Enteric Pathogens Research Articles

Potential probiotic Lactiplantibacillus plantarum strains alleviate TNF-α by regulating ADAM17 protein and ameliorate gut integrity through tight junction protein expression in in vitro model

BackgroundLactiplantibacillus species are extensively studied for their ability to regulate host immune responses and functional therapeutic potentials. Nevertheless, there is a lack of understanding on the mechanisms of interactions with the hosts during immunoregulatory activities.MethodsTwo Lactiplantibacillus plantarum strains MKMB01 and MKMB02 were tested for probiotic potential following Indian Council of Medical Research (ICMR) guidelines. Human colorectal adenocarcinoma cells such as HT-29, caco-2, and human monocytic cell THP-1 were also used to study the potential of MKMB01 and MKMB02 in regulating the host immune response when challenged with enteric pathogen Salmonella enterica typhimurium. Cells were pre-treated with MKMB01 and MKMB02 for 4 h and then stimulated with Salmonella. qRT-PCR and ELISA were used to analyze the genes and protein expression. Confocal microscopy and field emission scanning electron microscopy (FESEM) were used to visualize the effects. An Agilent Seahorse XF analyzer was used to determine real-time mitochondrial functioning.ResultsBoth probiotic strains could defend against Salmonella by maintaining gut integrity via expressing tight junction proteins (TJPs), MUC-2, and toll-like receptors (TLRs) negative regulators such as single Ig IL-1-related receptor (SIGIRR), toll-interacting protein (Tollip), interleukin-1 receptor-associated kinase (IRAK)-M, A20, and anti-inflammatory transforming growth factor-β and interleukin-10. Both strains also downregulated the expression of pro-inflammatory cytokines/chemokines interleukin-1β, monocyte chemoattractant protein (MCP)-1, tumor necrosis factor-alpha (TNF-α), interleukin 6, and nitric oxide (NO). Moreover, TNF-α sheddase protein, a disintegrin and metalloproteinase domain 17 (ADAM17), and its regulator iRhom2 were downregulated by both strains. Moreover, the bacteria also ameliorated Salmonella-induced mitochondrial dysfunction by restoring bioenergetic profiles, such as non-mitochondrial respiration, spare respiratory capacity (SRC), basal respiration, adenosine triphosphate (ATP) production, and maximal respiration.ConclusionsMKMB01 and MKMB02 can reduce pathogen-induced gut-associated disorders and therefore should be further explored for their probiotic potential.

Surveillance of Food and Waterborne Pathogens in North-East India: Protocol for a Laboratory-Based Sentinel Surveillance Study.

Food safety is a global concern, which is often underestimated owing to challenges in investigating foodborne diseases. These challenges arise from the increased globalization of the food trade, advancements in agricultural practices, and shifts in environmental factors. In North-East India, common diarrheal outbreaks from fermented foods pose significant health risks. Despite these challenges, systematic data on foodborne pathogens is lacking in India, highlighting a crucial gap in understanding food safety issues. The aim of this research protocol is to establish an integrated surveillance system to identify enteric pathogens circulating within humans, food animals, and the environment through a health approach in North-East India, and to conduct outbreak investigations. The Indian Council of Medical Research (ICMR) initiated a surveillance study across all 8 North-East states in India, employing a centralized digital database for data collation. The project aims to enhance theinfrastructure for microbial culture, antibiotic sensitivity testing, and molecular epidemiological studies. The study involves laboratory-based surveillance of foodborne pathogens in market foods, hospitalized diarrheal patients, poultry and animal farms, slaughterhouses, butcher shops, and diarrheal outbreaks. A standardized case report form ensures consistent data collection of age, sex, signs, symptoms, and admission dates for diarrheal cases. Stool and rectal swabs will undergo testing for pathogen identification and antimicrobial resistance. Similarly, samples of market foods, food animals, and the environment will be collected. Outbreaks confirmed by the Integrated Disease Surveillance Project (IDSP) will be thoroughly investigated following standardized guidelines. In phase I, 5 surveillance centers were established across 4 states (ie, Assam [Dibrugarh and Guwahati], Tripura, Sikkim, and Arunachal Pradesh) in 2020. Following an interim phase I data assessment and the successful establishment of a streamlined system for data procurement, investigation, recording, and analysis, along with the implementation of regular training and monitoring programs, phase II expansion was initiated in 2023-24. This includes the addition of 7 more centers (including 3 veterinary centers) in the remaining 4 states (ie, Manipur, Meghalaya, Mizoram, and Nagaland), eventually covering the entire North-Eastern Region of India. Food and waterborne diseases are a constant public health problem in many countries. Key challenges to the enhancement of food safety policy include the paucity of systematic data and awareness. With this background, ICMR's initiative is the first systematic surveillance study in the country to adopt a single health approach. Data obtained from this project will help to understand the risk of acquiring food and waterborne pathogens, their transmission pathways, and antimicrobial resistance patterns. The scientific evidence generated through this project will be helpful in formulating and strengthening food safety policy and in initiating government programs to protect the health of the nation. DERR1-10.2196/56469.

