Cheng Cheng

Background/Objectives: Enteropathogenic Escherichia coli O157:H7 infection disrupts intestinal homeostasis, causing dysbiosis, barrier dysfunction, and inflammation. This study aimed to evaluate the protective efficacy and mechanisms of a novel probiotic, Bacteroides thetaiotaomicron type strain ATCC 29148, isolated from goat feces, against E. coli O157:H7-induced colitis. Methods: This study assessed the protective potential of the probiotic strain Bacteroides thetaiotaomicronBT6 and BT7 in vitro for GI tolerance, adhesion, and no adverse effects were observed. For the in vivo experiment, male C57BL/6J mice were divided into groups treated with Bacteroides thetaiotaomicron (BT6), PBS, E. coli O157:H7, or a combination. We employed integrated analyses including 16S rRNA gene sequencing, antioxidant status, cytokine profiling, and short-chain fatty acid (SCFA) measurement. Results: In vitro, Bacteroides thetaiotaomicron (BT6 and BT7) showed high gastrointestinal tolerance (71.89–93.22% survival). In vivo, it significantly mitigated infection-associated weight loss and disease activity (p < 0.05). Probiotic treatment enhanced barrier integrity, reduced colonic inflammation, and modulated systemic immune responses, notably increasing anti-inflammatory IL-10 while decreasing pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 (p < 0.05). It also alleviated oxidative stress by reducing malondialdehyde (MDA) and elevating antioxidant enzymes (SOD, CAT, GSH) and ATP. Fecal SCFA profiling revealed increased propionic and butyric acid. 16S sequencing indicated that B. thetaiotaomicron (BT6) administration increased beneficial families (Lactobacillaceae, Muribaculaceae) and suppressed pathobionts. Conclusions: B. thetaiotaomicron (BT6) probiotic with potential for mitigating enteropathogenic infection, an effect mainly determined by its capacity to reestablish the intestinal epithelial barrier and enhance global host health, and modulating the inflammatory response.

Avian coccidiosis is an intestinal disease caused by Eimeria spp. infection. A deeper understanding of the interaction between host gut microbiota and the Eimeria parasite is crucial for developing alternative therapies to control avian coccidiosis. Here, we used full-length sequencing of 16S ribosomal RNA amplicons to compare changes in the gut microbiota of chickens infected with Eimeria tenella, Eimeria maxima, and Eimeria necatrix, aiming to identify both species-specific and common alterations in gut microbiota at 4 and 10 days post-infection. The result revealed that infection with all three Eimeria species led to a decrease in the abundance of the microbial genera Limosilactobacillus, Streptococcus, Alistipes, Lactobacillus and Phocaeicola, while the abundance of Bacteroides, Escherichia and Ligilactobacillus increased. Escherichia and Enterococcus were most abundant in the jejunum of the E. necatrix-infected group and in the cecum of the E. tenella-infected group, whereas Megamonas abundance was highest in the E. maxima-infected group. LEfSe analysis showed that infection with all three Eimeria species significantly reduced the abundance of 13 bacterial species, including Acetilactobacillus jinshanensis, Bacteroides ndongoniae, Barnesiella viscericola, Christensenella minuta, Enterocloster clostridioformis, Gemella haemolysans_A, Granulicatella adiacens, Lawsonibacter sp000177015, Limosilactobacillus reuteri, Limosilactobacillus reuteri_D, Limosilactobacillus vaginalis_A, Limosilactobacillus caviae, Limosilactobacillus vaginalis. Infection with E. tenella significantly increased the abundance of five bacterial species, including Bacteroides fragilis, Enterococcus cecorum, Helicobacter pylori, Methylovirgula ligni, and Phocaeicola sp900066445. Infection with E. maxima significantly increased the abundance of seven bacterial species, including Clostridioides difficile, Faecalibacterium prausnitzii, Mediterraneibacter torques, Muribaculum intestinale, Mediterraneibacter massiliensis, Phascolarctobacterium faecium, and Phocaeicola plebeius. Infection with E. necatrix significantly increased the abundance of seven bacterial species, including Alistipes sp900290115, Anaerotignum faecicola, Bacteroides fragilis_A, Escherichia coli, Harryflintia acetispora, Pseudoclostridium thermosuccinogenes, and Tidjanibacter inops_A. The results showed that Eimeria infection causes significant species- and time-dependent changes in the gut microbiota of chickens. These findings enhance our understanding of coccidiosis pathogenesis and offer potential targets for developing probiotics.

