Search

Short-chain fatty acids (SCFAs) are involved in important physiological processes such as gut health and immune response, and changes in SCFA levels can be indicative of disease. Despite the importance of SCFAs in human health and disease, reference values for fecal and plasma SCFA concentrations in healthy individuals are scarce. To address this gap in current knowledge, we developed a simple and reliable derivatization-free GC-TOFMS method for quantifying fecal and plasma SCFAs in healthy individuals. We targeted six linear- and seven branched-SCFAs, obtaining method recoveries of 73–88% and 83–134% in fecal and plasma matrices, respectively. The developed methods are simpler, faster, and more sensitive than previously published methods and are well suited for large-scale studies. Analysis of samples from 157 medically confirmed healthy individuals showed that the total SCFAs in the feces and plasma were 34.1 ± 15.3 µmol/g and 60.0 ± 45.9 µM, respectively. In fecal samples, acetic acid (Ace), propionic acid (Pro), and butanoic acid (But) were all significant, collectively accounting for 89% of the total SCFAs, whereas the only major SCFA in plasma samples was Ace, constituting of 93% of the total plasma SCFAs. There were no statistically significant differences in the total fecal and plasma SCFA concentrations between sexes or among age groups. The data revealed, however, a positive correlation for several nutrients, such as carbohydrate, fat, iron from vegetables, and water, to most of the targeted SCFAs. This is the first large-scale study to report SCFA reference intervals in the plasma and feces of healthy individuals, and thereby delivers valuable data for microbiome, metabolomics, and biomarker research.

Per- and polyfluoroalkyl substances (PFAS) are ubiquitous environmental contaminants with half-lives in humans in the range of years in case of the long-chain compounds, leading to accumulation and measurable levels in plasma. In contrast, short-chain and “alternative” PFAS have lower levels or are not detectable in humans with background exposure. This may be due to lower exposure, but also due to much shorter half-lives compared to long-chain compounds. To get better data on kinetics, a healthy volunteer orally ingested a mixture of fifteen predominantly 13C-labeled PFAS (“MPFAS”) in a pilot investigation (MPFBA, MPFPeA, MPFHxA, MPFHpA, MPFOA, MPFNA, MPFDA, MPFUdA, MPFDoA, PFBS, MPFHxS, MPFOS, DONA, HFPO-DA, 6:2FTS). After application, concentrations were measured over 450 days in plasma, urine and feces, using UHPLC-MS/MS analysis after extraction. The compounds were absorbed quickly and almost completely. Data analysis revealed volumes of distribution between 110 and 177 mL/kg bw for most compounds, but higher values for MPFDA, MPFUdA and MPFDoA (maximum of 354 mL/kg bw). Half-lives were found to vary extremely, from 0.5 days (MPFPeA) and 1.5 days (MPFHxA) to 51 days (PFBS) and 152 days (MPFHpA) in case of the short-chain and “alternative” compounds. For the long-chain compounds, half-lives in the range of several years were confirmed for MPFOA, MPFNA, MPFHxS and MPFOS, but with even higher chain-lengths of the carboxylic acids, the half-lives were found to decrease, with the shortest half-life for MPFDoA (295 days). Elimination from the body was completely explained by the urinary losses in case of the short-chain and “alternative” PFAS, and in part by the fecal losses in case of the long-chain PFCA. Overall, elimination kinetics seem to be determined by several different renal and gastrointestinal factors (fraction unbound in plasma, binding affinity to organic anion transporters causing netto secretion or reabsorption, fecal loss with mechanisms to be clarified).

Environmental pollution is widespread and poses significant risks to both human populations and diverse animal species. As part of conservation initiatives, many endangered animals are housed in zoos and breeding centers close to human activities, potentially exposing them to health threats arising from environmental contamination, such as mercury (Hg). By analyzing non-invasive fecal samples from 13 Chinese zoos, we assessed total mercury (THg) and methylmercury (MeHg) exposure in two non-human primate species: the endangered species golden snub-nosed monkey (Rhinopithecus roxellana), and the most widely distributed rhesus macaque (Macaca mulatta). The concentration of THg in golden snub-nosed monkeys (mean ± s.d.: 76.7 ± 35.6 ng/g; median: 71.8 ng/g, range: 11.2–200.0 ng/g, n = 71) was significantly higher than that in rhesus macaques (mean ± s.d.: 32.3 ± 54.7 ng/g; median: 17.5 ng/g, range: 7.4–409.0 ng/g, n = 98). No significant differences were observed in both MeHg and MeHg % between these two species. Age and sex did not predict fecal Hg. Among adults, approximately 85 % of sampled individuals exhibited hazard quotient (HQ) values exceeding 1, indicating mercury poses a health risk to captive primate populations. The average HQ for golden snub-nosed monkeys was 2.98 ± 1.68 (range: 0.46–8.21, n = 43), while for rhesus macaques was 1.84 ± 1.64 (range: 0.61–12.49, n = 71). Our findings suggest that conservation efforts for captive primates may be compromised by Hg pollution, particularly in golden snub-nosed monkeys. Further investigation and ongoing biomonitoring from an ecotoxicological perspective, are crucial to ensuring the health of captive primate populations.

Objective: This work aimed to explore the modified pathways impacted by changes in fecal metabolites among female obese adults during the human gut model period, both in the presence and absence of individual Triphala constituent fruit extract. Methods and results: The human gut model employed individual fruit extracts from Phyllanthus emblica, Terminalia bellerica, and T. chebula, comparing them to a control group. Fermentation used fecal samples from female obese adults over 24 h. Metabolite extraction, untargeted metabolomics, and pathway analysis identified metabolic changes. Treatments with P. emblica extract, T. bellerica extract, and T. chebula extract revealed the statistical detection of 128, 734, and 757 up-regulated metabolites, respectively, while 31, 90, and 92 down-regulated metabolites were identified. Pathway analysis revealed that P. emblica extract primarily influenced vitamin B6 metabolism, whereas the treatments with T. bellerica extract and T. chebula extract predominantly engaged glycine, serine, and threonine metabolism for metabolic regulation within the human gut model. Conclusion: By examining obese fecal metabolite changes and their association with metabolic pathway modulation through individual fruit extracts of Triphala constituents in a human gut model, this study provides a comprehensive understanding of Triphala components’ potential for managing obesity and its applications in the food industry.

ABSTRACT: The present study aimed to evaluate the effect of continuous butyrate administration in dairy calves' liquid diet considering diarrhea, metabolic profile, gastrointestinal development, and corporal growth. Immediately after birth, calves were randomly allocated into 2 groups of 62 calves (50 females and 12 males), with access to water and a solid feed ad libitum. The butyrate group (BG) received 4 g/d of sodium butyrate (Admix Easy, Adisseo) diluted in the whole milk, and the control group (CG) received whole milk with no supplementation. Sodium butyrate was administered from d 1 of life until the weaning at 90 d. Feces consistency was assessed daily for the first 30 d of life and characterized by scores from 0 to 4 (0 and 1 for normal, and 2, 3, and 4 for abnormal feces). Diarrhea was diagnosed when the animals had abnormal feces and fever. Morbidity, recurrence, mortality, and lethality data were recorded and compared between the groups. Average daily gain (ADG) and corporal growth (body weight, thoracic perimeter, height at the withers, and croup width) were evaluated weekly, from the first day to d 30, and later at 45, 60, and 90 d of life. Blood samples were taken weekly for up to 30 d to determine the circulating levels of total calcium, phosphorus, chloride, bicarbonate, glucose, β-hydroxybutyrate, and nonesterified fatty acids. The males were euthanized at 15 (n = 6 per group) and 30 d (n = 6 per group) for morphometric, histological, and gene expression analysis of the gastrointestinal tract. The results showed that the BG had a lower rate of morbidity (BG = 30% vs. CG = 50%) and recurrence (BG = 26.7% vs. CG = 60%) of diarrhea than the CG. In addition, the BG had abnormal feces for a shorter period (BG = 4.64 ± 0.47 d vs. CG = 8.6 ± 0.65 d). The ADG tended to be higher in BG than CG up to 30 and 60 d. Metabolic evaluations showed the lowest levels of glucose and highest levels of nonesterified fatty acids in BG. On d 30 of life, rumen papillae length, papilla area, duodenum villus length, and crypt depth were higher in BG than in CG. The duodenal gene expression at 30 d showed that animals with diarrhea episodes that did not receive butyrate had the highest levels of transcripts for the LCT and GLP2 genes. In addition, in different ways, both butyrate and neonatal diarrhea affected the gene expression of IGF1, SLC5A1, and AQP3. These results allow us to conclude that continuous supplementation with sodium butyrate improves gastrointestinal development, reduces the occurrence of diarrhea, and makes clinical conditions milder with faster recovery, favoring a higher ADG in the first 30 and 60 d of life. Based on these results, we conclude that sodium butyrate can be indicated for liquid diet supplementation to accelerate gastrointestinal tract development and prevent severe cases of neonatal diarrhea, tending to improve average daily gain until weaning.