Multiplex PCR detection of enteric pathogens in a community-based birth cohort in Ecuador: comparison of xTAG-GPP and TaqMan array card assays.

We compared the performance of two multiplex platforms, Luminex xTAG Gastrointestinal Pathogen PanelⓇ and TaqMan Array Card, against a panel of 14 enteric pathogen targets in a community-based birth cohort in Ecuador. We found high levels of agreement and similar prevalence estimates across most pathogens.

Enteric Pathogen Detection Using Multiplex PCR Assay in Kidney Transplant Recipients with Diarrhea-Retrospective Before-After Study.

Diarrhea is a frequent complication after kidney transplantation, however the etiology is often not identified. Multiplex PCR assays may increase the detection of diarrheal pathogens among kidney transplant recipients (KTRs), leading to improved management. This was a retrospective before-after study, conducted in a high-volume transplant center. In September 2017, multiplex PCR assay was introduced. We reviewed all hospitalized KTRs with diarrhea during 1/2015-8/2017 (pre-GI PCR, n = 111) and 9/2017-12/2021 (GI PCR, n = 159) and followed them for 3years. We performed univariate and multivariate analysis for predictors of pathogen identification, introducing the study period as an independent variable. Among 270 hospitalized KTRs with diarrhea, 64 (24%) had an identified diarrheal pathogen. The proportion of KTRs with an identified pathogen increased from 20% (13/64) in the pre-GI PCR to 80% (51/64) post GI PCR (p < 0.01). Of 51 KTRs with an identified pathogen in the post GI PCR, 44 (86%) were diagnosed using GI PCR. GI PCR was more likely used in younger KTRs with more recent transplantation and higher creatinine level at admission. The most common non-C. difficile diarrheal pathogens in the post-GI PCR cohort were enteropathogenic Escherichia coli (n = 23, 58%), norovirus (n = 11, 28%), and Campylobacter (n = 11, 28%). Implementing GI PCR significantly increased the detection and identification of GI pathogens (odds ratio [OR] = 21, CI 95% 10-44; p < 0.001). Infectious etiologies of diarrhea were identified in a higher proportion of KTRs after the implementation of GI PCR. This emphasizes the importance of integrating this diagnostic tool into diarrhea workup in KTRs.

Spatial–Temporal Patterns in the Enteric Pathogen Contamination of Soil in the Public Environments of Low- and Middle-Income Neighborhoods in Nairobi, Kenya

Public spaces in countries with limited societal development can be contaminated with feces containing pathogenic microbes from animals and people. Data on contamination levels, spatial distribution, and the diversity of enteric pathogens in the public settings of low- and middle-income neighborhoods are crucial for devising strategies that minimize the enteric infection burden. The objective of this study was to compare spatial–temporal differences in the detection rate and diversity of enteric pathogens in the public spaces of low- and middle-income neighborhoods of Nairobi, Kenya. TaqMan array card (TAC) molecular assays were employed to analyze soil samples for 19 enteropathogens, along with a selective bacterial culture for pathogenic Enterobacteriaceae. An observational assessment was conducted during every site visit to document the hygienic infrastructure and sanitation conditions at the sites. We detected at least one pathogen in 79% (127/160) and ≥2 pathogens in 67.5% (108/160) of the soil samples tested. The four most frequently detected pathogens were EAEC (67.5%), ETEC (59%), EPEC (57.5%), and STEC (31%). The detection rate (91% vs. 66%) and mean number of enteric pathogens (5 vs. 4.7) were higher in low-income Kibera than in middle-income Jericho. The more extensive spatial distribution of pathogens in Kibera resulted in increases in the detection of different enteric pathogens from within-site (area &lt; 50 m2) and across-site (across-neighborhood) movements compared to Jericho. The pathogen detection rates fluctuated seasonally in Jericho but remained at sustained high levels in Kibera. While better neighborhood conditions were linked with lower pathogen detection rates, pathogenic E. coli remained prevalent in the public environment across both neighborhoods. Future studies should focus on identifying how the sources of pathogen contamination are modified by improved environmental sanitation and hygiene and the role of these contaminated public environments in enteric infections in children.

