The objective of this study was to investigate the relationships between ruminal microbial communities and carcass traits associated with adipose accumulation in two Tibetan sheep breeds—Gannan and Tianzhu. A total of twenty Tibetan sheep (ten from each breed) were slaughtered, and samples of ruminal contents along with carcass trait data were collected for analysis. Ruminal microbial DNA was analyzed by 16S rRNA gene sequencing, and correlations between microbial composition and carcass traits were examined using correlation analysis and one-way ANOVA. The results showed that marbling score (p = 0.001) and longissimus lipid content (p = 0.007) were positively correlated with the Chao1 richness index, indicating that individuals with higher intramuscular fat content had greater ruminal microbial species richness. At the phylum level, Rikenellaceae RC9 gut group, Ruminococcaceae NK4A214 group were negatively correlated (p ≤ 0.05) with the above fat traits, whereas the abundance of the bacterial family Ruminococcus 1 was positively correlated with marbling score (p = 0.002). Stratified analysis by marbling grade further revealed associations with microbial richness (p ≤ 0.063), diversity (p = 0.044), and Ruminococcus 1 abundance (p < 0.001). However, microbial metabolic pathway prediction showed no significant differences (p ≥ 0.05) between the high- and low-marbling groups. In addition, several microbial taxa were positively correlated (p ≤ 0.05) with rib fat thickness and yield grade. In summary, ruminal microbial composition was closely associated with variations in carcass fat traits. Notably, most of the bacterial taxa associated with intramuscular and subcutaneous fat deposition did not overlap, suggesting that microbial metabolites may regulate fat deposition by influencing distinct adipogenic pathways in the host.

Extracellular vesicles (EVs) carry a wealth of functionally active molecules and play essential physiological roles in intercellular communication. However, little is known about the milk-derived EVs (mEVs) in yak colostrum (YC) and mature milk (YM). Herein, the Astral-DIA proteomics technique was applied to analyze and compare mEV proteins in yak milk. A total of 2311 mEV proteins were identified in yak milk, including 185 significant differentially expressed mEV proteins between both groups. Gene Ontology (GO) analysis showed that the differentially expressed mEV proteins were primarily linked to the innate immune response, calcium ion binding, and the extracellular space. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that complement and coagulation cascades were key pathways involved. This study offers a comprehensive understanding of the composition and functional properties of mEV proteins in yak milk across lactation stages and provides valuable guidance for the future development and application of mEV proteins.

Lignocellulosic biomass is a promising feedstock for biofuel production, but its complex structure, including cellulose and hemicellulose, challenges efficient enzymatic hydrolysis. CelyA, a novel cellulase from the yak rumen, has potential for improving biomass degradation and biofuel production. The CelyA gene was cloned, expressed, and purified. Biochemical characterization included assessments of pH, temperature, and salt tolerance. The enzyme's substrate specificity was tested on crystalline cellulose, CMC, and other polysaccharides. In vitro rumen fermentation was performed to evaluate its effect on fiber digestibility and microbial community composition. Biofuel production was tested by hydrolyzing maize, rice, and wheat straw. CelyA demonstrated optimal activity at pH 6.0 and 40.0 °C and maintained moderate stability across a wide pH range (3.0-12.0), retaining measurable activity even under strongly acidic and alkaline conditions. The enzyme demonstrated excellent salt tolerance, retaining 91.0 % activity in 1.0 M NaCl. CelyA efficiently degraded maize straw in hydrolysis assays, producing 7.2 µmol/L of reducing sugars. In vitro rumen fermentation with CelyA increased fiber digestibility by 8.3 % for maize straw, 14.5 % for rice straw, and 2.7 % for wheat straw. Gas production also increased significantly, with maize straw showing a 91.3 % increase. 16S rRNA sequencing revealed selective enrichment of Ruminococcus and Prevotella, key cellulolytic microbes. CelyA demonstrates strong potential for biofuel production, efficiently hydrolyzing lignocellulosic biomass and enhancing ruminal fiber digestibility. Its stability, salt tolerance, and substrate specificity make it a valuable enzyme for biofuel production and livestock feed optimization.

