise of antimicrobial resistant (AMR) bacteria is a major health concern, especially with regard to members of the ESKAPE group, to which vancomycin-resistant (VRE) Enterococcus faecium belongs. Phage therapy has emerged as a novel alternative for the treatment of AMR infections. This, however, relies on the isolation and characterisation of a large collection of phages. This work describes the exploration of human faeces as a source of new E. faecium-infecting phages. Phage vB_EfaH_163 was isolated and characterised at the microbiological, genomic, and functional levels. vB_EfaH_163 phage, a new member of Herelleviridae, subfamily Brockvirinae, has a dsDNA genome of 150,836 bp that does not harbour any virulence factors or antibiotic resistance genes. It infects a wide range of E. faecium strains of different origins, including VRE strains. Interestingly, it can also infect Enterococcus faecalis strains, even some that are linezolid-resistant. Its capacity to control the growth of a clinical VRE isolate was shown in broth culture and in a Galleria mellonella animal model. The discovery and characterisation of vB_EfaH_163 increases the number of phages that might be used therapeutically against AMR bacteria.

Prevalence and risk factors of hookworm infections among vegetable growers in Anhui Province from 2018 to 2019

To investigate the prevalence and risk factors of hookworm infections among vegetable growers in Anhui Province, so as to provide the evidence for formulating the control strategy and interventions for hookworm human infections. A total of 1 179 vegetable growers were randomly sampled from 7 counties in Anhui province according to the natural ecological function areas in 2018 and 2019, and the basic features and risk factors of hookworm infections were collected from all subjects using a self-designed questionnaire, including living environment, vegetable planting and fertilization, type of household crops, animal feeding, hygiene and labor habits. In addition, stool samples were collected from all participants, and each stool sample was subjected to twice tests with the Kato-Katz technique. The risk factors of hookworm infections were identified among vegetable growers with the Bayesian log-binomial regression model. The prevalence of hookworm infection was 3.90% among the 1 179 vegetable growers. The prevalence of hookworm infection was 10.07% (15/149) among unemployed subjects, 5.32% (31/583) among farmers, and 0 among full-time vegetable growers (0/377) and subjects with other occupations (0/70), and there was a significant difference among subjects in terms of occupation (χ2=36.37, P < 0.01). Bayesian log-binomial regression analysis showed that fertilization with human feces [relative risk (RR)=18.25, 95% confidence interval (CI): (4.31, 77.17)], barefoot labor in vegetable fields [RR=2.86, 95% CI: (1.43, 5.70)], and living in plain ecological areas along the middle and lower reaches of the Yangtze River [RR=4.19, 95% CI: (1.19, 14.81)] were independent risk factors of hookworm infections among vegetable growers. Stratified analysis showed that fertilization with human feces, barefoot labor in vegetable fields, drinking well water or mountain spring, and living in plain ecological areas along the middle and lower reaches of the Yangtze River were independent risk factors of hookw, orm infection among farmers, and living in plain ecological areas along the middle and lower reaches of the Yangtze River and growing rice were independent risk factors of hookw, orm infection among unemployed individuals. The prevalence of hookworm infection is high among vegetable growers in Anhui Province, and fertilization with human feces, barefoot labor in vegetable fields and living in plain ecological areas along the middle and lower reaches of the Yangtze River are risk factors of hookw, orm infections among vegetable growers.

COVID-19 surveillance in wastewater: An epidemiological tool for the monitoring of SARS-CoV-2.

The coronavirus disease 2019 (COVID-19) pandemic has prompted a lot of questions globally regarding the range of information about the virus's possible routes of transmission, diagnostics, and therapeutic tools. Worldwide studies have pointed out the importance of monitoring and early surveillance techniques based on the identification of viral RNA in wastewater. These studies indicated the presence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in human feces, which is shed via excreta including mucus, feces, saliva, and sputum. Subsequently, they get dumped into wastewater, and their presence in wastewater provides a possibility of using it as a tool to help prevent and eradicate the virus. Its monitoring is still done in many regions worldwide and serves as an early "warning signal"; however, a lot of limitations of wastewater surveillance have also been identified.

Research advances of microplastics and potential health risks of microplastics on terrestrial higher mammals: a bibliometric analysis and literature review.

Microplastics (MPs) have become increasingly serious global problems due to their wide distribution and complicated impacts on living organisms. To obtain a comprehensive overview of the latest research progress on MPs, we conducted a bibliometric analysis combined with a literature review. The results showed that the number of studies on MPs has grown exponentially since 2010. Recently, the hotspot on MPs has shifted to terrestrial ecosystems and biological health risks, including human health risks. In addition, the toxic effects, identification and quantification of MPs are relatively new research hotspots. We subsequently provide a review of MPs studies related to health risks to terrestrial higher mammals and, in particular, to humans, including detection methods and potential toxicities based on current studies. Currently, MPs have been found existing in human feces, blood, colon, placenta and lung, but it is still unclear whether this is associated with related systemic diseases. In vivo and in vitro studies have demonstrated that MPs cause intestinal toxicity, metabolic disruption, reproductive toxicity, neurotoxicity, immunotoxicity through oxidative stress, apoptosis and specific pathways, etc. Notably, in terms of combined effects with pollutants and neurotoxicity, the effects of MPs are still controversial. Future attention should be paid to the detection and quantification of MPs in human tissues, exploring the combined effects and related mechanisms of MPs with other pollutants and clarifying the association between MPs and the development of pre-existing diseases. Our work enhances further understanding of the potential health risks of MPs to terrestrial higher mammals.