Objective: The incidence and prevalence of inflammatory bowel disease (IBD) in pediatric patients are increasing. Currently, the diagnostic method for IBD is inconvenient, expensive, and difficult. S100A12, a type of calcium-binding protein, detected in the feces of patients with IBD has recently been suggested as a promising diagnostic tool. Hence, the authors aimed to evaluate the accuracy of fecal S100A12 in diagnosing IBD in pediatric patients by performing a meta-analysis. Methods: The authors performed a systematic literature search in five electronic databases for eligible studies up to July 15, 2021. Pooled diagnostic accuracies of fecal S100A12 were analyzed as the primary outcomes. Secondary outcomes were standardized mean difference (SMD) of fecal S100A12 levels between IBD and non-IBD groups and a comparison of diagnostic accuracies between fecal S100A12 and fecal calprotectin. Results: Seven studies comprising 712 children and adolescents (474 non-IBD controls and 238 IBD cases) were included. Fecal S100A12 levels were higher in the IBD group than in the non-IBD group (SMD = 1.88; 95% confidence interval [CI] = 1.19–2.58; p < 0.0001). Fecal S100A12 could diagnose IBD in pediatric patients with a pooled sensitivity of 95% (95% CI = 88%–98%), specificity of 97% (95% CI = 95%–98%), and area under the receiver operating summary characteristics (AUSROC) curve of 0.99 (95% CI = 0.97–0.99). Fecal S100A12 specificity and AUSROC curve values were higher than those of fecal calprotectin (p < 0.05). Conclusion: Fecal S100A12 may serve as an accurate and non-invasive tool for diagnosing pediatric IBD.

Pigs are natural host to various zoonotic pathogens including viruses. In this study, we analyzed the viral communities in the feces of 89 piglets with diarrhea under one month old which were collected from six farms in Jiangsu Province of the Eastern China, using the unbiased virus metagenomic method. A total of 89 libraries were constructed, and 46937894 unique sequence reads were generated by Illumina sequencing. Overall, the family Picornaviridae accounted for the majority of the total reads of putative mammalian viruses. Ten novel virus genomes from different family members were discovered, including Parvoviridae (n = 2), Picobirnaviridae (n = 4) and CRESS DNA viruses (n = 4). A large number of phages were identified, which mainly belonged to the order Caudovirales and the family Microviridae. Moreover, some identified viruses were closely related to viruses found in non-porcine hosts, highlighting the potential for cross-species virus dissemination. This study increased our understanding of the fecal virus communities of diarrhea piglets and provided valuable information for virus monitoring and preventing.

Benzylalkyldimethylammonium (BACs), dialkyldimethylammonium (DDACs), and alkyltrimethylammonium compounds (ATMACs) are quaternary ammonium compounds (QACs) used widely as biocides, disinfectants, and sanitizers. Owing to their toxicity, human exposure to this class of chemicals is a concern. Pet animals are sentinels of human exposure to several indoor environmental chemicals. For the first time, we measured 7 BACs, 6 DDACs, 6 ATMACs, and 8 metabolites of BACs in urine and feces of pet dogs and cats from New York State, USA. We found widespread occurrence of QACs in feces, with median concentration of ∑All (sum concentration of all 27 QAC analytes) at 9680 and 1260 ng/g dry weight (dw) in dog and cat feces, respectively. BACs were the most abundant compounds among the four types of QACs, accounting for 64 % and 57 % of ∑All in dog and cat feces, respectively, followed by DDACs (33 % and 34 %, respectively), ATMACs (4 % and 9 %, respectively), and BAC metabolites (0.2 % and 0.3 %, respectively). However, in urine, only ω-carboxylic acid metabolites of BACs were found at median concentrations at 2.08 and 0.28 ng/mL in dogs and cats, respectively. Samples collected from animal shelters contained elevated levels of QACs than those from homes of pet owners. A significant positive correlation was found among the four types of QACs analyzed, which suggested usage of these chemicals in combination as mixtures. Based on the concentrations measured in feces, and through a reverse dosimetry approach, the median cumulative daily intakes (CDIs) of QACs were estimated to be 49.4 and 4.75 µg/kg body weight (BW)/day for dogs and cats, respectively. This study provides first evidence that pet dogs and cats are exposed to QACs at significant levels that warrant further attention.

To develop successful ex-situ breeding programs for the conservation of threatened wildlife, an understanding of reproductive endocrinology in the species of interest is essential. Fecal hormone metabolite patterns are widely used in this regard, but limited information is available regarding how these reflect blood hormone profiles. This study aimed to longitudinally monitor and compare circulating progestagen (sP) and estrogen (sE) concentrations, as well as fecal progestagen (fPM) and estrogen (fEM) metabolite concentrations with observed reproductive behaviors of African lionesses. For 18 months, blood sampling (n = 309) from five captive lionesses took place 1–7 times per week during positive reinforcement training. In parallel, over a period of 9 months, fecal samples (n = 129) from four of these lionesses were searched for daily and collected when found. Daily behavioral monitoring of all females enabled classification of reproductive stage according to presence/absence of characteristic behavioral events (i.e., “allowing mount”, “copulation”, “flirting run”, “lordosis”, “purring”, and “rolling”). Competitive enzyme immunoassays were used for steroid quantification. In total, 11 an-ovulatory and 16 ovulatory cycles (7 pregnancies and 9 pseudo-pregnancies) were assessed. When compared, the pattern of fecal hormone metabolite concentrations matched reproductive behavior more reliably than circulating steroid concentrations. Both sP and fPM patterns were correlated (n = 51; r = 0.86) and enabled identification of luteal phases, which helped to distinguish between pregnant and non-pregnant females. In contrast, detection of estrus by measurement of sE and fEM was not accurate regardless of the matrix evaluated.

Cranial radiotherapy is an important treatment for intracranial and head and neck tumors. To investigate the effects of cranial irradiation (C-irradiation) on gut microbiota and metabolomic profile, the feces, plasma and cerebral cortex were isolated after exposing mice to cranial X-ray irradiation at a dose rate of 2.33 Gy/min (5 Gy/d for 4 d consecutively). The gut microorganisms and metabolites were detected by 16 S rRNA gene sequencing method and LC-MS method, respectively. We found that compared with sham group, the gut microbiota composition changed at 2 W and 4 W after C-irradiation at the genus level. The fecal metabolomics showed that compared with Sham group, 44 and 66 differential metabolites were found to be annotated into metabolism pathways at 2 W and 4 W after C-irradiation, which were significantly enriched in the arginine and proline metabolism. Metabolome analysis of serum and cerebral cortex showed that, at 4 W after C-irradiation, the expression pattern of metabolites in serum samples of mice was similar to that of sham group, and the cerebral cortex metabolites of the two groups were completely separated. KEGG functional analysis showed that serum and brain tissue differential metabolites were respectively enriched in tryptophan metabolism, and arginine proline metabolism. The correlation analysis showed that the changes of gut microbiota genera were significantly correlated with the changes of metabolism, especially Helicobacter, which was significantly correlated with many different metabolites at 4 W after C-irradiation. These data suggested that C-irradiation could affect the gut microbiota and metabolism profile, even at relatively long times after C-irradiation.

Triphala is a mixture of tree fruits obtained from Terminalia chebula, Terminalia bellerica, and Phyllanthus emblica. It is one of the Ayurveda medicinal recipes used to treat health diseases such as obesity. The chemical composition analysis of Triphala extracts obtained from an equal portion of three fruits was performed. The contents of total phenolic compounds (62.87 ± 0.21 mg gallic acid equivalent/mL), total flavonoids (0.24 ± 0.01 mg catechin equivalent/mL), hydrolyzable tannins (177.27 ± 10.09 mg gallotannin equivalent/mL), and condensed tannins (0.62 ± 0.11 mg catechin equivalent/mL) were observed in Triphala extracts. The 1 mg/mL of Triphala extracts was applied to batch culture fermentation which contained the feces from voluntarily obese female adults (body mass index of 35.0–40.0 kg/m2) for 24 h. The extraction of DNA and metabolites was each conducted on the samples obtained from batch culture fermentation within and without Triphala extracts treatment. The 16S rRNA gene sequencing and untargeted metabolomic analysis were carried out. There was no statistically significant difference between Triphala extracts and control treatments on the changes in microbial profiles (p-value