Gut barrier dysfunction is associated with dysbiosis of the gut microbiota and its metabolites, which is closely linked to the pathogenesis of ulcerative colitis (UC). Recent studies have demonstrated that Sulforaphane (SFN) exerts beneficial effects on UC. However, the role of the gut microbiota and microbial metabolism in the anti-UC mechanisms of SFN remains inadequately understood. In this study, we observed that SFN administration significantly improved the pathological phenotype, restored gut barrier integrity, and reduced colon inflammation in dextran sulfate sodium (DSS)-induced colitis mice. Gut microbiota analysis illustrated that SFN administration rebalances the alterations in gut microbiota composition, including genera such as Turicibacter, Lactobacillus and Bacteroides, in DSS-induced mice. Furthermore, untargeted metabolomics analysis indicated that the levels of microbial arachidonic acid metabolism, as well as the metabolism of alanine, aspartate, and glutamate, and glutathione metabolism in the gastrointestinal tract, were significantly altered in DSS-induced mice. Interestingly, SFN treatment significantly restore the alterations in glutathione metabolism and the levels of associated metabolites. Additionally, we observed that the MAPK/NF-κB signaling pathway, regulated by glutathione metabolism, was inhibited in the colon of DSS-induced mice following SFN treatment. Collectively, these results suggest that SFN can alleviate DSS-induced colitis in mice by restoring dysregulated gut microbiota and glutathione metabolism, thereby modulating the MAPK/NF-κB signaling pathway, enhancing intestinal barrier function and reducing colonic inflammation. Importantly, our findings elucidate a novel mechanism by which SFN improves gut barrier function, highlighting its potential to advance the development of SFN-derived therapeutics for the clinical management of colitis.

As one of the most representative natural products among flavonoids, quercetin (QUE) has been reported to exhibit beneficial effects on gut health in recent years. In this study, we utilized a dextran sulfate sodium (DSS)-induced colitis mice model to explore the protective effects and underlying mechanisms of QUE on colitis. Our data demonstrated that QUE oral gavage administration significantly ameliorates the symptoms and histopathological changes associated with colitis. Additionally, the concentration of mucin-2, the number of goblet cells, and the expression of tight junction proteins (such as ZO-1, Occludin, and Claudin-1) were all found to be increased. Furthermore, QUE treatment regulated the levels of inflammatory cytokines and macrophage polarization, as well as the oxidative stress-related pathway (Nrf2/HO-1) and associated enzymes. Additionally, 16S rDNA sequencing revealed that QUE treatment rebalances the alterations in colon microbiota composition (inlcuding Bacteroidaceae, Bacteroides, and Odoribacter) in DSS-induced colitis mice. The analysis of network dynamics reveals a significant correlation between gut microbial communities and microenvironmental factors associated with inflammation and oxidative stress, in conjunction with the previously mentioned findings. Collectively, our results suggest that QUE has the potential to treat colitis by maintaining the mucosal barrier, modulating inflammation, and reducing oxidation stress, which may depend on the reversal of gut microbiota dysbiosis.

This study analyzed the flavor compounds in the meat of four indigenous breeds of Chinese sheep through the use of gas chromatography–ion mobility spectrometry (GC-IMS). GC-IMS provided information on the characteristics and strength of 71 volatile flavor compounds (monomers and dimers), with aldehydes, alcohols and ketones being the most abundant in all types of sheep meat. The compounds with higher intensity peaks in the sheep meat were aldehydes (n-nonanal, octanal, heptanal, 3-methylbutanal, and hexanal), alcohols (1-octen-3-ol, hexanol, and pentanol), ketones (3-hydroxy-2-butanon, 2-butanone, and 2-propanone), esters (methyl benzoate), and thiazole (trimethylthiazole). The volatile flavor components in the meat of the different breeds of sheep obtained via GC-IMS were further differentiated using principal component analysis. In addition, orthogonal partial least squares discriminant analysis (OPLS-DA) and variable importance on projection (VIP) were used to determine the characteristic flavor compounds in the meats of different breeds of sheep, and 21 differentially volatile components were screened out based on having a VIP above 1. These results indicate that GC-IMS combined with multivariate analysis is a convenient and powerful method for characterizing and discriminating sheep meat.