This research was conducted to evaluate the safety and effects of two probiotic candidate strains, Enterococcus faecium (E. faecium) and B. fragilis (BF), which were isolated from healthy sheep, on various factors such as intestinal structure, growth performance, organ indices, and gut microbiota in female C57BL/6J mice. A total of fifty mice were categorized into five groups: BF108, BF109, EF108, EF109, and a control group. The results indicated that mice in the EF108 and BF109 groups experienced a significant increase in body weight by day 28 compared to the control group (p < 0.05). Additionally, the liver index in the BF108 and EF108 groups was significantly lower (p < 0.05) than that of the control group. The length of the colon was also significantly greater (p < 0.05) in the EF109 and BF109 groups relative to the control group. According to the linear discriminant analysis effect size (LEfSe), the presence of probiotics led to an increased relative abundance of Pediococcus acidilactici in the EF109 and BF109 groups, while Lactiplantibacillus plantarum was the most prevalent strain in the EF108 and BF108 groups. Finally, Spearman correlation analysis demonstrated a positive association between Bacillota and Bacteroidota as well as Verrucomicrobiota. Additionally, Bacteroidota displayed positive relationships with Verrucomicrobiota and Planctomycetota, while showing a negative correlation with Actinomycetota and Gemmatimonadota, supporting its potential use as next-generation probiotics. In summary, our results indicate probiotics may have a role in promoting intestinal health by enhancing intestinal shape and histological characteristics.

Background: As critical nodes for zoonotic disease transmission, the microbial community composition in sheep slaughterhouse environments directly impacts meat safety and public health. While existing studies have focused on single-pathogen detection, there remains a lack of systematic understanding of the diversity, distribution patterns, and transmission risks of pathogenic microorganisms in complex environments. This study aims to elucidate the microbial community characteristics of mutton slaughterhouse environments using metagenomics approaches, with a focus on analyzing the ecological distribution of zoonotic pathogens and their potential threats. Materials, Methods & Results: A total of 12 environmental samples (n=3 per zone) were collected from a sheep slaughterhouse in Suzhou, Jiangsu Province, including slaughter workshop, slaughter tools, pre-slaughter holding area, and production wastewater. Samples were preserved in liquid nitrogen, and total DNA was extracted using the HiPure Stool DNA Kit. Paired-end sequencing (PE150) was performed on the Illumina HiSeq platform. Raw reads were quality-filtered (Cutadapt), and host-contaminant sequences removed (BWA). Clean reads were assembled with MEGAHIT, and unigenes predicted via Prodigal. Clustering by MMseq2 generated a non-redundant gene set, which was annotated against the NCBI NT database using Diamond to obtain taxonomic assignments. Species abundance was calculated at phylum, genus, and species levels, and principal component analysis (PCA) assessed variation between zones. At the phylum level, microbial communities were dominated by Proteobacteria (31.39% - 75%), Firmicutes (11.89% - 43.66%), Bacteroidota (9.15% -41.61%), and Actinobacteria (3% - 14.69%), with significant clustering by functional zones. Genus-level analysis revealed Chryseobacterium, Comamonas, Acinetobacter, Psychrobacter, and Prevotella as top taxa across samples. Species-level profiling detected 6 categories of zoonotic pathogens, notably Staphylococcus aureus (7.07% relative abundance in slaughter tools), Mycobacteroides abscessus (3.56% in slaughter workshop), Vibrio cholerae, and Klebsiella pneumoniae. Slaughter tools harbored the highest pathogen load; Lactococcus garvieae accounted for 10.39%. Production wastewater contained elevated levels of aquatic pathogens, suggesting environmental dissemination risks. An upset plot showed 116 species shared across all samples, with unique species counts ranging from 88 to 505 per sample. Discussion: This study elucidates the ecological niches of drug-resistant and opportunistic pathogens in a sheep slaughterhouse, identifying production wastewater and slaughter tools as high-risk contamination sources. High abundances of Staphylococcus aureus and Mycobacteroides abscessus underscore occupational exposure risks for abattoir workers, while detection of Vibrio cholerae highlights vulnerabilities in wastewater treatment and potential for zoonotic outbreaks. The prevalence of biofilm-forming Firmicutes on tools indicates that conventional disinfectants may be insufficient. Horizontal transfer of antibiotic resistance genes among dominant phyla could exacerbate public health threats. We recommend implementing targeted disinfection protocols, routine metagenomic monitoring for early pathogen detection, stricter PPE use and worker training, and optimization of wastewater pH control. Future studies should expand geographic sampling and integrate functional metagenomics to unravel resistance mechanisms and cross-species gene flow. These strategies will strengthen slaughterhouse biosafety and mitigate zoonotic disease transmission. Keywords: sheep slaughterhouse, public health, environment microbiology, zoonoses, pathogens, metagenomics.