Human excreta can be used as biofertilizers due to their nutrient and organic matter content. Nevertheless, the behavior of microorganisms should be investigated, as enteric pathogens can accumulate in the soil. Therefore, we evaluated the survival and transport of two enteric bacteria (E. coli and Salmonella enterica) and two enteric virus surrogates (MS2 and ΦX-174 bacteriophages) when applied as contaminants of four biofertilizers on Lactuca sativa production. The study was carried out in lysimeters outdoors with repacked silt loam soil, using a randomized block design (RBD) with six treatments and three replicates. Biofertilizers were urea-treated feces, feces composted with organic waste, stored liquid urine, and struvite and were inoculated at 106–107 cfu/pfu g−1 of E. coli and bacteriophages strains and 107–109 of S. enterica. Soil amended with composted feces exhibited the lowest decimal reductions (T90) for the microorganisms evaluated, with 10.2 days for E. coli, 11.9 days for S. enterica, 22.2 days for MS2, and 3.7 days for ΦX-174. Urea-treated feces temporarily hindered the growth and survival of E. coli and S. enterica in the soil. However, both bacteria were present after one month, while MS2 showed a stable concentration with this biofertilizer. E. coli presented a stable behavior when applied with urine, whereas MS2 was stable with urea-treated feces, urine, and struvite. ΦX-174 and S. enterica were not considered good representatives of most resistant enteric pathogens’ behavior in the soil during food production since they were not detected in leachate and showed a rapid die-off in soil. E. coli leached faster and in higher concentrations than MS2, which presented higher concentrations in deeper soil layers, from 10 to 20 cm. Therefore, using urine, struvite, and urea-treated faces implied a higher chance of infection, as a decimal reduction time was not observed during the 36 days of the experiment. As a result, in short-term cultures, it is recommended that additional hygiene barriers are taken to avoid infection since E. coli and MS2 were still detected at the end of the experiment. An interval of 90 days between fertilizing and harvesting is recommended to prevent pathogen infection. These results are the basis for a quantitative risk assessment of human excreta-based biofertilizers for food production.

Understanding snake prey breadth, within forest ecosystems, may elucidate their ecological role. We determined the prey items of 35 individuals of four snake species (11 through induced regurgitation analysis and 24 through molecular fecal analysis) collected from five mountain forests between May 2016 and October 2018, located within South Korea. Snake prey breadth spanned across six small mammal (4 rodents and 2 shrews) and five amphibian (2 urodeles and 3 anurans) species. Small mammals were more frequently detected as prey items than amphibians, but the proportion between small mammal and amphibian prey varied by different snake species. Two rodent prey species, Mus musculus and Craseomys regulus, made up 68.6% of the total amount of prey consumed (24 of 35 prey items). Elaphe dione and Gloydius intermedius samples contained only small mammal prey remains, whereas Rhabdophis lateralis and G. ussuriensis appeared to predate both small mammals and amphibians. These findings show snakes’ role in controlling small mammal population sizes in mountain forests and highlight the importance of conserving snake biodiversity native to South Korean mountain forests.

Here, using viral metagenomic method a novel picornavirus (named UJS-2019picorna, GenBank accession number OP821762) was discovered in fecal and blood samples of experimental rabbits (Oryctolagus cuniculus). The complete genome size of UJS-2019picorna is 7832 bp excluding the poly(A)-tail, with GC content of 44.00% and a nucleotide composition of 28.0% A, 28.0% U, 21.5% G, and 22.5% C. The viral genome has a typical picornavirus organization pattern from the 5′-3′ direction: VPg-5′ UTR-(L)-P1, (VP4–VP2–VP3–VP1)–P2, (2 A-2B–2C)–P3, (3 A-3B–3C-3D)-3′ UTR-poly(A). The P1 region of UJS-2019picorna is related to Erbovirus with amino acid identity of 37.31%, while the P2 and P3 regions are the closest to Bopivirus with amino acid identity of 35.66%–39.53%. According to the Picornaviridae Study Group guidelines, UJS-2019picorna should be presumed to be a new genus belonging to the Picornaviridae family. Epidemiologic study revealed that this novel picornavirus was prevalent in a cohort of experimental rabbits, with prevalence rate of 23.68% (9/38) in feces and 18.4% (7/38) in blood samples. Further work is required to elucidate whether this virus is pathogenic to rabbits and whether it has influence on studies using rabbits as experimental animal.

ABSTRACT: Polymerase chain reaction (PCR) method was coupled with a DNA extraction to enumerate Campylobacter spp. from poultry gastrointestinal tract samples. Three experiments were conducted that included: 1) Development of a DNA standard curve related to bacterial DNA primers; 2) Design of a cell/genomic DNA extraction protocol to isolate Campylobacter spp. DNA from complex samples such as poultry feces; and 3) Comparison of PCR quantification to standard plate count methodology. The standard curve using primers for Campylobacter spp. was created for DNA extracted from environmental isolates with a linear range (R2 > 0.95) and with a high specificity for C. coli and C. jejuni recovered from poultry, swine and laboratory isolates. A 2-step extraction process of bacterial DNA from poultry feces was developed in which the cells were first concentrated using a gradient-centrifugation step followed by comparison of 4 DNA extraction methods. Two commercial DNA extraction methods (Zymo Research Quick DNA, and Invitrogen magnetic separation), a traditional phenol-chloroform DNA extraction method using proteinase K to inactivate DNAses, and an in-house isolation method for DNA extraction based on chaotropic salts were used. The middle gradient layer recovered 89% to 98% of the bacteria cells from the sample, with recovery dependent upon the Campylobacter genus. The 4 DNA extractions methods recovered 112 to 302 ug/nL of DNA. Finally, the qPCR and standard plate methods were highly correlated for enumerating Campylobacter spp. in the 2.0 to 8.0-log CFU range. Analyses of the results from this study demonstrate that the combination of the standard curve for Campylobacter spp. DNA primers, the gradient cell concentration method and DNA extraction techniques with qPCR can be used to enumerate Campylobacter spp. from poultry samples with findings similar those of traditional plate count methodology.

The ghost bat (Macroderma gigas) is the largest microbat in Australia and occupies a large but contracting range. The ghost bat is listed as Vulnerable with a decreasing population trend by the IUCN Red List of Threatened Species. They occupy caves, disused mine adits and rock crevices as daytime roosts but distinguishing which are preferred as maternal roosts is challenging as sampling is difficult, and bats are easily disturbed while roosting. Identification of maternal roosts is a priority for conservation and management purposes, and therefore non-invasive hormone analysis was investigated as a potential tool for the future identification of the cave preferences of pregnant individuals. To validate fecal progesterone metabolite analysis techniques, fecal samples were collected from group housed female ghost bats at Perth Zoo between October and January, during the expected parturition period, over three years. Fecal samples were weighed (0.025 g), extracted with 2.5 ml of 80% methanol and analyzed for progesterone metabolite levels by enzyme-immunoassay. Significant elevations in fecal progesterone metabolite levels were detected in a subset of samples collected from co-housed females prior to parturition but not after, providing biological validation of the hormone analysis techniques. Over the three years, four pups were born with birth dates ranging from early November to late December. The mean fecal progesterone metabolite levels of non-pregnant females was significantly lower than those of pregnant females (206.5 ± 102.9 ng/g and 7003.6 ± 6078.0 ng/g respectively). These techniques can be used to monitor and evaluate the reproductive health of ghost bat populations over time and help identify factors influencing maternity roost fidelity. This data will provide valuable information for the conservation and management of the ghost bat and has the potential to be applied to other bat species.

To date, microplastic is ubiquitously encountered in the environment. Studies analyzing microplastic in terrestrial ecosystems, including animal feces and feed, are few. Microplastic quantification method validation and harmonization are not yet far developed. For the analysis of small microplastic, approximately