Ruminant animals naturally emit methane gas owing to anaerobic microbial fermentation in the rumen, and these gases are considered major contributors to global warming. Scientists worldwide are attempting to minimize methane emissions from ruminant animals. Some of these attempts include the manipulation of rumen microbes using antibiotics, synthetic chemicals, dietary interventions, probiotics, propionate enhancers, stimulation of acetogens, manipulation of rumination time, vaccination, and genetic selection of animals that produce low methane (CH4). The majority of synthetic additives are harmful to both beneficial rumen microbes and the host or only temporarily affect methanogenesis. Phytogenic feed additives (PFAs) have recently emerged as the best alternatives to antibiotics and synthetic chemicals because of growing public concerns regarding drug resistance and the negative impacts of antibiotics and synthetic chemicals on humans, livestock, and the environment. These additives reduce methane production and improve the volatile fatty acid profile. In this review, we provide an overview of PFA sources and how their bioactive components affect the rumen microbiome to reduce methane emissions. Additionally, we highlight the mechanisms of action of PFAs as a whole, as well as some of their bioactive components. We also review some selected trees, herbs, shrubs, and forages and their roles in reducing methane emissions.

BackgroundInflammatory bowel disease (IBD) is a chronic and relapsing inflammatory disorders of the gastrointestinal tract, and currently the treatment options of IBD remain limited due to its complex pathogenesis. Quercetin (QUE), one of the representative flavonoids with various bioactivities, has many beneficial effects on gut health. However, little is known of its function on colitis. In this study, the protective effects and underlying mechanisms of QUE on colitis were investigated using dextran sulfate sodium (DSS)-induced mice by using 16S rDNA sequencing, immunofluorescence histochemistry, enzyme-linked immunoassay, respectively.ResultOur datas revealed that QUE administration by oral gavage not only ameliorates the symptoms and histopathological change of colitis, but also increases the concentration of mucin2, numbers of goblet cells and the expression of tight junction proteins, including ZO-1, Occludin, and Claudin-1. In addition, we also found that macrophage in colon were skewed to M1 macrophage polarization by DSS treatment, but QUE treatment markedly promoted M2 polarization in mice colon, while the levels of inflammatory cytokines (TNF-α, IL-1β, IL-6 and IL-10) were also changed, and received after QUE treatment. In addition, Immunofluorescence results also revealed that QUE upregulated the Nrf2/HO-1 signaling pathway, while the level of oxidative-stress enzymes (superoxide dismutase, glutathione, and catalase) were also reversed by QUE treatment. Furthermore, 16S rDNA sequencing revealed that QUE treatment partially restored the alterations in the gut microbiota in colitis mice, mainly by increasing the abundance of potentially beneficial bacteria (such as Odoribacter) whereas decreasing the abundance of potentially harmful bacteria (such as Bacteroidaceae and Bacteroides).ConclusionQUE had the capacity to treat colitis by maintaining the mucosal barrier, modulating inflammation, decreasing oxidation stress, and reverting gut microbiota changes. This research provides an ideal, healthful, and sustainable approach for IBD treatment.

Oral ingestion of probiotics is a promising approach to relieving inflammatory disease through regulating the gut microbiota. A newly discovered strain, Lactobacillus rhamnosus CY12 (LCY12), obtained from cattle-yak milk, displayed numerous probiotic properties. These included enhanced viability in low pH and bile environments, adhesion capabilities, and potent antimicrobial effects. The research aimed to explore the beneficial impacts of the novel LCY12 strain on colitis in mice induced by dextran sulfate sodium (DSS) and to elucidate the underlying molecular mechanisms. The results of the study showed that administration of LCY12 effectively helped to reduce the negative effects of DSS-induced body weight loss, disease activity index score, colon length shortening, loss of goblet cells, and overall histopathological scores in the intestines. Simultaneously, LCY12 administration significantly alleviated intestinal inflammation and safeguarded intestinal barrier integrity by enhancing IL-10 levels, while dampening IL-6, IL-1β, and TNF-α production. Additionally, LCY12 boosted the presence of tight junction proteins. Furthermore, LCY12 hindered the TLR4/MyD88/NF-κB signaling pathway by downregulating TLR4 and MyD88 expression, inactivating phosphorylated IκBα, and preventing translocation of NF-κB p65 from the cytoplasm to the nucleus. The LCY12 also increased specific intestinal microbial communities and short-chain fatty acid (SCFA) production. Altogether, LCY12 oral administration alleviated colitis induced with DSS in mice by improving intestinal barrier function and regulating inflammatory cytokines, SCFA production, and intestinal microbiota.