Oral supplementation with lactic acid bacteria improve the intestinal epithelial barrier and gut microbiota of broiler chicks to alleviate Salmonella Enteritidis infection

Lactic acid bacteria (LAB) play a key role in regulating the balance of gut microbiota and serve as a suitable alternative to antibiotics. This study aims to evaluate the characteristics of 2 LAB isolates Lactiplantibacillus plantarum Lp71 (L. plantarum Lp71) and Enterococcus faecium Ef72 (E. faecium Ef72), and their roles in alleviating Salmonella Enteritidis infection. Sixty 1-day-old chicks were randomly divided into 4 groups which treated with or without L. plantarum Lp71 and E. faecium Ef72 mixture for 21 d, and then intestinal samples were collected for gut microbiota analysis, pathological and immunohistochemical analysis at 24 h post infection with or without Salmonella Enteritidis on the 22nd d. The results showed that L. plantarum Lp71 and E. faecium Ef72 had the ability to anti-acid and anti-bile salt. Salmonella Enteritidis infection damaged the intestinal epithelial barrier and reduced the expression level of tight junction proteins (ZO-1, Claudin-1, Occludin). Oral supplementation with L. plantarum Lp71 and E. faecium Ef72 mixture could alleviated the damages to intestinal epithelial barrier by Salmonella Enteritidis infection. Salmonella Enteritidis could cause abnormal Akkermansia muciniphila proliferation and decrease the diversity of cecal microbiota in chicks. These conditions could have further led to reduce gut microbiota health index (GMHI), and improve microbial dysbiosis index (MDI). Moreover, oral supplementation with L. plantarum Lp71 and E. faecium Ef72 mixture could effectively prevent the aforementioned infection outcomes and increase the abundance proportions of the several key functions in metabolic pathways metabolic pathways such as transcription and signal transduction mechanisms. In summary, L. plantarum Lp71 and E. faecium Ef72 could be the probiotics candidates that used to prevent the damage from enteric pathogens such as Salmonella Enteritidis in broiler chicks.

Demographic, Behavioural, and Biological Factors Seen in Men Who Have Sex with Men with Salmonella spp.: A Systematic Review

Background: The sexual transmissibility of enteric pathogens, including Salmonella spp., has been described in men who have sex with men (MSM). However, the factors seen in MSM with Salmonella spp. are poorly understood. Method: We aimed to systematically review the literature to explore any factors seen in MSM with Salmonella spp. (MSM). We searched six databases—Medline, PubMed, CINAHL, Embase, Emcare, and Global Health—in April 2024 for manuscripts which contained primary peer-reviewed data in English and the measurement of any risk factors observed in MSM with Salmonella spp. This review was registered on PROSPERO (CRD42023472864). Results: Eleven manuscripts were included in the final review and highlighted demographic (living with HIV), behavioural (oral–anal sex, receptive and penetrative anal sex, hand licking to stimulate their partner, group sex, non-condom use), and biological (co-infection with CMV, Mycobacterium avium complex, Strongyloides stercoralis, Blastocystis hominis, Klebsiella spp. Herpes simplex virus, Cytomegalovirus, Cryptosporidium, Histoplasmosis, Shigella spp.; previous infection with Treponema pallidum, Neisseria gonorhoeae, Chlamydia trachomatis and hepatitis B; and antimicrobial treatment failure) factors seen in MSM with Salmonella spp. Conclusion: Despite a limited number of manuscripts and individuals, this review highlighted some potential demographic, behavioural, and biological factors implicated in the transmission of Salmonella spp. in MSM. These data will provide insights for future guidelines, public health control strategies, and research.

Magnitude of enteric pathogens associated with diarrhea and antibiotic resistance of enteric bacterial pathogens isolated among children under 5 years of age in Bule Hora town, West Guji, Ethiopia.

