Orally Administered Lactobacillus rhamnosus CY12 Alleviates DSS-Induced Colitis in Mice by Restoring the Intestinal Barrier and Inhibiting the TLR4-MyD88-NF-κB Pathway via Intestinal Microbiota Modulation.

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-1ß, 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.

Discovery of an antitumor compound from xenorhabdus stockiae HN_xs01.

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 IC50 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.

Precision enzyme discovery through targeted mining of metagenomic data.

Metagenomics has opened new avenues for exploring the genetic potential of uncultured microorganisms, which may serve as promising sources of enzymes and natural products for industrial applications. Identifying enzymes with improved catalytic properties from the vast amount of available metagenomic data poses a significant challenge that demands the development of novel computational and functional screening tools. The catalytic properties of all enzymes are primarily dictated by their structures, which are predominantly determined by their amino acid sequences. However, this aspect has not been fully considered in the enzyme bioprospecting processes. With the accumulating number of available enzyme sequences and the increasing demand for discovering novel biocatalysts, structural and functional modeling can be employed to identify potential enzymes with novel catalytic properties. Recent efforts to discover new polysaccharide-degrading enzymes from rumen metagenome data using homology-based searches and machine learning-based models have shown significant promise. Here, we will explore various computational approaches that can be employed to screen and shortlist metagenome-derived enzymes as potential biocatalyst candidates, in conjunction with the wet lab analytical methods traditionally used for enzyme characterization.

Plateau pika fecal microbiota transplantation ameliorates inflammatory bowel disease manifestations in a mouse model of colitis.

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.

Forage lignocellulose is an important factor in driving the seasonal dynamics of rumen anaerobic fungi in grazing yak and cattle.

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.

Heritability and recursive influence of host genetics on the rumen microbiota drive body weight variance in male Hu sheep lambs.

BACKGROUND: Heritable rumen microbiota is an important modulator of ruminant growth performance. However, no information exists to date on host genetics-rumen microbiota interactions and their association with phenotype in sheep. To solve this, we curated and analyzed whole-genome resequencing genotypes, 16S rumen-microbiota data, and longitudinal body weight (BW) phenotypes from 1150 sheep. RESULTS: A variance component model indicated significant heritability of rumen microbial community diversity. Genome-wide association studies (GWAS) using microbial features as traits identified 411 loci-taxon significant associations (P < 10-8). We found a heritability of 39% for 180-day-old BW, while also the rumen microbiota likely played a significant role, explaining that 20% of the phenotypic variation. Microbiota-wide association studies (MWAS) and GWAS identified four marker genera (Bonferroni corrected P < 0.05) and five novel genetic variants (P < 10-8) that were significantly associated with BW. Integrative analysis identified the mediating role of marker genera in genotype influencing phenotype and unravelled that the same genetic markers have direct and indirect effects on sheep weight. CONCLUSIONS: This study reveals a reciprocal interplay among host genetic variations, the rumen microbiota and the body weight traits of sheep. The information obtained provide insights into the diverse microbiota characteristics of rumen and may help in designing precision microbiota management strategies for controlling and manipulating sheep rumen microbiota to increase productivity. Video Abstract.

Antimicrobial resistance and virulence profiles of staphylococci isolated from clinical bovine mastitis.

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.

Prevalence and Characterization of Serratia marcescens Isolated from Clinical Bovine Mastitis Cases in Ningxia Hui Autonomous Region of China.

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.

Lignocellulose degradation by rumen bacterial communities: New insights from metagenome analyses.

Ruminant animals house a dense and diverse community of microorganisms in their rumen, an enlarged compartment in their stomach, which provides a supportive environment for the storage and microbial fermentation of ingested feeds dominated by plant materials. The rumen microbiota has acquired diverse and functionally overlapped enzymes for the degradation of plant cell wall polysaccharides. In rumen Bacteroidetes, enzymes involved in degradation are clustered into polysaccharide utilization loci to facilitate coordinated expression when target polysaccharides are available. Firmicutes use free enzymes and cellulosomes to degrade the polysaccharides. Fibrobacters either aggregate lignocellulose-degrading enzymes on their cell surface or release them into the extracellular medium in membrane vesicles, a mechanism that has proven extremely effective in the breakdown of recalcitrant cellulose. Based on current metagenomic analyses, rumen Bacteroidetes and Firmicutes are categorized as generalist microbes that can degrade a wide range of polysaccharides, while other members adapted toward specific polysaccharides. Particularly, there is ample evidence that Verrucomicrobia and Spirochaetes have evolved enzyme systems for the breakdown of complex polysaccharides such as xyloglucans, peptidoglycans, and pectin. It is concluded that diversity in degradation mechanisms is required to ensure that every component in feeds is efficiently degraded, which is key to harvesting maximum energy by host animals.

Paeoniflorin alleviates inflammation in bovine mammary epithelial cells induced by Staphylococcus haemolyticus through TLR2/NF-κB signaling pathways.

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.

Efficiency of an alkaline, thermostable, detergent compatible, and organic solvent tolerant lipase with hydrolytic potential in biotreatment of wastewater.