The gut microbiome plays a crucial role in maintaining animal health and is influenced by various factors, including light exposure; however, the response in laying hens of the gut microbiome to intermittent light regimes and the related impact on antibiotic resistance genes (ARGs) remain poorly understood. In this study, we divided 20-week-old laying hens into two groups. These groups were exposed to either continuous normal light or intermittent light for 8 weeks. The feces and cecal contents of laying hens were collected for analysis. Metagenomic analysis of both feces and cecal content samples revealed significant shifts in the microbial composition and abundance of ARGs under intermittent light exposure compared to normal light exposure (P < 0.05). Furthermore, metabolomic analysis of the cecal contents revealed substantial alterations in the abundance and composition of ARGs and mobile genetic elements (MGEs) in response to intermittent light exposure (P < 0.05). Network analysis revealed intricate co-occurrence patterns among bacterial communities, metabolites, and ARGs, highlighting correlations between Bacteroidetes species, ARGs, and metabolites. Although certain bacterial species showed differential associations, the dominant bacteria carrying ARGs or MGEs had relatively low numbers, suggesting that other bacterial communities may have had a greater influence on ARG dissemination. Moreover, our observations highlight the crucial role of metabolites as mediators between bacterial communities and ARGs, providing novel insights into the dynamics of antibiotic resistance development. Our findings underscore the impact of intermittent light exposure on ARG proliferation in poultry farming and emphasize interconnections among ARGs, bacterial communities, and metabolic pathways. The results underscore the importance of considering both microbial communities and metabolic processes to understand antibiotic resistance in agricultural settings., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Little is known about exposure of humans and companion animals to liquid crystal monomers (LCMs), which are extensively used in digital displays. We determined the concentrations of 52 LCMs in feces of humans, pet dogs and cats from New York State, USA, using gas chromatography-high resolution mass spectrometry (GC-HRMS). Twenty-four, eight, and six LCMs, that were mainly fluorinated, were detected in human, dog, and cat feces, respectively. ∑LCMs concentrations in the feces of humans (mean: 8.01 ng/g dry weight [dw]) were significantly higher (p < 0.05) than those of dogs (mean: 1.82 ng/g dw) and cats (mean: 1.24 ng/g dw) and with concentrations measured as high as 39.8 ng/g dw. Rel-4'-((1r,1'r,4 R,4'R)-4'-ethyl-[1,1'-bi(cyclohexan)]-4-yl)-3,4-difluoro-1,1'-biphenyl (RELEEBCH or 2bcHdFB) was found at the highest detection frequency (DF) among LCMs analyzed in human (DF: 89 %), dog (DF: 28 %), and cat (DF: 50 %) feces, although this compound accounted only < 4 % of ∑LCM concentrations. The mean cumulative daily intakes of ∑LCMs, calculated through a reverse dosimetry approach, were 71.7, 87.5, and 10.7 ng/kg body weight (bw)/day for humans, dogs, and cats, respectively. This study provides evidence of exposure of both humans and pets to LCMs, highlighting the importance of assessing sources of exposure and associated health risks., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Microplastics (MPs) are found in a wide range of ecosystems, from the Arctic to the deep ocean. However, there is no data on their presence in terrestrial mammals that inhabit the Selva Maya. The aim of this study is to detect the presence of MPs in the feces of the Baird's tapir (Tapirus bairdii) from the region of Calakmul, located in the Yucatan Peninsula, Mexico. We analyzed 129 fecal samples collected during 2017 and 2018, obtaining 57 and 72 samples during the rainy and dry seasons respectively. Sixty-eight percent of the samples contained 743 MPs with a mean of 19.3 ± 28.1 MPs/kg of dry weight (DW) feces in both years. An inter-annual variation in the average abundance of microplastic was observed during the two-year period (2017-2018), with a 72 % increase in these plastic particles in feces. Fourteen polymers were identified, with ethylene vinyl acetate (EVA), polypropylene (PP) and polyester (PES) being the most abundant during both years. Although the effects of MPs on the health of tapirs are not known, their presence is cause for concern. There is an urgent need for the implementation of appropriate plastic waste management programs in communities of the Selva Maya to diminish the consumption of MPs in species including humans where they pose a significant risk to health. ENVIRONMENTAL IMPLICATIONS: The use of plastics worldwide is increasing every day, so the presence of microplastics is and will continue to be a major environmental problem. It is known that contaminants can adhere to plastics, making them hazardous materials. Microplastics can contaminate remote areas such as Biosphere Reserves. Terrestrial species such as the tapir can ingest microplastics, putting their health at risk. Knowing the dispersion of microplastics is very important in order to manage them properly, taking into account their emission sources and type of polymer., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Trichuris spp. are nematode parasites infecting wild ruminants in zoological institutions worldwide. These helminths cause significant morbidity in giraffe (Giraffa camelopardalis) and other hoofstock located in zoological institutions throughout the United States. Historically, studies and institutions have used a variety of nematode detection methods with various flotation solutions. Optimization of Trichuris egg detection is necessary for monitoring collections. Fecal and soil optimized protocols were generated in this study using samples containing Trichuris eggs from multiple semi free-ranging zoological institutions. First, Sheather's sugar (specific gravity (SG) 1.27), sucrose (SG 1.40), magnesium sulfate (SG 1.26), and zinc sulfate (SG 1.18) were compared as flotation solutions by quantitative eggs per gram using a modified Stoll method. Then a soil recovery method was optimized comparing Tween 20, sodium hydroxide, Dawn™ (Procter and Gamble) detergent, and sodium chloride as liberating solutions to free eggs from the soil. We found that Sheather's sugar and sucrose solutions were the most effective for Trichuris egg detection, and either sodium hydroxide or sodium chloride liberated eggs from soil.

Fecal waste is a significant source of antimicrobial resistance (AMR) pollution and provides valuable insights into the AMR development in animal and human populations within the "One health" framework. Various genetic elements, including antibiotic resistance genes (ARGs), biocide and metal resistance genes (BMGs), mobile genetic elements (MGEs), and virulence factor genes (VFGs), are crucial AMR risk determinants (ARDs). However, few studies focused on compositional characteristics of ARDs in different feces. Here, we analyzed 753 public metagenomes from human, pig, chicken, and cattle feces, revealing significant differences in ARD richness and abundance across fecal types, notably lowest in cattle samples. Tetracycline, multi-metal, and -biocide resistance genes were dominant resistome. A few core genes contributed to 25.6%-91.1% of gene abundance, and their correlations were stronger in cattle samples. Procrustes analysis showed that microbial composition had higher correlations with ARGs (M 2  = 0.579) and BMGs (M 2  = 0.519). Gammaproteobacteria was identified as major ARD-hosts especially in human and pig feces, and they mainly carried multi-resistance genes. MGEs exhibited direct positive effects on ARGs and BMGs, indirectly impacting VFGs. Utilizing random forest methods, we identified 42 indicator genes for tracking AMR pollution originating from fecal sources in the environments. This study offers new insights into understanding and controlling the AMR pollution of fecal waste from human and food animals., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Lassa arenavirus (LASV) is the cause of Lassa Fever in humans in West Africa. The multimammate mouse (Mastomys natalensis) is a reservoir host of LASV and the primary source of human infections. Humans are assumed to become infected due to contact with this animal or its excretions. Thus far, the available literature does not describe the sampling of feces as a means to detect LASV in M. natalensis populations. More evidence is needed to know if feces of naturally infected M. natalensis can be LASV-positive and an exposure risk to humans. This study sampled feces deposits in households from three villages in the LASV-endemic region of Faranah, Guinea. PCR analysis found 10 out of 88 samples to be positive for LASV, and sequencing showed clustering to previously identified Yarawelia and Dalafilani strains. We conclude that feces sampling is a viable, non-invasive method for the determination and sequencing of LASV strains.