Diarrhea is the second leading cause of morbidity and mortality worldwide among all ages, and one of the leading causes of poor health and premature death in the developing world. Microorganisms, such as viruses, bacteria, and parasites, are responsible for enteric infections among children. Excessive and inappropriate use of antimicrobial drugs and poor infection control practices have transformed antimicrobial resistance into a serious threat to public health worldwide. Therefore, it is essential to investigate the prevalence of enteric pathogens and the antimicrobial susceptibility patterns of bacterial pathogens. Assess the prevalence of enteric pathogens associated with diarrhea in children under 5 years at the Bule Hora Health Facility in 2021. An institution-based cross-sectional study was conducted from May 2021 to July 2021 in a Bule Hora Health Centre and Hospital. A total of 422 children with diarrhea were included in the study in the outpatient department. Sociodemographic and associated factors were evaluated using a pre-tested questionnaire. A sufficient amount of stool specimens was collected following standard microbiological procedures. An antigen detection kit was used to diagnose rotaviruses, parasites were examined using microscopy, and bacterial identification was carried out by culture and biochemical tests. The antibiotic susceptibility test of the bacterial isolates was performed using the Kirby-Bauer disc diffusion method. The data were analyzed using SPSS version 20. The p-value less than 0.05 was declared statistically significant. The overall prevalence of enteropathogens in children under 5 years of age was 17%, with no mixed infections detected. Of this, 7.8% of the children's stool samples contained bacterial pathogens, 5% tested positive for rotavirus, and 4.2% contained intestinal parasites. Unprotected drinking water sources, poor carrier hand washing practices, and poor cleaning of utensils for child feeding were factors significantly associated with the prevalence of enteropathogens. Bacterial isolates have shown a high prevalence of resistance to amoxicillin. Therefore, it is important to take steps to reduce the spread of enteric pathogens among children under 5 years practicing good hygiene, ensuring sanitation, and providing clean drinking water. We recommend performing antimicrobial susceptibility tests before prescribing treatment to children with diarrhea.

A-277 Viability of multi-drug resistant organisms in a fecal collection and transport system stored at ultra-low temperature

Abstract Background Epidemiological studies are constantly investigating new, easier ways to collect and store patient specimens that allow for flexible testing windows and long-term storage as well as accurate quantification of organisms of interest. The FecalSwabTM system (Copan, Brescia, Italy) is intended for the collection and preservation of rectal and fecal specimens. This system includes a preservation medium which serves to maintain the viability of enteric pathogens during transport to a clinical laboratory for up to 72 hours at 4oC. The objective of this study was to assess the continued viability of multi-drug resistant organisms (MDROs) suspended in the FecalSwab sample format when frozen at -80°C and the impact of freeze-thaw cycles. Methods A total of 40 organisms were chosen spanning methicillin-resistant Staphylococcus aureus (MRSA, n=10), carbapenem-resistant Pseudomonas aeruginosa (CRPA, n=5), carbapenem-resistant Enterobacterales (CRE, n=5), vancomycin-resistant Enterobacterales (VRE, n=10) and extended-spectrum beta-lactamase containing Enterobacterales (ESBL, n=10). Each organism was spiked into FecalSwab at standardized concentrations. At each time point, FecalSwab media was plated onto blood agar and colonies were quantified. Each of the 40 organisms were assessed one-week post-freeze and over the course of freeze-thaw cycles at days 3 and 5. The survival of each MDRO was evaluated by calculating a difference between the concentration of organisms recovered (CFU per 10uL media) at each time point and the concentration of organisms confirmed present at the time of inoculation. Percent recovery was calculated from this difference. Results One week after freezing, the percent recovery for MRSA, VRE, ESBL, CRE, and CRPA averaged to 102%, 201%, 17.8%, 7.6%, and 0.8%, respectively (Kruskal Wallis p &amp;lt; 0.001), with significant differences noted between gram positive and gram negative organisms (Kruskal Wallis p &amp;lt;0.001). Conclusions Epidemiological studies interested in MDRO surveillance rely on methodologies that allow for accurate quantification of viable organisms. In this study, we’ve evaluated the viability of several multi-drug resistant organisms when frozen at -80°C in the FecalSwab. The FecalSwab system is intended for the collection of rectal swab and fecal specimens and to preserve the viability of enteric pathogenic bacteria for testing in the clinical laboratory. Although it was not specifically designed to preserve MDROs, it performed well with MRSA and VRE, but less well with CRE, CRPA and ESBL organisms. When using the FecalSwab system, the observed gram-specific differences may inform the suitability of this system depending on the organism(s) of interest.