Atmospheric and room temperature plasma (ARTP) is an emerging artificial mutagenesis breeding technology. In comparison to traditional physical and chemical methods, ARTP technology can induce DNA damage more effectively and obtain mutation strains with stable heredity more easily after screening. It possesses advantages such as simplicity, safety, non-toxicity, and cost-effectiveness, showing high application value in microbial breeding. This article focuses on ARTP mutagenesis breeding of actinomycetes, specifically highlighting the application of ARTP mutagenesis technology in improving the performance of strains and enhancing the biosynthetic capabilities of actinomycetes. We analyzed the advantages and challenges of ARTP technology in actinomycetes breeding and summarized the common features, specific mutation sites and metabolic pathways of ARTP mutagenic strains, which could give guidance for genetic modification. It suggested that the future research work should focus on the establishment of high throughput rapid screening methods and integrate transcriptomics, proteomics, metabonomics and other omics to delve into the genetic regulations and synthetic mechanisms of the bioactive substances in ARTP mutated actinomycetes. This article aims to provide new perspectives for actinomycetes breeding through the establishment and application of ARTP mutagenesis technology, thereby promoting source innovation and the sustainable industrial development of actinomycetes.

Xenorhabdus , known for its symbiotic relationship with Entomopathogenic nematodes (EPNs), belongs to the Enterobacteriaceae family. This dual-host symbiotic nematode exhibits pathogenic traits, rendering it a promising biocontrol agent against insects. Our prior investigations revealed that Xenorhabdus stockiae HN_xs01, isolated in our laboratory, demonstrates exceptional potential in halting bacterial growth and displaying anti-tumor activity. Subsequently, we separated and purified the supernatant of the HN_xs01 strain and obtained a new compound with significant inhibitory activity on tumor cells, which we named XNAE. Through LC-MS analysis, the mass-to-nucleus ratio of XNAE was determined to be 254.24. Our findings indicated that XNAE exerts a time- and dose-dependent inhibition on B16 and HeLa cells. After 24 h, its IC 50 for B16 and HeLa cells was 30.178 µg/mL and 33.015 µg/mL, respectively. Electron microscopy revealed conspicuous damage to subcellular structures, notably mitochondria and the cytoskeleton, resulting in a notable reduction in cell numbers among treated tumor cells. Interestingly, while XNAE exerted a more pronounced inhibitory effect on B16 cells compared to HeLa cells, it showed no discernible impact on HUVEC cells. Treatment of B16 cells with XNAE induced early apoptosis and led to cell cycle arrest in the G2 phase, as evidenced by flow cytometry analysis. The impressive capability of X. stockiae HN_xs01 in synthesizing bioactive secondary metabolites promises to significantly expand the reservoir of natural products. Further exploration to identify the bioactivity of these compounds holds the potential to shed light on their roles in bacteria-host interaction. Overall, these outcomes underscore the promising potential of XNAE as a bioactive compound for tumor treatment.

Purpose: This study aimed to investigate the prevalence and genetic characterization of enterococcal isolates (Enterococcus faecalis, Enterococcus faecium and Enterococcus hirae) isolated from clinical bovine mastitis cases in Ningxia, China. Patients and Methods: The enterococci were identified by 16S rRNA amplification and sequencing. Antimicrobial resistance was determined by disc diffusion method. Virulence and antimicrobial resistance genes were detected by PCR assays. Results: Overall, 198 enterococcal isolates were identified from 2897 mastitis samples, including 137 (4.7%) E. faecalis, 50 (1.7%) E. faecium and 11 (0.4%) E. hirae. E. faecalis, E. faecium and E. hirae isolates showed high resistance to tetracycline (92.7%, 68.0%, 90.9%), followed by erythromycin (86.9%, 76.0%, 72.7%). The multidrug-resistant strains of E. faecalis and E. faecium were 29 (21.2%) and 13 (26.0%), respectively. The resistance of E. faecalis, E. faecium and E. hirae isolates to tetracycline is mainly attributed to the presence of tetL (alone or combined with tetM and/or tetK), the erythromycin resistance to ermB (alone or combined with ermC and/or ermA). Moreover, cpd (94.2%), gelE (77.4%), efaAfs (93.4%), and esp (79.6%) were the most common virulence genes in E. faecalis. In E. faecium, except for the gene efaAfs (82.0%), other virulence genes are rarely found. Only two strains of E. hirae carrying asa1 gene were detected. Conclusion: The results of this study can provide a reference for the prevention and treatment of bovine mastitis caused by enterococci.