Globally, safe sanitation has improved significantly in the last two decades, but unsafe child feces disposal remains a growing challenge in many regions, exposing household members and communities to infectious pathogens. The drivers associated with child feces disposal in several contexts including humanitarian settings are not well understood. This study investigated child feces disposal (CFD) practices and associated factors in low- and middle-income countries, including in humanitarian settings. Data from 352,173 women in 34 countries, collected between 2012 and 2021 through Demographic and Health Surveys (DHS), were used. We utilized multivariate logistic regression to assess CFD practices among children under two years old and the factors linked to these practices. We incorporated data from the United Nations High Commissioner for Refugees (UNHCR) regarding refugee camps' locations in the analysis. Time series and local spatial autocorrelation analyses were run to examine changes in safe CFD practices over time and space, respectively. Results showed minimal improvement in safe child feces disposal over the past decade, with 55.6 % of respondents in non-humanitarian settings and 38.1 % in humanitarian settings improperly disposing of feces. Improper CFD significantly correlated with increased odds of diarrhea in non-humanitarian settings (OR 1.09 95 % CI: 1.05-1.13) but not in humanitarian settings (OR 1.14 95 % CI: 0.53-2.49). The most significant factors (p < 0.05) associated with safe CFD included being in the richest wealth quintile (OR 3.27 95 % CI: 3.06-3.49), having basic education (OR 1.28 95 % CI: 1.22-1.33), children eating solid food (OR 1.53 95 % CI: 1.48-1.57), improved sanitation access (OR 1.88 95 % CI: 1.81-1.96), and listening to radio at least weekly (OR 1.40 95 % CI: 135-1.46). Policymakers and development partners must include safe CFD guidelines in national policies and programs, as well as prioritize investments in household-level sanitation and educate caregivers about safe CFD practices., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Diarrheagenic E. coli (DEC) can cause severe diarrhea and is a public health concern worldwide. Cattle are an important reservoir for this group of pathogens, and once introduced into the abattoir environment, these microorganisms can contaminate consumer products. This study aimed to characterize the distribution of DEC [Shiga toxin-producing E. coli (STEC), enteroinvasive E. coli (EIEC), enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), and enteroaggregative E. coli (EAEC)] from extensive and intensive cattle production systems in Brazil. Samples (n = 919) were collected from animal feces (n = 200), carcasses (n = 600), meat cuts (n = 90), employee feces (n = 9), and slaughterhouse water (n = 20). Virulence genes were detected by PCR in 10% of animal samples (94/919), with STEC (n = 81) as the higher prevalence, followed by EIEC (n = 8), and lastly EPEC (n = 5). Animals raised in an extensive system had a higher prevalence of STEC (average 48%, sd = 2.04) when compared to animals raised in an intensive system (23%, sd = 1.95) (Chi-square test, P < 0.001). From these animals, most STEC isolates only harbored stx2 (58%), and 7% were STEC LEE-positive isolates that were further identified as O157:H7. This study provides further evidence that cattle are potential sources of DEC, especially STEC, and that potentially pathogenic E. coli isolates are widely distributed in feces and carcasses during the slaughter process., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Using waste from sewage systems, particularly human excreta, could save resources and increase soil fertility, contributing to nutrient management. However, because of the pathogenic content in human feces, this resource can pose health risks to farmers and consumers. Therefore, this work analyzed the behavior of the microorganisms: Escherichia coli ATCC13706 and human adenovirus (HAdV-2) in the soil and the internal part of the plant tissue during the vegetative stage after applying spiked composted human feces as biofertilizer. In a greenhouse, we simulated the application of the biofertilizer in lettuce cultivation by spiking three concentrations of E. coli (6.58, 7.31, and 8.01 log 10 CFU.g -1 ) and HAdV-2 (3.81, 3.97, and 5.92 log 10 PFU.g -1 ). As a result, we achieved faster decay in soil at higher concentrations of E. coli. We estimated linear decay rates of -0.07279, -0.09092, and -0.115 days, corresponding to T 90 s of 13.7, 11.0, and 8.6 days from higher to smaller concentrations of E. coli, respectively. The estimated periods for the inactivation of 4 logarithmic units of E. coli bacteria in soil are longer than the cultivation period of lettuce for all concentrations studied. Concerning the bacterial contamination in plants, we found E. coli in the internal part of the leaves at the highest concentration tested during the first three weeks of the experiment. Furthermore, HAdV-2 was found in roots at a stable concentration of 2-2.3 log 10 PFU.g -1 in five of the six samples analyzed. Therefore, bacterial infection could pose a risk, even if fresh greens are washed before consumption, especially for short-term cultures. Regarding viral infection, a positive result in the roots after disinfection may pose a risk to root and tubercule vegetables. These discoveries highlight the importance of conducting comprehensive evaluations of hygiene practices in incorporating organic amendments in crops, explicitly aiming to minimize the risk of post-contamination., Competing Interests: Declaration of competing interest The authors declare that none of their known personal or financial conflicts could have possibly affected the research described in this work., (Copyright © 2024 Elsevier B.V. All rights reserved.)

During hospitalization horses may develop gastrointestinal conditions triggered by a stress-associated weak local immune system. The prospective, clinical trial was conducted to find out whether fecal immunoglobulin A (IgA) concentrations could be determined in hospitalized horses and how they changed during hospitalization and in response to various stressors. Samples were obtained from 110 horses and a control group (n = 14). At arrival in the hospital, horses were categorized into pain grades (1-5), and elective versus strenuous surgery (> 2 hours, traumatic and emergency procedures). Feces were collected on day 1, day 2, day 3, and day 7 in all horses. Blood samples were obtained at the same intervals, but additionally after general anaesthesia in horses undergoing surgery (day 2). IgA concentration in feces was determined by ELISA and measured in optical density at 450nm. The control group showed constant IgA concentrations on all days (mean value 0.30 OD 450 ±SD 0.11, 1.26 mg/g; n = 11). After general anaesthesia fecal IgA concentrations decreased considerably independent of duration and type of surgery (P < 0.001 for elective and P = 0.043 for traumatic surgeries). High plasma cortisol concentrations were weakly correlated with low fecal IgA on the day after surgery (P = 0.012, day 3, correlation coefficient r = 0.113). Equine fecal IgA concentrations showed a decline associated with transport, surgery, and hospitalization in general, indicating that stress has an impact on the local intestinal immune function and may predispose horses for developing gastrointestinal diseases such as enterocolitis., Competing Interests: Declaration of competing interest None of the authors has any financial or personal relationships that could inappropriately influence or bias the content of the paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Enterococci are widely distributed in dairy sector. They are commensals of the gastrointestinal tract of animals, thus, via fecal contamination, could reach raw milk and dairy products. The aims of this study were: 1) to investigate the enterococcal diversity in cow feces and milk samples and 2) to evaluate the antibiotic resistance (AR) of dairy-related enterococci and their ability to transfer resistance genes. E. faecalis (59.9%), E. faecium (18.6%) and E. lactis (12.4%) were prevalent in milk, while E. faecium (84.2%) and E. hirae (15.0%) were dominant in bovine feces. RAPD-PCR highlighted a high number of Enterococcus biotypes (45 from milk and 37 from feces) and none of the milk strains exhibited genetic profiles similar to those of feces biotypes. A high percentage of enterococci isolated from milk (71%) were identified as multidrug resistant and resistance against streptomycin and tetracycline were widespread among milk strains while enterococci from feces were commonly resistant to linezolid and quinupristin/dalfopristin. Only E. faecalis strains were able to transfer horizontally the tetM gene to Lb. delbrueckii subsp. lactis. Our results indicated that Enterococcus biotypes from milk and bovine feces belong to different community and the ability of these microorganisms to transfer AR genes is strain-dependent., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

This research investigated the variations in the occurrence of Salmonella, STEC O157:H7 and non-O157 in the beef production chain in Colombia affected by seasons, hypothesizing that pathogen prevalence will be highest in the rainy season owing to soil moisture promoting bacteria multiplication and transfer between animals. To test this hypothesis, samples were obtained from five abattoirs, which represent 50% of the beef production in this country. A total of 1017 samples were collected, from which 606 were bovine feces, 206 were hide swabs, and 205 corresponded to carcass post-intervention. From the 1017 samples, 49.9% (n = 507) were collected during dry season, while 50.1% (n = 510) during rainy season. All samples (n = 1017) underwent screening for E. coli O157:H7 and Salmonella, while only a proportion of fecal samples (n = 339) were screened for the big six STEC serogroups and their virulence markers. The effect of season, age of animal and sex of animal were correlated with the prevalence results. A total of 84.7% of fecal samples carried virulence genes associated to STEC (stx or eae), suggesting that testing and control should be increased for the big-six STEC compared to E. coli O157:H7. Pathogen prevalence in feces was found to be 8.3%, 5.0%, and 51.0% for Salmonella, E. coli O157:H7 and STEC non-O157, respectively. Hides had a prevalence of 15.0% and 6.8% of Salmonella and E. coli O157:H7, respectively. Carcasses post-intervention were found to have 4.4% and 2.5% prevalence of Salmonella and E. coli O157:H7, respectively. A seasonal effect was found for fecal samples. E. coli O157 and non-O157 STEC shedding were significantly higher (P ≤ 0.05) during rainy season compared to dry season. In contrast, hides and carcasses were more likely to present lower incidence of pathogens during rainy months compared to dry season; however, it was significant only for Salmonella on carcasses with estimated odds of detection almost six times higher in the dry season relative to the rainy season (OR = 5.90, 95% CI 1.18–29.57).

Measuring bile acids in feces has an important role in disease prevention, diagnosis, treatment, and can be considered a measure of health status. Therefore, the primary aim was to develop a sensitive, robust, and high throughput liquid chromatography tandem mass spectrometry method with minimal sample preparation for quantitative determination of bile acids in human feces applicable to large cohorts. Due to the chemical diversity of bile acids, their wide concentration range in feces, and the complexity of feces itself, developing a sensitive and selective analytical method for bile acids is challenging. A simple extraction method using methanol suitable for subsequent quantification by liquid chromatography tandem mass spectrometry has been reported in, “Extraction and quantitative determination of bile acids in feces” [1]. The data highlight the importance of optimization of the extraction procedure and the stability of the bile acids in feces post-extraction and prior to analysis and after several freeze-thaw cycles.