Asymptomatic enteric pathogen carriage and its association with proton pump inhibitors use in men who have sex with men in Taiwan, 2019-2022

ObjectivesCurrently recognized risk factors for sexually transmitted enteric infections (STEIs) among men who have sex with men (MSM) include oroanal sex, multiple sexual partners, and chemsex. This study aimed to investigate the prevalence of the asymptomatic carriage of enteric pathogens in men who have sex with men (MSM) and to identify the associated risk factors. MethodsQuestionnaires were completed by 375 MSM in Taiwan from December 2019 to November 2022. Fecal samples were analyzed by multiplex PCR to determine whether seven enteric pathogens, including Entamoeba histolytica, Giardia duodenalis, Shiga toxin-producing Escherichia coli, Cryptosporidium, Campylobacter, Salmonella, and Shigella species, were present. ResultsAmong 375 fecal samples from asymptomatic MSM, 27 (7.2%) fecal samples tested positive for at least one enteric pathogen. The recent use of proton pump inhibitors (PPIs) was significantly associated with asymptomatic fecal carriage (22.2% vs. 2.0%, P < 0.001). G. duodenalis (2.1%, 8 cases), E. histolytica (1.6%, 6 cases), and Shigella species (1.3%, 5 cases) were commonly detected. Oroanal sex and PPI use were associated with the asymptomatic carriage of enteric pathogens. Specifically, Shigella, Salmonella, or Campylobacter carriage was significantly correlated with PPI use. In contrast, rectal gonorrhea was associated with multiple sexual partners and prior syphilis. ConclusionsRecent use of PPIs was associated with the asymptomatic carriage of enteric pathogens. Therefore, targeted education about the appropriate use of PPIs is necessary to mitigate the risk of STEIs among MSM.

Herd-level occurrence and risk factors associated with respiratory and enteric pathogens from dairy calves in Ontario: A cross-sectional study

This cross-sectional herd-level study aimed to determine the occurrence of and risk factors for pathogens associated with neonatal calf diarrhea and bovine respiratory disease on Ontario dairy farms. From April to August 2022, a convenience sample of 100 dairy farms were visited once. A questionnaire covering farm biosecurity, calving and colostrum management, preweaning nutrition, and housing was administered on-farm. At each farm visit, approximately 5 calves between 2 and 35 d old were randomly selected for fecal sampling. Furthermore, approximately 5 calves between 21 to 122 d old were randomly selected for nasopharyngeal sampling. In total, 363 fecal samples (from 83 dairy farms) and 390 nasopharyngeal swab samples (from 80 dairy farms) were collected. Fecal samples were analyzed individually using a multiplex PCR to identify bacterial and parasitic enteric pathogens. Nasopharyngeal swabs were analyzed as one pooled sample per farm using bacterial culture and real-time PCR. The most common enteric pathogens detected at herd-level were Cryptosporidium parvum (67.4%) and Escherichia coli K99+ (13.2%). The most common respiratory pathogens detected at herd-level were Pasteurella multocida (62.5%), bovine coronavirus (42.5%), and Mycoplasma bovis (21.2%). Multivariable logistic models were built to explore associations between the most common pathogens and herd-level predictors selected from the questionnaire. Herd positivity for C. parvum was positively associated with having more than 61 preweaned calves per year and feeding mainly whole milk to calves. The presence of M. bovis was positively associated with herds that combined manual and automatic milk-feeding systems, and the presence of bovine coronavirus was positively associated with having more than 98 preweaned calves during the year. Univariable Poisson regression models were built to explore the association between the most common pathogens and preweaning calf mortality. Herds that were positive for C. parvum, M. bovis, or bovine coronavirus had a greater risk of preweaning calf mortality. These results provide insights for future research on pathogens associated with NCD and BRD and offer guidance for veterinarians and dairy farmers in implementing disease control measures in dairy calf herds.

Molecular Typing of Cryptosporidium in Cattle in Federal Capital Territory, Nigeria

Cryptosporidium is an enteric pathogen with worldwide distribution in humans and animals with cattle as their reservoirs. Molecular tools have been very useful in determining the epidemiology of this zoonotic pathogen of public health importance. Despite the importance of this pathogen to humans and animals, there is limited published information on the risk factors of its epidemiology in the Federal Capital Territory of Nigeria. Positive faecal samples of 400 cattle in 46 farms located in 6 area councils of the FCT that were earlier screened for Cryptosporidium on light microscopy were genotyped by nested PCR using primers that targeted 18S ssUrRNA gene and followed by RFLP. The PCR-RFLP products were further sub genotyped by nested PCR and GP 60 KDa gene amplification. The nucleotides of the amplicons were subsequently sequenced. The overall PCR detection rate was 5.5%. Analysis of the 18S rRNA gene of the PCR-RFLP fragments revealed Cryptosporidium ryanae (18.3%), C. andersoni (8.5%) and C. parvum (4.2%). Analysis of the nucleotide sequence of the GP 60KDa gene of all the C. parvum detected showed all to be 100% subtype family IIa and 100% sub genotype IIaA18G3R1. In conclusion, this study has established the presence of Cryptosporidium in cattle in the FCT. The detection of the zoonotic C. parvum, its subtype IIa and its allele IIaA18G3R1 is an indication of source of zoonotic transmission of the parasite in the FCT, Nigeria.