Ruminant animals naturally emit methane gas owing to anaerobic microbial fermentation in the rumen, and these gases are considered major contributors to global warming. Scientists worldwide are attempting to minimize methane emissions from ruminant animals. Some of these attempts include the manipulation of rumen microbes using antibiotics, synthetic chemicals, dietary interventions, probiotics, propionate enhancers, stimulation of acetogens, manipulation of rumination time, vaccination, and genetic selection of animals that produce low methane (CH4). The majority of synthetic additives are harmful to both beneficial rumen microbes and the host or only temporarily affect methanogenesis. Phytogenic feed additives (PFAs) have recently emerged as the best alternatives to antibiotics and synthetic chemicals because of growing public concerns regarding drug resistance and the negative impacts of antibiotics and synthetic chemicals on humans, livestock, and the environment. These additives reduce methane production and improve the volatile fatty acid profile. In this review, we provide an overview of PFA sources and how their bioactive components affect the rumen microbiome to reduce methane emissions. Additionally, we highlight the mechanisms of action of PFAs as a whole as well as some of their bioactive components.

IntroductionPostpartum period of dairy cattle is an important phase of their life mainly associated with the changes in physiology, rumen function, and energy metabolism. Studies have shown that gut microbial composition undergoes drastic changes during the postpartum period. However, little is known about the temporal variations in digestive tract microbiota in postpartum Tibetan cattle. The aim of this study was to investigate the temporal variations in blood metabolites, ruminal fermentation, and microbial community of oral, rumen, and gut in lactating Tibetan cattle during postpartum.MethodsWe collected blood, saliva, rumen fluid, and fecal samples from lactating Tibetan cattle during 1st week (1 W), the 2nd week (2 W), the 1st month (1 M), and the 2nd month (2 M) of the postpartum period. The microbiota of saliva, rumen fluid, and fecal samples were assessed using 16S rRNA sequencing. The rumen volatile fatty acid and blood parameters were also quantified.ResultsThe content of volatile fatty acids (VFAs) and blood parameters showed opposite tendency to each other and reached to stability at 2 M. Rumen microbiota showed the highest alpha diversity compared to other two sites. At phylum level, the oral cavity was dominated by Proteobacteria, while most dominant phylum in rumen and feces were Firmicutes and Bacteroidetes, respectively. The dominant genera in oral cavity were Moraxella and Bibersteinia, while genera Prevotella 1 and Ruminococcaceae UCG-005 were dominant in rumen and fecal samples, respectively.DiscussionMicrobial network analysis revealed that most of the active genera in all networks belonged to phylum Firmicutes, indicating the importance of this phyla during postpartum period of lactating cattle. The functional analysis revealed distinct division of labor among three gastrointestinal sites associated with defense, fatty acid synthesis, and maintaining health of host. All in all, our findings provide insights into the metabolic and microbial changes of lactating Tibetan cattle and help to the improvement of the management strategies.