Waste from infected individuals can contaminate the soil and water of the community without adequate sanitation facilities, increasing the risk of infection for other individuals. The objective of this study was to assess child feces disposal status and associated factors in community-led total sanitation (CLTS) adopted districts of the Jimma Zone, Ethiopia. This study analyzed comparative cross-section secondary data from 756 households systematically selected in two chosen districts (Kersa and Omo Nadda) of Jimma Zone, Oromia Regional State, Ethiopia in 2018 by our research team. The data used was obtained by interviewing mothers/caregivers using a pre-tested structured questionnaire. About 80% of mothers/caregivers of CLTS adopted kebeles (the smallest administrative structure) and 58.7% of non-CLTS kebeles had practiced safe child feces disposal practices. Compared to their counterparts, the use of unimproved latrine form (AOR=0.45; 95% CI: 0.24–0.83) and location of drinking water storage containers elevated above the floor (OR=0.02; 95% CI: 0.05–0.88) had a negative association with safe disposal practices of child feces. However, among mothers/caregivers indexed as medium (AOR=1.64; 95% CI: 1.03–2.61), rich (AOR=2.17; 95% CI: 1.29–3.66), live with children under five vulnerable to diarrhea (AOR= 1.54; 95% CI: 1.02–2.52) and owned toilet without a handwashing facility (AOR= 2.44; 95% CI: 1.49–3.98), the likelihood of safe child feces disposal practices was increased. In conclusion, our study shows that the study participants' safe child feces disposal status was comparatively higher in CLTS adopted kebeles, but the difference was not significant in the final model.

Black soldier fly larvae (BSFL) have great potential in livestock manure disposal. However, the changes in metal speciation, microbial communities, potential pathogens during the manure transformation process by BSFL is still largely uncharacterized, as well as the underlying metal tolerance mechanism of larval gut microbiome. Here we used BSFL to convert pig manure (PM) into larval feces (BF), and investigated the metal and microbial changes in the conversion process. Physicochemical parameters (e.g. pH, electrical conductivity, total nitrogen, total phosphorus and total potassium) in PM were significantly altered compared to BF. After conversion, less than 10% of Cu and Zn were accumulated in larval bodies. The bioavailable fraction of Cu (88.3%–86.2%) and Zn (80.6%–82.3%) occupied as the primary form in PM and BF. Genera Enterococcus, Clostridium_sensu_stricto_1, Terrisporobacter and Romboutsia were substantially enriched in the final BSFL gut (GF) compared with initial gut (GI). BSFL transformation substantially reduced pathogen abundances (decreased by 89%) derived from pig manure. Functional genes involved in metal homeostasis and resistance (e.g. CutC, pcoC, cusR, zurR and zntB) were obviously strengthened (by 2.3–7.7 folds) in GF than in GI, which might partly explain the metal tolerance ability of BSFL during the livestock manure transformation process.

Sapovirus (SaV) is a nonenveloped RNA virus that causes acute gastroenteritis in humans. Although SaV is a clinically important pathogen in children, an effective vaccine is currently unavailable. The capsid protein VP1 of SaVs forms the outer shell of the virion and is highly diverse, as often seen in the virion-surface proteins of RNA viruses, creating an obstacle for vaccine development. We here report a unique phenomenon pertaining to the variation of SaV VP1. Phylogenetic and information entropy analyses using full-length VP1 sequences from a public database consistently showed that the amino acid sequences of the VP1 protein have been highly conserved over more than 40 years in the major epidemic genotype GI.1 but not in GI.2. Structural modeling showed that even the VP1 P2 subdomain, which is arranged on the outermost shell of the virion and presumably exposed to anti-SaV antibodies, remained highly homogeneous in GI.1 but not in GI.2. These results suggest strong evolutionary constraints against amino acid changes in the P2 subdomain of the SaV GI.1 capsid and illustrate a hitherto unappreciated mechanism, i.e., preservation of the VP1 P2 subdomain, involved in SaV survival. Our findings could have important implications for the development of an anti-SaV vaccine., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