S1378 Corticosteroids Prior to Antimicrobials Do Not Increase the Risk of Adverse Outcomes Among Individuals Hospitalized for IBD Flares Complicated by Enteric Pathogens

S1378 Corticosteroids Prior to Antimicrobials Do Not Increase the Risk of Adverse Outcomes Among Individuals Hospitalized for IBD Flares Complicated by Enteric Pathogens

Greywater recycling for diverse collection scales and appliances: enteric pathogen log-removal targets and treatment trains

In light of increasingly diverse greywater reuse applications, this study proposes risk-based log-removal targets (LRTs) to aid the selection of treatment trains for greywater recycling at different collection scales, including appliance-scale reuse of individual greywater streams. An epidemiology-based model was used to simulate the concentrations of prevalent and treatment-resistant reference pathogens (protozoa: Giardia and Cryptosporidium spp., bacteria: Salmonella and Campylobacter spp., viruses: rotavirus, norovirus, adenovirus, and Coxsackievirus B5) in the greywater streams for collection scales of 5-, 100-, and a 1000-people. Using quantitative microbial risk assessment (QMRA), we calculated LRTs to meet a health benchmark of 10-4 infections per person per year over 10’000 Monte Carlo iterations. LRTs were highest for norovirus at the 5-people scale and for adenovirus at the 100- and 1000-people scales. Example treatment trains were designed to meet the 95% quantiles of LRTs. Treatment trains consisted of an aerated membrane bioreactor, chlorination, and, if required, UV disinfection. In most cases, rotavirus, norovirus, adenovirus and Cryptosporidium spp. determined the overall treatment train requirements. Norovirus was most often critical to dimension the chlorination (concentration × time values) and adenovirus determined the required UV dose. Smaller collection scales did not generally allow for simpler treatment trains due to the high LRTs associated with viruses, with the exception of recirculating washing machines and handwashing stations. Similarly, treating greywater sources individually resulted in lower LRTs, but the lower required LRTs nevertheless did not generally allow for simpler treatment trains. For instance, LRTs for a recirculating washing machine were around 2-log units lower compared to LRTs for indoor reuse of combined greywater (1000-people scale), but both scenarios necessitated treatment with a membrane bioreactor, chlorination and UV disinfection. However, simpler treatment trains may be feasible for small-scale and application-scale reuse if: (i) less conservative health benchmarks are used for household-based systems, considering the reduced relative importance of treated greywater in pathogen transmission in households, and (ii) higher log-removal values (LRVs) can be validated for unit processes, enabling simpler treatment trains for a larger number of appliance-scale reuse systems.

Epiphytic, Attached, and Internal Escherichia coliO157:H7 Subpopulations Associating With Romaine Lettuce Are Strain‐Dependent and Affected by Relative Humidity and Pre‐ and Postharvest Plant State

ABSTRACTRomaine lettuce is susceptible to Escherichia coli O157:H7 contamination. We evaluated strain and pre‐ and postharvest lettuce product differences in E. coli O157:H7 subpopulation distribution on romaine lettuce at two relative humidity (RH) levels. Plants of romaine lettuce cultivar 'Carlsbad' harvested and processed 'Carlsbad' leaves, and store‐bought ready‐to‐eat romaine lettuce were inoculated with E. coli O157:H7 reference strain EDL933 and romaine lettuce outbreak strain 2705C. Using four processing methods, we determined pathogen cell fractions representing All (entire population), Epiphytic (loosely attached cells), Strongly Attached + Internal (excluding loosely attached cells), and Internal (excluding epiphytic cells) subpopulations. Preharvest, 80% RH favored subpopulations in each cell fraction, compared to 40% RH (p &lt; 0.01 for both strains) and yielded 92%–100% internalization incidence of E. coli O157:H7, compared to 50%–57% at 40% RH. Levels of internal EDL933 cells were also 1.1 log higher than 2705C cells from plants kept at 80% RH (p &lt; 0.001). While EDL933 had lower measures of Strongly Attached + Internal cells compared to All and Epiphytic fractions (p &lt; 0.01), 2705C yielded no difference. Taken together, data suggest that the lettuce outbreak strain had a higher propensity for strong attachment to leaves and EDL933 internalized more successfully. Moreover, the Strongly Attached + Internal fractions of both strains were lower on preharvest 'Carlsbad' compared to 'Carlsbad' processed leaves (p &lt; 0.01), suggesting that E. coli O157:H7 attached less strongly to preharvest plants than postharvest cut and stored leaves of the same variety. Our study uncovers important factors influencing cultivar‐ and strain‐specific differences in association and internalization of enteric pathogens on leafy greens.