Inflammatory bowel disease (IBD) is a serious global public health concern. Although the pathogenesis of the disease is currently unknown, it has been reported to be associated with both intestinal microbiota and inflammatory mediators. There is evidence suggesting that the feces of the Plateau pika is useful for treating gastrointestinal injuries and pain. Although fecal microbiota transplantation is highly efficacious intervention for IBD prevention, however, potential the transfer of pathogenic microbes or toxic substances is potentially hazardous. Fortunately, micropore filtering of the donor feces can minimize the risk of bacterial infection allowing retention of the therapeutic effects of the residual bacteriophages. Here, we demonstrated that Plateau pika feces not only alleviated the IBD symptoms but also promoted optimal structure and composition of the intestinal microbiota. Additionally, Plateau pika feces transfer also enhanced phenotypic features, such as, body-weight, disease activity index, and histological scores. In conclusion, Plateau pika feces was found to protect mice against colitis induced by dextran sodium sulfate by reducing inflammation and regulating microbial dysbiosis. These findings suggest the potential of Plateau pika feces as an alternative therapy for IBD.

Staphylococcus haemolyticus (S. haemolyticus) is one of the most common coagulase-negative staphylococci (CoNS) isolates from bovine mastitis. Paeoniflorin (PF) shows anti-inflammatory effects on different inflammatory diseases in vitro studies and in vivo animal experiments. In this study, the viability of bovine mammary epithelial cells (bMECs) was detected by the cell counting kit-8 experiment. Subsequently, bMECs were induced with S. haemolyticus, and the induction dosage was determined. The expression of pro-inflammatory cytokines and toll-like receptor (TLR2) and nuclear factor kappa-B (NF-κB) signaling pathway-related genes were investigated by quantitative real-time PCR. The critical pathway proteins were detected by western blot. The results showed that the multiplicity of infection (MOI; the ratio of bacteria to bMECs) 5:1 of S. haemolyticus for 12 h could cause cellular inflammation, which was selected to establish the inflammatory model. Incubation with 50 μg/ml PF for 12 h was the best intervention condition for cells stimulated by S. hemolyticus. Quantitative real-time PCR and western blot analysis showed that PF inhibited the activation of TLR2 and NF-κB pathway-related genes and the expression of related proteins. Western blot results showed that PF suppressed the expression of NF-κB unit p65, NF-κB unit p50, and MyD88 in bMECs stimulated by S. haemolyticus. The inflammatory response pathway and molecular mechanism caused by S. haemolyticus on bMECs are related to TLR2-mediated NF-κB signaling pathways. The anti-inflammatory mechanism of PF may also be through this pathway. Therefore, PF is expected to develop potential drugs against CoNS-induced bovine mastitis.

Anaerobic fungi (AF) inhabit the gastrointestinal tract of ruminants and play an important role in the degradation of fiber feed. However, limited knowledge is available on seasonal dynamics and inter-species differences in rumen AF community in yak and cattle under natural grazing systems. Using the random forests model, the null model, and structural equation model, we investigated the seasonal dynamics and key driving factors of fiber-associated rumen AF in grazing yak and cattle throughout the year on the Qinghai-Tibet Plateau (QTP). We found that the richness and diversity of rumen AF of grazing yak and cattle in cold season were significantly higher than those in warm season (P < 0.05). We identified 12 rumen AF genera, among which , Cyllamyces, and Orpinomyces were predominant in the rumen of both grazing yak and cattle. LEfSe and random forest analysis showed that Feramyces, Tahromyces, and Buwchfawromyces were important seasonal indicator of rumen AF in grazing yak (P < 0.05), and Caecomyces, Cyllamyces, and Piromyces in grazing cattle (P < 0.05). Null model analysis revealed that the dynamic changes of rumen AF community structure were mainly affected by deterministic factors. Notably, mantel test and structural equation model revealed that forage physical-chemical properties, including dry matter (DM), neutral detergent fiber (NDF), and hemicellulose contents (HC) were the key factors driving the seasonal variations of the rumen AF community (P < 0.05). The results revealed that forage lignocellulose was probably an important factor affecting the seasonal dynamics and inter-species differences of the rumen AF community under natural grazing conditions. IMPORTANCE The seasonal dynamics of rumen anaerobic fungi in nature grazing yak and cattle were determined during cold and warm seasons based on pasture nutritional quality and environmental data sets. The main driving factors of anaerobic fungi in yak and cattle rumen were explored by combining random forest and structural equation models. In addition, the dynamic differences in the composition of the anaerobic fungi community in the yak and cattle in different seasons were characterized. It was found that some rumen anaerobic fungi have contributed to high fiber degradation rate in yak. These novel findings improve our understanding of the association of environmental and dietary seasonal variations with anaerobic fungal community, facilitating yak adaptation to high altitude.