This research investigated the in-situ decay rates of four human wastewater-associated markers (Bacteroides HF183 (HF183), Lachnospiraceae Lachno3 (Lachno3), cross-assembling phage (crAssphage), pepper mild mottle virus (PMMoV) and three enteric viruses (human adenovirus 40/41 (HAdV 40/41), enterovirus (EV) and human norovirus GII (HNoV GII) in two estuarine water environments (Davidson Park (DP) and Hen and Chicken Bay (HCB) in temperate Sydney, NSW, Australia, employing qPCR and RT-qPCR assays. The study also aimed to compare decay rates observed in mesocosms with previously published laboratory microcosms, providing insights into the persistence of markers and viruses in estuarine environments. Results indicated varying decay rates between DP and HCB mesocosms, with HF183 exhibiting relatively faster decay rates compared to other markers and enteric viruses in sunlight and dark mesocosms. In DP mesocosms, HF183 decayed the fastest, contrasting with PMMoV, which exhibited the slowest. Sunlight induced higher decay rates for all markers and viruses in DP mesocosms. In HCB sunlight mesocosms, HF183 nucleic acid decayed most rapidly compared to other markers and enteric viruses. In dark mesocosms, crAssphage showed the fastest decay, while PMMoV decayed at the slowest rate in both sunlight and dark mesocosms. Comparisons with laboratory microcosms revealed faster decay of markers and enteric viruses in laboratory microcosms than the mesocosms, except for crAssphage and HAdV 40/41 in dark, and PMMoV in sunlight mesocosms. The study concludes that decay rates of markers and enteric viruses vary between estuarine mesocosms, emphasizing the impact of sunlight exposure, which was potentially influenced by the elevated turbidity at HCB estuarine waters. The generated decay rates contribute valuable insights for establishing site-specific risk-based thresholds of human wastewater-associated markers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Recent studies have highlighted the presence of antibiotic resistance genes (ARGs) in Antarctica, which are typically indicative of human activity. However, these studies have concentrated in the Antarctic Peninsula region, and relatively less is known about ARG prevalence in East Antarctica, where human activity levels are lower compared to the Antarctic Peninsula. In addition, the mechanisms of ARG transmission to Antarctica through natural or anthropogenic pathways remain unclear. In this study, we analyzed the fecal samples of Adélie penguins and South polar skuas by using high-throughput sequencing and microfluidic quantitative PCR to detect potential pathogens and ARGs at their breeding colonies near Syowa Station in East Antarctica. These results revealed the presence of several potential pathogens in the fecal matter of both bird species. However, the HF183 marker, which indicates human fecal contamination, was absent in all samples, as well as seawater sampled near the breeding colonies. This suggests that the human fecal contamination was negligible in our study area. In addition to pathogens, we found a significant number of ARGs and metal resistance genes in the feces of both Adélie penguins and South polar skuas, with higher detection rates in skuas than in penguins. To better understand how these birds acquire and transmit these genes, we analyzed the migratory patterns of Adélie penguins and South polar skuas by geolocator tracking. We found that the skuas migrate to the tropical and subtropical regions of the Indian Ocean during the austral winter. On the other hand, Adélie penguins exhibited a more localized migration pattern, mainly staying within Antarctic waters. Because the Indian Ocean is considered one of the major reservoirs of ARGs, South polar skuas might be exposed to ARGs during their winter migration and transfer these genes to Antarctica., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Early-life stress (ELS) has been robustly associated with a range of poor mental and physical health outcomes. Recent studies implicate the gut microbiome in stress-related mental, cardio-metabolic and immune health problems, but research on humans is scarce and thus far often based on small, selected samples, often using retrospective reports of ELS. We examined associations between ELS and the human gut microbiome in a large, population-based study of children. ELS was measured prospectively from birth to 10 years of age in 2,004 children from the Generation R Study. We studied overall ELS, as well as unique effects of five different ELS domains, including life events, contextual risk, parental risk, interpersonal risk, and direct victimization. Stool microbiome was assessed using 16S rRNA sequencing at age 10 years and data were analyzed at multiple levels (i.e. α- and β-diversity indices, individual genera and predicted functional pathways). In addition, we explored potential mediators of ELS-microbiome associations, including diet at age 8 and body mass index at 10 years. While no associations were observed between overall ELS (composite score of five domains) and the microbiome after multiple testing correction, contextual risk - a specific ELS domain related to socio-economic stress, including risk factors such as financial difficulties and low maternal education - was significantly associated with microbiome variability. This ELS domain was associated with lower α-diversity, with β-diversity, and with predicted functional pathways involved, amongst others, in tryptophan biosynthesis. These associations were in part mediated by overall diet quality, a pro-inflammatory diet, fiber intake, and body mass index (BMI). These results suggest that stress related to socio-economic adversity - but not overall early life stress - is associated with a less diverse microbiome in the general population, and that this association may in part be explained by poorer diet and higher BMI. Future research is needed to test causality and to establish whether modifiable factors such as diet could be used to mitigate the negative effects of socio-economic adversity on the microbiome and related health consequences., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Septic tank systems (STS) in rural catchments represent a potential source of microbial pollution to watercourses; however, data concerning the risk of faecal indicator organism (FIO) export from STS to surface waters are scarce. In the absence of empirical data, elicitation of expert judgements can provide an alternative approach to aid understanding of FIO pollution risk from STS. Our study employed a structured elicitation process using the Sheffield Elicitation Framework to obtain expert judgements on the proportion of FIOs likely to be delivered from STS to watercourses, based on 36 scenarios combining: (i) septic tank effluent movement risk, driven by soil hydro-morphological characteristics; (ii) distance of septic tank to watercourse; and (iii) degree of slope. Experts used the tertile method to elicit a range of values representing their beliefs of the proportion of FIOs likely to be delivered to a watercourse for each scenario. The experts judged that 93 % of FIOs would likely be delivered from an STS to a watercourse under the highest risk scenario that combined (i) very high STS effluent movement risk, (ii) STS distance to watercourse <10 m, and (iii) a location on a steep slope with gradient >25 %. Under the lowest risk scenario, the proportion of FIOs reaching a watercourse would likely reduce to 5 %. Expert confidence was high for scenarios that represented extremes of risk, while uncertainty increased for scenarios depicting intermediate risk conditions. The behavioural aggregation process employed to obtain a consensus among the experts proved to be useful for highlighting both areas of strong consensus and high uncertainty. The latter therefore represent priorities for future empirical research to further improve our understanding of potential pollution risk from septic tanks and in turn enable better assessments of potential threats to water quality in rural catchments throughout the world where decentralised wastewater systems are common., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Outdoor defecation by people experiencing homelessness is frequently perceived as a potentially large source of human fecal pollution and a significant source of health risk in urban waterbodies with recreational contact. The goal of this study was to count the number of people experiencing homelessness and quantifies their sanitation habits in an urban river corridor setting, then use this information for estimating human fecal pollutant loading on a watershed scale. Two types of census counts were conducted including periodic point-in-time counts over six years and weekly counts of encampments. While the population census varied from count-to-count, the range of population estimates in the river corridor varied from 109 to 349 individuals during the six-year span, which mirrored the weekly counts of encampments. A face-to-face survey of people experiencing homelessness assessed the sanitation habits of the unsheltered population (N = 63), including outdoor defecation frequency and containment practices. Overall, 95 % of survey respondents reported defecating outdoors; 36 % practiced outdoor defecation between 4 and 7 days/week and 27 % practiced outdoor defecation <1 day/week. Of those that did practice outdoor defecation, 75 % contained their feces in a bucket or bag, thereby limiting fecal material contributions to the river; 6.7 % reported defecating on low ground near the river that could wash off when flood waters rise during a storm event. Only a single survey respondent reported defecating directly into the river. Based on literature values for average HF183 output for an adult human, and the average rainfall in the urban watershed, the total watershed contribution of HF183 averaged 1.2 × 10 10 gene copies per storm event (95 % CI: 0.9 × 10 10 -1.6 × 10 10 ) along the 41 km stretch of river in this study. This human fecal loading estimate is at least two orders of magnitude less than cumulative HF183 loading from all human sources measured at the bottom of the watershed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The flushing of wet wipes down toilets leads to blockages of sewerage systems. This, together with unregulated sewage discharge, often results in increasing numbers of wet wipes washing up onto beaches. However, it is unclear how long wet wipes can persist on the beach and whether they pose a prolonged public health risk if contaminated by E. coli. In this mesocosm study, three types of wet wipes (plastic containing, and home and commercially compostable) colonised with E. coli were buried in beach sand and their degradation, tensile strength, and concentration of E. coli was quantified over 15 weeks. Wet wipes containing plastic remained largely intact for 15 weeks, whilst both compostable wet wipes fragmented and degraded. Importantly, E. coli persisted on all three wet wipe types, representing localised reservoirs of E. coli in the sand, which could present a human health risk at the beach., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Enterocytozoon bieneusi is one of the 17 microsporidian species pathogenic to humans in low and high-income countries, inducing both symptomatic and asymptomatic intestinal infections, independently of the immunological condition of the infected individual. Faecal-oral transmission occurs in a broad hosts range, including several animal species, but the parasite's zoonotic potential remains still unclear. Few studies are available in Italy regarding E. bieneusi presence in humans and no data on its genetic variability are so far reported. In this investigation, through the ITSr RNA sequences analysis, we provided the first E. bieneusi molecular characterization from symptomatic patients in Italy. Faecal samples from 410 patients sent for routine analyses to the Unit of Parasitology, Policlinico Tor Vergata, Rome, and resulted positive for E. bieneusi to a cartridge-based molecular test for qualitative detection (Novodiag® Stool Parasites assay), were collected. DNA was extracted, endpoint PCR performed and then sequences obtained for 3/410 patients (0.7 %). Genotype A (N = 1), genotype C (N = 1) and genotype K (N = 1) were identified, all belonging to phylogenetic Group 1. One patient (identified as genotype A) showed positivity to the same genotype previously characterized after a two-month period. Additional investigations are required, within a One Health framework, to review the importance of a zoonotic potential linked to E. bieneusi in human populations, animals and environmental reservoirs worldwide., Competing Interests: Declaration of competing interest The Authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Goat coccidiosis compromises animal welfare, reduces productivity and may cause mortality and delayed growth rates in goat kids around the weaning period worldwide. This field study was conducted to evaluate the efficacy of metaphylactic treatments with two doses of toltrazuril (20 or 40 mg/kg body weight - BW, p. o.), at different timing, in kids naturally infected with Eimeria spp. A total of 97 healthy goat kids (Majorera milk aptitude breed) were divided into five groups, depending on the age of treatment (2 or 7 weeks). One group remained untreated as a negative control until the end of the study. Faecal oocyst shedding, faecal consistency, and body weight of the animals were monitored at day 0 and at weekly intervals. Counts of oocysts per gram of faeces (OPG) were determined by a modified McMaster technique. Morphometric identification of Eimeria species was carried out on individual faecal samples from each experimental group after oocyst sporulation. Goat kids treated at two weeks of age maintained OPG values close to zero during the 5 weeks post-treatment and, overall, had lower faecal oocyst counts than untreated control animals. No significant differences were observed between the two doses of toltrazuril used in two-week-old treated animals. By contrast, when treatment was carried out at seven weeks of age, the dose of 40 mg/kg BW of toltrazuril reduced oocyst levels for longer and to a greater extent than the 20 mg/kg dose. Irrespectively of the treatment and dose, toltrazuril delayed the appearance of pathogenic Eimeria species, i. e. Eimeria ninakohlyakimovae and Eimeria arloingi. As a whole, Eimeria christenseni, with a rather moderate pathogenicity, was highly predominant throughout the study period, including the untreated control group, which was probably the reason why clinical signs of coccidiosis were barely observed throughout the experiment. Under these circumstances, the positive effect of toltrazuril on body weight condition observed in some treated groups was difficult to correlate to the timing and doses. Metaphylactic treatments with 20 mg/kg BW toltrazuril given at two weeks of age are sufficient to control oocyst excretion in goat kids; whereas if administered later in 7-week-old animals, thereby coinciding with the frequently observed peak of oocyst elimination in goat kids under field conditions, a higher dose might be advisable to prevent environmental contamination with infectious oocysts., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The objective of this study was to compare preharvest monitoring strategies by evaluating three different sampling methods in the lairage area to determine pathogen recovery for each sampling method and incoming pathogen prevalence from the cattle to inform in-plant decision making. Samples were gathered over a 5-month period, from February to June 2022, at a harvesting and processing facility located in Eastern Nebraska. Sampling methods included (i) fecal pats, (ii) boot swabs, and (iii) MicroTally swab. A total of 329 samples were collected over the study period (fecal pats: n = 105, boot swabs: n = 104, and MicroTally swabs: n = 120). Specific media combinations, an incubation temperature of 42°C, and incubation timepoints (18-24 h) were utilized for each matrix and the prevalence of Salmonella, Escherichia coli O157:H7, and six non-O157 Shiga-toxin producing E. coli (STEC) was evaluated using the BAX system Real-Time PCR assay. Overall, results from the study concluded that boot swabs were an effective sampling method for pathogen detection in the cattle lairage area. Boot swabs (97.1%) were statistically more likely to detect for Salmonella (p < 0.05) when compared to fecal pats (67.6%) and MicroTally swab (77.5%) methods. For E. coli O157:H7 and STEC - O26, O121, O45, and O103 prevalence, boot swabs were significantly better at detecting for these pathogens (p < 0.05) than MicroTally swabs (OR = 3.16 - 11.95) and a comparable sampling method to fecal pats (OR = 0.93 - 2.01, p > 0.05). Lastly, all three sampling methods detected a very low prevalence for E. coli O111 and O145; therefore, no further analysis was conducted. The boot swab sampling method was strongly favored because they require little training to implement, are inexpensive, and they do not require much sampling labor; therefore, would be a simple and effective sampling method to implement within the industry to evaluate pathogen prevalence preharvest., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Fasciola hepatica causes liver fluke disease in production animals and humans worldwide. Faecal egg counts (FEC) are the most common diagnostic tool for the diagnosis of liver fluke disease. However, FEC has low sensitivity and is often unreliable for the detection of patent infection. In this study, loop-mediated isothermal amplification (LAMP) was optimised and evaluated for the detection of Fasciola hepatica infection, with the aim of increased sensitivity and making it suitable for on-farm application. LAMP was initially conducted under laboratory conditions, optimised to enable visual detection using calcein dye. DNA extraction based on bead-beating was developed to enable on-farm application. LAMP results were compared to FEC and polymerase chain reaction (PCR). Under laboratory conditions, LAMP was conducted using two incubation methods: a conventional PCR thermocycler and a field-deployable LAMP instrument. When compared to a 'rigorous' FEC protocol consisting of multiple counts using a comparatively large volume of faeces and with infection confirmed post-mortem, LAMP was highly sensitive and specific (using silica membrane DNA extraction sensitivity 88 %, specificity 100 %; using sieving and beat-beating DNA extraction sensitivity 98.9 %, specificity 100 %). When applied on-farm, LAMP was compared to conventional FEC, which suggested high sensitivity but low specificity (sensitivity 97 %, specificity 37.5 %). However, further analysis, comparing field LAMP results to laboratory PCR, suggested that the low specificity was likely the outcome of the inability of conventional FEC to detect all true F. hepatica positive samples. Based on the high sensitivity and specificity of LAMP compared to a 'rigorous' FEC protocol and its ability to be used in field settings, the study demonstrates the potential of LAMP for diagnosing F. hepatica infection in agriculture., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Tanjina Bari reports financial support was provided by Australian Government Department of Education., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