Floppy Kid Syndrome (FKS) is a common and serious disease in goats, with incidence rates ranging from 10 to 50% and mortality rates between 20 and 60%. This study aimed to investigate the etiology of FKS through blood biochemical analysis and metagenomic sequencing. Blood biochemical analysis revealed metabolic disorders in FKS-affected goats, including acidosis and hypoglycemia. Metagenomic analysis showed marked gastric and gut dysbacteriosis, characterized by an increase in pathogenic bacteria such as Escherichia coli and Staphylococcus aureus, alongside a significant reduction in probiotic like Lactobacillus amylovorus. Furthermore, species diversity and richness were notably lower in FKS-affected goats compared to healthy goats. Based on these findings, we infer that FKS is a multifactorial disease caused by gastric and gut dysbacteriosis. The immaturity of the digestive system in newborn goats, combined with environmental stressors (such as sudden changes in weather), leads to gastric and gut dysbacteriosis, with a significant reduction in probiotic and an overgrowth of pathogenic bacteria. The dysbacteriosis, along with the inability to properly digest excessive milk intake, contributes to the accumulation of undigested milk in the digestive tract, creating an environment conducive to pathogenic bacteria growth. The fermentation of milk and the production of excessive lactic acid by pathogenic bacteria are absorbed into the bloodstream, causing acidosis and hypoglycemia. These metabolic disorders, in conjunction with the dysbacteriosis and systemic dysfunction, lead to the onset of FKS. These results underscore the critical role of gastric and gut dysbacteriosis in the pathogenesis of FKS, highlighting the need for targeted preventive and therapeutic strategies.

With the growing demand for sheep, the sheep farming industry has developed rapidly. However, lamb diarrhea, a disease with high mortality rates, significantly hampers the industry’s growth. Traditional antibiotic treatments often disrupt the Intestinal microbiota, induce antibiotic resistance, and cause adverse side effects, highlighting the urgent need to develop alternative therapies. Bacteroides fragilis, a candidate next-generation probiotic, has been closely associated with intestinal health. This study investigated the growth characteristics and probiotic effects of a sheep-derived Bacteroides fragilis isolate, focusing on its efficacy in alleviating lamb diarrhea and infectious intestinal diseases. The experiments demonstrated that the Bacteroides fragilis isolate grows well under mildly acidic conditions (pH 6–8), exhibits some tolerance to bile salts, and has survival rates of 38.89% and 92.22% in simulated gastric and intestinal fluids, respectively, indicating its potential as a probiotic. In a mouse model, Bacteroides fragilis intervention significantly alleviated colonic inflammation caused by Enterohemorrhagic Escherichia coli infection, enhanced tight junction protein expression, mitigated oxidative stress, and improved intestinal barrier function, with high-dose interventions showing superior effects. In lamb trials, Bacteroides fragilis intervention stopped diarrhea in four out of five lambs, partially restored intestinal microbiota diversity, and reduced the abundance of potential pathogens such as Aerococcus suis and Corynebacterium camporealensis. Therefore, Bacteroides fragilis exhibited remarkable effects in regulating intestinal homeostasis, alleviating inflammation, and promoting recovery from diarrhea.