Staphylococci, mainly including Staphylococcus aureus and coagulase-negative staphylococci (CNS), are one of the most common pathogens causing bovine mastitis worldwide. In this study, we investigated the antimicrobial resistance and virulence profiles of staphylococci from clinical bovine mastitis in Ningxia Hui Autonomous Region of China. Antimicrobial resistance was determined by disc diffusion combined with E-test method. Genes of antimicrobial resistance and virulence factors were determined by PCR. A total of 332 staphylococcal isolates were confirmed from 1,519 mastitic milk samples, including 172 S. aureus and 160 CNS isolates. Fifteen CNS species were identified, with S. chromogenes being the most frequent found (49.4%), followed by S. equorum (13.8%). Noticeably, 2 S. agnetis isolates were found among the CNS isolates. To our knowledge, this is the first report documenting the presence of S. agnetis from bovine mastitis in China. The S. aureus and CNS isolates showed high resistance against penicillin, followed by erythromycin and tetracycline. Multidrug resistance was found in 11.6 and 16.3% of the S. aureus and CNS isolates, respectively. Resistance to penicillin was attributed to the presence of blaZ, erythromycin resistance to ermC (alone or combined with ermB) and tetracycline resistance to tetK (alone or combined with tetM). Notably, one S. equorum isolate and one S. saprophyticus isolate were both methicillin-resistant and mecA positive. Additionally, all S. aureus isolates carried the adhesin genes fnbpA, clfA, clfB, and sdrC, and most of them contained cna and sdrE. Conversely, only a few of the CNS isolates carried clfA, cna, and fnbA. Regarding toxin genes, all S. aureus isolates harbored hlb, and most of them were hlg positive. The lukE-lukD, lukM, sec, sed, sei, sen, seo, tst, seg, seh, and sej were also detected with low frequencies. However, no toxin genes were observed in CNS isolates. This study reveals high species diversity of staphylococci from clinical bovine mastitis in Ningxia Hui Autonomous Region of China. The findings for the genetic determinants of antimicrobial resistance and virulence factor provide valuable information for control and prevention of staphylococcal bovine mastitis.

Purpose:This study aimed to investigate the prevalence and genetic characterization of Serratia marcescens isolates from clinical bovine mastitis in Ningxia Hui Autonomous Region of China.Methods:S. marcescens was identified by the polymerase-chain reaction of 16S rRNA gene and sequencing. Antimicrobial susceptibility was tested by the disk diffusion method. Genes of resistance and virulence were determined by the PCR.Results:Overall, S. marcescens were confirmed from 32 of 2897 (1.1%) mastitis milk samples. These isolates showed high resistance to cefazolin (30/32, 93.8%) and chloramphenicol (28/32, 87.5%). A 12.5% (4/32) of the isolates displayed multidrug resistance (MDR). The most prevalent resistant genes found in S. marcescens were TEM (32/32, 100%) and CTX-M (24/32, 75.0%; CTX-M-15, 14/32, 43.8%; CTX-M-14, 8/32, 25.0%; CTX-M-65, 2/32, 6.3%) for extended-spectrum beta-lactamase, cmlA (28/32, 87.5%) and floR (16/32, 50.0%) for chloramphenicol resistance, SIM-1 (2/32, 6.3%) for carbapenemases, and sdeB (28/32, 87.5%), sdeY (26/32, 81.3%), sdeR (26/32, 81.3%) and sdeD (20/32, 62.5%) for efflux pumps. Moreover, all isolates carried virulence genes flhD, entB, and kpn, and most of them contained mrkD (30/32, 93.8%), ycfM (26/32, 81.3%), bsmB (26/32, 81.3%), pigP (26/32, 81.3%), kfu (24/32, 75.0%) and shlB (24/32, 75.0%).Conclusion:To our knowledge, this is the first report of genetic determinants for antimicrobial resistance and virulence in S. marcescens isolated from bovine mastitis cases in China. These findings are useful for developing strategies for prevention and treatment of bovine mastitis caused by S. marcescens in China.