A cross-sectional study of associations between the 13C-sucrose breath test, the lactulose rhamnose assay, and growth in children at high risk of environmental enteropathy

BackgroundEnvironmental enteropathy’ (EE) is common among children who are highly exposed to enteric pathogens in low-resource settings. We optimised and validated a stable isotope-based breath test of intestinal sucrase activity (13C-SBT) as a non-invasive test of carbohydrate digestion and metabolism. ObjectivesThe primary objective of this study was to assess the relationship between the 13C-SBT and the lactulose/rhamnose ratio (LR) and growth in children. Secondary objectives were to assess the relationship between the 13C-SBT and additional biomarkers of EE. We also characterised the relationship between the 13C-SBT and child sex anddietary diversity, and household socioeconomic status and food security. MethodsIn this cross-sectional study, 12-to-15-month-old children were recruited in Bangladesh, India, Kenya, and Peru. Children were assessed with a 4-hour 13C-SBT and a 90-minute LR test. Plasma was collected for the determination of citrulline and the kynurenine/tryptophan ratio. Length and weight were measured, and other variables were assessed through questionnaires. For a subset of children, anthropometry was re-measured after three months. inear regression was used to examine associations corresponding to each objective. ResultsThree sites generated 13C-SBT breath curves that enabled pooled analysis. Differences in 13C-SBT breath curves, LR ratios, and other EE biomarkers were observed between sites. No associations were observed for 13C-SBT summary measures and LR, or child growth (e.g., association between LR and cumulative percent dose recovered at 90 minutes (cPDR90): -0.39, 95%CI: -1.79, 0.70). Length-for-age and weight-for-age were positively associated with the time to 50% of dose recovered (T50) (0.05, 95%CI: 0.01, 0.09, and 0.05, 95%CI: 0.02, 0.07, respectively), and dietary diversity was associated with T50 and cPDR90(-0.10, 95%CI: -0.18, -0.02 and 2.67, 95%CI: 0.47, 4.88, respectively). ConclusionsIn children at risk of EE there were no associations between the 13C-SBT, LR or other EE biomarkers encompassing different pathophysiological domains of EE. Trial Registryhttps://clinicaltrials.gov/study/NCT04109352

Evaluation of physical, chemical, and microbiological characteristics of waste stabilization ponds, Giza, Egypt

Wastewater treatment plants (WWTPs) contain a diverse array of microbes, underscoring the need for regular monitoring to ensure treatment efficacy and protect health. However, detailed studies on waste stabilization ponds (WSPs) are scarce. This study evaluates a full-scale WSP, located in Giza Governorate, Egypt, including anaerobic, facultative, and maturation ponds, examining an array of parameters such as enteric viruses, microeukaryotes (protozoa and algae), bacterial indicators, bacterial pathogens, and physicochemical characteristics. Utilizing multivariate statistical models, we identified significant distinctions in physicochemical parameters and microbial communities, primarily driven by treatment stages rather than temporal variations. In addition, seven viruses (human rotavirus, adenovirus, norovirus, astrovirus, hepatitis A virus, polyomavirus, and papillomavirus) were detected during the different stages (inlet, anaerobic, facultative, and outlet) of the WSP, except norovirus and papillomavirus were absent in the outlet stage. The viral log means reductions ranged from 1.24 to 5.94, depending on the stage and virus type. The removal efficiency of bacterial pathogens was more than 99%. High throughput 18S rRNA gene amplicon sequencing indicated the dominance of animal parasitic Apicomplexa species and Vermamoeba spp. in the WSP. Network analysis indicated significant roles for Ciliophora in virus reduction. Notably, the maturation pond's outlet was dominated by Spirulina maxima, whose mat-forming tendencies may inhibit pathogen removal by providing protective shelters. Although the WSP effectively reduced pathogen levels, the high initial loads resulted in considerable concentrations in the final effluent, posing ongoing public health concerns. This study highlights the imperative of including pathogen standards in national regulations for wastewater reuse.