In the pigeon industry, treating and preventing diarrhea is vital because it is a serious health problem for pigeons. This study investigated the incidence of diarrhea in 3 pigeon farms in Shanghai, and analyzed the microflora through 16S rDNA high-throughput sequencing. Four strains of Escherichia coli (E. coli) isolated from pigeon diarrhea feces were administered via gavage to healthy pigeons, with each pigeon receiving 2 × 10 8 CFU. Pigeons that developed diarrhea after E. coli challenge were treated with 3 g of Lactobacillus salivarius SNK-6 (L. salivarius SNK-6) health sand (1.6 × 10 7 CFU/g). Then, a mass feeding experiment expanded to 688 pairs of pigeons with 3 replicates, each receiving 3 g of health sand containing L. salivarius SNK-6 (1.6 × 10 7 CFU/g) every 2 wk, and fecal status monitored and recorded. The study found that the relative abundance of the Lactobacillus genus and L. salivarius in feces from pigeons with diarrhea was significantly lower than in normal pigeon feces (P < 0.05). In contrast, E. coli showed a higher abundance and diversity in feces from pigeons with diarrhea than in normal feces (P < 0.05). Three out of the 4 isolated E. coli strains caused pigeon diarrhea, resulting in a significant reduction in microbial diversity in fecal samples (P < 0.05). Both the small group attack experiment and the mass-fed additive experiment in pigeon farms demonstrated that feeding L. salivarius SNK-6 effectively cured and prevented diarrhea. Pigeons fed with L. salivarius SNK-6 exhibited no diarrhea, while the control group had a 10% diarrhea rate. In summary, a deficiency of Lactobacillus or a high abundance of E. coli in the intestine could easily cause pigeon diarrhea. Feeding L. salivarius SNK-6 could treat pigeon diarrhea, and continuous supplementation could maintain stable preventive effects., (Copyright © 2024. Published by Elsevier Inc.)

Blastocystis is a ubiquitous intestinal protist in humans and animals worldwide. The traditional livestock free-roaming raising system in rural communities increases the risk of infection with contact with a wider range of pathogens transmitted via the faecal-oral route associated with that wildlife-livestock-human interface. However, no studies have been conducted to determine the occurrence and subtype distribution of Blastocystis in livestock in Portugal. Here, we collected 180 faecal samples from herbivore livestock (cattle, goats, horses, and sheep) in different regions of the country to investigate Blastocystis prevalence and subtype diversity using PCR and next-generation amplicon sequencing. Blastocystis was present in 40.6% (73/180; 95% CI: 33.31-48.11) of the samples (goats, 81.0%; sheep, 60.9%; cattle, 32.2%). None of the horse samples were Blastocystis-positive. Eighteen subtypes were detected (ST1-ST3, ST5-ST7, ST10, ST13, ST14, ST21, ST23-ST26, ST30, ST42-ST44). Mixed infections were detected in 97.3% of the Blastocystis-positive samples. Potentially zoonotic subtypes were identified in 75.0%, 96.4%, and 100% of the Blastocystis-positive specimens collected from cattle, sheep, and goats, respectively. These results demonstrate that cattle, sheep, and goats harbour a high diversity of Blastocystis subtypes in the study regions. Importantly, our data provide novel molecular evidence strongly suggesting that some Blastocystis STs/ST subgroups may have differential host specificity., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Toxoplasma gondii is a widespread and specialized intracellular protozoan pathogen that affects one third of the world' s population, posing a great threat to public health. As the definitive host, cats excrete oocysts and play a crucial role in the transmission of toxoplasmosis. The current diagnostic tools usually require bulky equipment and expertize, which hinders the efficient diagnosis and intervention of Toxoplasma infection in cats. In this study, we combined (RPA) with clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technique to establish an easier method for the detection of T. gondii oocysts in cat fecal samples. The sensitivity, specificity, and practicability of the established RPA-CRISPR/Cas9 method were evaluated using a lateral flow strip, with the limitation of detection determined at 10 plasmid copies/μL (corresponding to about one oocyst), cross reactivity to none of Giardia lamblia, Cryptosporidium sp., Microsporidium biberi and Blastocystis hominis that also commonly found in cats, and comparable performance in detecting T. gondii in clinical samples to conventional PCR amplification. This RPA-CRISPR/Cas9 method provides an alternative to conventional molecular tools used in the clinical diagnosis of Toxoplasma infection in cats and other animals., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Our understanding of anthelmintic resistance in the gastrointestinal nematodes of Australian cattle relies exclusively on small-scale phenotypic reports utilising traditional faecal egg count reduction tests. This approach is not readily scalable to establish the national prevalence of resistance, nor is it conducive of routine longitudinal surveillance for the emergence of resistance in its early stages. This study introduces the benefits of applying mixed amplicon metabarcoding longitudinally for timely and cost-efficient molecular surveillance of multiple anthelmintic resistance mutations, as they emerge on farms. Using opportunistically collected faecal samples from a cattle herd in central west New South Wales (2019-2023), we detected the early emergence of Haemonchus spp. levamisole-resistant S168T shortly after levamisole introduction, while benzimidazole-resistant allele frequencies remained constant. Additionally, we observed the possible spill-over of resistant Haemonchus contortus from sheep, along with variations in faecal burdens and species diversity influenced by climate stochasticity and host immunity. This study emphasises the power of molecular diagnostics for farm-level anthelmintic resistance management, providing essential evidence to support its integration into routine surveillance programmes., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Editorial Board of Veterinary Parasitology (JŠ)., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: An in vivo/in vitro ileal fermentation assay using growing pigs has been developed but not yet formally validated., Objectives: This study aimed to validate the in vivo/in vitro ileal fermentation assay by comparing in vitro fermentation values with those obtained in vivo in growing pigs. The effect of raising pigs under different environmental conditions was also investigated., Methods: Thirty piglets (1.59 ± 0.31 kg body weight, mean ± standard deviation) were subjected to 1 of 3 treatments: artificially reared (AR) (nonfarm, laboratory housing conditions) from postnatal day (PND) 7 (AR group), inoculated orally with human infant fecal extracts from birth until PND 8 and AR (AR+ group), or conventionally reared on a farm (control group). Starting at PND 7, the AR and AR+ pigs received human infant formula for 3 wk, followed by a human-type diet for 5 wk. Control pigs were weaned on the farm and, on PND 63, relocated to the laboratory animal facility. From PND 63, all pigs received a human-type diet. On PND 78, pigs were killed, after which ileal digesta were collected to perform an in vitro ileal fermentation (in vitro organic matter [OM] fermentability and organic acid production) and to determine digesta microbial composition and dietary OM fermentability in vivo., Results: The rearing regimen resulted in only a few differences in ileal microbial taxonomic composition. The rearing regimen generally did not affect the in vitro production of individual organic acids. The in vivo and in vitro OM fermentability of proximal ileal digesta (19.7 ± 2.04%; mean ± SEM) was similar (P > 0.05) for the AR and control pigs but not for the AR+ pigs., Conclusions: The control-rearing regimen was preferred over AR or AR+ because of ease of implementation. The in vitro ileal fermentation assay accurately predicted the in vivo OM fermentability., (Copyright © 2024 American Society for Nutrition. Published by Elsevier Inc. All rights reserved.)