Mapping opportunity structure: exploring Dalit-led litigation on manual scavenging

Given a disproportionate scholarly focus on a high-profile litigation on manual scavenging – the manual cleaning of human faeces – in India, this paper aims to explore the experiences of NGOs led by Dalits – formerly “untouchables” – in navigating the judiciary to eradicate manual scavenging. It does so by first exploring the frequency and characteristics of court cases on manual scavenging, and second by exploring obstacles and opportunities that affect whether and how Dalit actors involve courts in their efforts. Through a mixed- and multi-method research, drawing on participatory field research, data from Right to Information requests, court judgments and interviews, the paper operationalises the concept of legal opportunity structure and shows that the choice to litigate is made carefully and is shaped by practical concerns. Notably, caste normativity both pushes Dalit actors towards litigation, and shapes how Dalit actors can engage with the judiciary. This power of caste to shape recourse to state law has serious implications for the idea that law and the judiciary are neutral and available to all equally. While previous literature focuses on public interest litigation and high-profile court cases on manual scavenging, this paper provides the first dataset on manual scavenging litigation, shows that such cases are the minority, and foregrounds experiences of Dalit actors.

Unraveling the influence of human fecal pollution on antibiotic resistance gene levels in different receiving water bodies using crAssphage indicator gene.

Discharged wastewater treatment plant (WWTP) effluents can contaminate receiving water bodies with human feces and alter the abundance of antibiotic resistance genes (ARGs). In this study, we examined the co-occurrence of ARGs, human fecal pollution indicator crAssphage, and antibiotics in human feces and a series of connected receiving water bodies affected by human feces, including water from different treatment units of a WWTP, river, lake, and tap waters. Results showed that crAssphage was detected in 68.2 % of the studied water bodies, confirming widespread human fecal contamination. Both ARG and crAssphage abundances exhibited a distance-decay effect from the emission source to the receiving environment. Interestingly, the detected ARG abundance in the water bodies was significantly correlated with crAssphage abundance but not with the residual antibiotic concentration, demonstrating that the presence of ARG could largely be explained by the extent of fecal pollution, with no clear signs of antibiotic selection. In addition, 14 ARGs co-shared by human feces and water bodies were significantly correlated with crAssphage. Furthermore, a close evolutionary relationship was observed between the blaTEM-1 gene from human feces and aquatic environments. These results imply a potential ARG exchange between human feces and receiving water bodies. Overall, this study provides important insights into the distribution and sources of ARGs in water bodies affected by human fecal contamination.

Сlinical-Epidemiological, Immunological, Microbiological Aspects and Prevalence of Blastocystosis among the Population in the Republic of Azerbaijan

Blastocystis hominis is currently one of the most common parasites found in human feces - up to 10% of the population in the developed countries and approximately 50-60% of cases are found particularly in the developing countries. Nevertheless blastocystosis is one of the least studied infectious diseases among intestinal protozoans in the Republic of Azerbaijan. The aim of this study was to assess morbidity of blastocystosis, to determine its clinical picture, its role in the immune system and in the intestinal microflora. Methods. This study involved a total of 8002 patients who applied to the Parasitology Clinic of the Research Institute of Medical Prevention named after V.Yu. Akhundova in Baku, Azerbaijan. As part of the study, the feces of the participants were examined microscopically, using ELISA and PCR methods. Results. This study has presented that course of infection is mostly asymptomatic, although patients could experience abdominal pain, nausea, diarrhea, foul-smelling stool, skin rashes, allergy symptoms, hypersalivation and unpleasant breath. The increase of IL8 was detected among most patients which has indicated negative effect on the immune system, moreover it has caused quantitative and qualitative changes in the normal microflora of the intestines. Conclusions. Based on the results we came to conclusion that blastocystosis has been very widespread among population of Republic of Azerbaijan and it is still vital to continue in-depth study.

Purification of Bacterial-Enriched Extracellular Vesicle Samples from Feces by Density Gradient Ultracentrifugation.

Commensal microbiota has huge impact on the maintenance of human health, its dysregulation being associated with the development of a plethora of diseases. Release of bacterial extracellular vesicles (BEVs) is a fundamental mechanism of systemic microbiome influence on the host organism. Nevertheless, due to the technical challenges of isolation methods, BEV composition and functions remain poorly characterized. Hereby, we describe the up-to-date protocol for isolation of BEV-enriched samples from human feces. Fecal extracellular vesicles (EVs) are purified through the orthogonal implementation of filtration, size-exclusion chromatography (SEC), and density gradient ultracentrifugation. EVs are first separated from bacteria, flagella, and cell debris by size. In the next steps, BEVs are separated from host-derived EVs by density. The quality of vesicle preparation is estimated via immuno-TEM (transmission electron microscopy) for the presence of vesicle-like structures expressing EV markers and via NTA (nanoparticle tracking analysis) for assaying particle concentration and size. Distribution of EVs of human origin in gradient fractions is estimated using antibodies against human exosomal markers with Western blot and ExoView R100 imaging platform. The enrichment for BEVs in vesicle preparation is estimated by Western blot for the presence of bacterial OMVs (outer membrane vesicles) marker and OmpA (outer membrane protein A). Taken together, our study describes a detailed protocol for EV preparation with enrichment for BEVs from feces with a purity level suitable for bioactivity functional assays.