Towards geospatially-resolved public-health surveillance via wastewater sequencing

Wastewater is a geospatially- and temporally-linked microbial fingerprint of a given population, making it a potentially valuable tool for tracking public health across locales and time. Here, we integrate targeted and bulk RNA sequencing (N = 2238 samples) to track the viral, bacterial, and functional content over geospatially distinct areas within Miami Dade County, USA, from 2020-2022. We used targeted amplicon sequencing to track diverse SARS-CoV-2 variants across space and time, and we found a tight correspondence with positive PCR tests from University students and Miami-Dade hospital patients. Additionally, in bulk metatranscriptomic data, we demonstrate that the bacterial content of different wastewater sampling locations serving small population sizes can be used to detect putative, host-derived microorganisms that themselves have known associations with human health and diet. We also detect multiple enteric pathogens (e.g., Norovirus) and characterize viral diversity across sites. Moreover, we observed an enrichment of antimicrobial resistance genes (ARGs) in hospital wastewater; antibiotic-specific ARGs correlated to total prescriptions of those same antibiotics (e.g Ampicillin, Gentamicin). Overall, this effort lays the groundwork for systematic characterization of wastewater that can potentially influence public health decision-making.

Assessing antimicrobial resistance profiles of Salmonella enterica in the pork production system.

Introduction. Salmonella enterica is a significant enteric pathogen affecting human and livestock health. Pork production is a common source of Salmonella contamination, with emerging multidrug resistance (MDR) posing a global health threat.Gap statement. Salmonella contamination and antimicrobial resistance (AMR) profiles in the pig production chain are underreported.Aim. To investigate the prevalence of S. enterica in the pig production chain and characterise their AMR profiles.Methodology. We collected 485 samples from pig farms, a standard pig abattoir and retail markets in Patthalung and Songkhla provinces in southern Thailand. Antimicrobial susceptibility testing was performed on these samples, and AMR profiles were determined.Results. S. enterica was detected in 68.67% of farm samples, 45.95% of abattoir samples and 50.67% of retail market samples. Analysis of 264 isolates, representing 18 serotypes, identified S. enterica serotype Rissen as the most prevalent. The predominant resistance phenotypes included ampicillin (AMP, 91.29%), tetracycline (TET, 88.26%) and streptomycin (STR, 84.47%). Over 80% of isolates showed resistance to three or more antimicrobial classes, indicating MDR. The AMP-STR-TET resistance pattern was found in nearly 70% of all MDR isolates across the production chain.Conclusions. The high prevalence of MDR is consistent with extensive antimicrobial use in the livestock sector. The presence of extensively resistant S. enterica highlights the urgent need for antimicrobial stewardship. Strengthening preventive strategies and control measures is crucial to mitigate the risk of MDR Salmonella spreading from farm to fork.

Multilevel factors drive child exposure to enteric pathogens in animal feces: A qualitative study in northwestern coastal Ecuador.

Exposure to animal feces and associated enteric pathogens poses significant risks to child health. However, public health strategies to mitigate enteric infections among children largely aim to reduce exposure to human feces, overlooking transmission pathways related to animal feces. In this study we examine if and how children are exposed to enteric pathogens in animal feces in northwestern coastal Ecuador. We conducted qualitative interviews with mothers of children aged 10-18 months that owned (n = 32) and did not own (n = 26) animals in urban and rural communities. Using thematic analysis, we identified community, household, and child behavioral factors that influence exposure. We also compared child exposure by household animal ownership. Our findings revealed myriad opportunities for young children to be exposed to enteric pathogens in many locations and from multiple animal sources, regardless of household animal ownership. Animal feces management practices (AFM) used by mothers, such as rinsing feces into ditches and throwing feces into surrounding areas, may increase environmental contamination outside their homes and in their communities. Unsafe AFM practices were similar to unsafe child feces management practices reported in other studies, including practices related to defecation location, feces removal and disposal, environmental contamination cleaning, and handwashing. Findings suggest that animal feces may contaminate the environment along similar pathways as human feces. Identification and incorporation of safe AFM practices, similar to those developed for child feces management, would 1) mitigate child exposure to enteric pathogens by reducing animal feces contamination in domestic and public spaces; and 2) enable an integrated approach to address enteric pathogen exposure pathways related to animal and child feces.