Background: Mycoplasma infections are widespread globally, causing respiratory and extrapulmonary diseases in animals, particularly in sheep and goats. Symptoms include coughing, nasal discharge, respiratory distress, and pneumonia. Several Mycoplasma species like Mycoplasma ovipneumoniae (MO), Mycoplasma capricolum ssp. capripneumoniae (MCCP), Mycoplasma mycoides ssp. capri (MMC), and Mycoplasma arginini (MA) are identified as causes. MA was first identified in sheep with respiratory diseases in 1972, and its presence has been consistently reported. In China, MA isolation from respiratory samples was achieved in 1991, correlating with pulmonary lesions. Sheep and goats are economic resources in China. In recent years, China encourages the sheep and goat breeding model to convert into scale. Nevertheless, sheep and goats farming are threatened by Mycoplasma, which greatly slowed down the production. Currently, there is a scarcity of data and reports regarding Mycoplasma infection in sheep and goat farming in Jiangsu province. This study aims to determine the prevalence and species of Mycoplasma in sheep and goats in Jiangsu province. The results may contribute to the prevention and control of Mycoplasma infection in Jiangsu region, and will also expand our understanding of the distribution of these respiratory diseases in sheep and goats. Materials, Methods & Results: The study collected 920 nasal swab samples from sheep and goats in six cities in Jiangsu province. Following sample processing, DNA extraction was carried out using the lysis boiling method for both clinical samples and reference strains. Subsequently, PCR amplification was conducted utilizing universal and specific primers to detect and identify Mycoplasma species. Finally, the positive PCR products underwent sequencing and were compared for phylogenetic analysis. The results show that through using PCR specific primers to amplify genes associated with the Mycoplasma, 109 (11.85%) were confirmed to be Mycoplasma positive out of 920 samples that were collected from clinically diseased (470) and healthy (450) sheep and goats from Jiangsu province, China. 109 positive samples were obtained and classified as MO in 78.9% and MA in 21.1%. No MCCP and MMC were detected, and there was no evidence of mixed Mycoplasma infection. The phylogenetic analysis of the partial sequences of positive samples based on the MO p113 gene and MA ADI gene showed a high degree of similarity with some sequences of Mycoplasma in GenBank. Discussion: Among clinically healthy animals, 28 out of 450 samples (6.22%) tested positive, while among those with respiratory diseases, 81 out of 470 samples (17.23%) were positive, indicating a higher risk of respiratory disease in infected sheep and goats. Mycoplasma was found even in asymptomatic sheep and goats, suggesting it may not be the sole cause of respiratory symptoms. Immunized animals had lower infection rates, indicating some effectiveness of current vaccines, though overall efficacy remains modest. Regional variations in infection rates may stem from differences in vaccination and hygiene practices. Out of 109 positive samples, 86 were identified as MO infection and 23 as MA infection, while no infections of MCCP and MMC were detected. In this study, MO isolates lacked a central umbilicus in colonies, while MA isolates resembled "fried egg like" morphology. MO P113 gene sequencing revealed lower homology (86.0%) with certain strains compared to ATCC 29419, GZ-QX1, and NCTC 10151. This discrepancy could be ascribed to genetic mutations or disparities in pathogenic strains. MA ADI gene sequencing showed high homology (> 99.1%) with ATCC 23838, NCTC 10129, and HAZ145 strains. This indicates that prevalent Mycoplasma linked to respiratory diseases in Jiangsu province are MO and MA. Keywords: ovine, caprine, small ruminants, Mycoplasma, epidemiology, sequencing, hylogenic analysis.

In order to investigate the infection status and genotypes of Enterocytozoon bieneusi (E. bieneusi) in sheep and goats in Jiangsu Province, a total of 786 fresh fecal samples from 18 farms across five regions in Jiangsu were collected and examined for the presence of E. bieneusi, and the genotype of E. bieneusi was examined using nested-PCR and sequencing of the ribosomal internal transcribed spacer. The results showed that E. bieneusi was detected in the fecal samples of sheep and goats in all regions, with infection rates ranging from 23.65% to 42.81%. The overall infection rate was 36.51% (287/786). The infection rate of E. bieneusi showed no significant difference between sheep and goats, as well as among different ages of animals (p > 0.05), but showed a significant difference in sheep and goats with different health conditions (p < 0.05). The positive products were amplified and cloned and subjected to sequenced analysis. Six genotypes, BEB6, CHG2, CHG3, CHC8, CHG14, and COS-I, were found. Phylogenetic analysis indicated that the six genotypes belonged to Group 2, which had previously been described as a non-zoonotic group.