Discharging the tannery wastewater into the environment is a serious challenge worldwide due to the release of severe recalcitrant pollutants such as oil compounds and organic materials. The biological treatment through enzymatic hydrolysis is a cheap and eco-friendly method for eliminating fatty substances from wastewater. In this context, lipases can be utilized for bio-treatment of wastewater in multifaceted industrial applications. To overcome the limitations in removing pollutants in the effluent, we aimed to identify a novel robust stable lipase (PersiLipase1) from metagenomic data of tannery wastewater for effective bio-degradation of the oily wastewater pollution. The lipase displayed remarkable thermostability and maintained over 81 % of its activity at 60 °C.After prolonged incubation for 35 days at 60°C, the PersiLipase1 still maintained 53.9 % of its activity. The enzyme also retained over 67 % of its activity in a wide range of pH (4.0 to 9.0). In addition, PersiLipase1 demonstrated considerable tolerance toward metal ions and organic solvents (e.g., retaining >70% activity after the addition of 100 mM of chemicals). Hydrolysis of olive oil and sheep fat by this enzyme showed 100 % efficiency. Furthermore, the PersiLipase1 proved to be efficient for biotreatment of oil and grease from tannery wastewater with the hydrolysis efficiency of 90.76 % ± 0.88. These results demonstrated that the metagenome-derived PersiLipase1 from tannery wastewater has a promising potential for the biodegradation and management of oily wastewater pollution.

Circadian orchestration of host and gut microbiota in infection.

Circadian rhythms are present in almost every organism and regulate multiple aspects of biological and physiological processes (e.g. metabolism, immune responses, and microbial exposure). There exists a bidirectional circadian interaction between the host and its gut microbiota, and potential circadian orchestration of both host and gut microbiota in response to invading pathogens. In this review, we summarize what is known about these intestinal microbial oscillations and the relationships between host circadian clocks and various infectious agents (bacteria, fungi, parasites, and viruses), and discuss how host circadian clocks prime the immune system to fight pathogen infections as well as the direct effects of circadian clocks on viral activity (e.g. SARS-CoV-2 entry and replication). Finally, we consider strategies employed to realign normal circadian rhythmicity for host health, such as chronotherapy, dietary intervention, good sleep hygiene, and gut microbiota-targeted therapy. We propose that targeting circadian rhythmicity may provide therapeutic opportunities for the treatment of infectious diseases.

Ascorbic Acid-Mediated Modulation of Antibiotic Susceptibility of Major Bovine Mastitis Pathogens.

This study aimed to investigate the effects of ascorbic acid on antibiotic susceptibility of major bovine mastitis pathogens, including Staphylococcus aureus, Streptococcus dysgalactiae, Streptococcus uberis, Streptococcus agalactiae, and Escherichia coli. Minimum inhibitory concentrations (MICs) were determined by E-test method. The presence of 10 mM ascorbic acid decreased the MICs of penicillin and ampicillin while increased the MICs of erythromycin, kanamycin, streptomycin, and ciprofloxacin for all tested strains. Besides, ascorbic acid specifically reduced the MICs of tetracycline for gram-positive bacteria and chloramphenicol for gram-negative bacteria. This study highlights that ascorbic acid is a potential modulator of antibiotic activity against the major bovine mastitis pathogens.

Regulation of yak longissimus lumborum energy metabolism and tenderness by the AMPK/SIRT1 signaling pathways during postmortem storage.

AMPK can activate nicotinamide phosphoribosyltransferase (NAMPT), increasing the ratio of oxidized nicotinamide adenine dinucleotide (NAD+)/reduced nicotinamide adenine dinucleotide (NADH) ratio, leading to the activation of the energy receptor SIRT1. This pathway is known as the AMPK/SIRT1 signaling pathway. SIRT1 deacetylates and activate LKB1, which is activated by phosphorylation of AMPK (Thr172) and inhibited by phosphorylase-mediated dephosphorylation of AMPK. At the same time, increased AMP/ATP and NAD+/NADH ratios lead to the activation of AMPK and SIRT1. SIRT1 and AMPK can activate each other forming a positive feedback loop, which can strengthen catabolism and weaken anabolism thus maintaining energy homeostasis of energy metabolism. At present, there has been no systematic study on AMPK-associated signaling cascades in stored yak meat and details of the AMPK/SIRT1 signaling under these conditions are not known. In this study, NAD+, NADH were added to yak longissimus thoracic muscles to study AMPK pathway regulation by AMPK/SIRT1 signaling. NAD+ significantly increased the activity of AMPK and glycolysis during postmortem maturation, increased the rate of energy metabolism, and increased the expression of AMPK protein, indicating that NAD+ increased energy metabolism in the stored muscle by promoting AMPK activity. NADH treatment inhibited both AMPK activation and glycolysis, together with increasing the pH in the muscle. The results showed that SIRT1 activation elevated the activity of AMPK, leading to its phosphorylation and the activation of glycolysis. Thus, AMPK activity was found to increase in yak meat as an adaptation to hypoxic conditions. This allows more effective regulation of energy production and improves the tenderness of the meat.

Neutrophil Extracellular Traps Mediate Bovine Endometrial Epithelial Cell Pyroptosis in Dairy Cows with Endometritis.

Neutrophils are involved in the development of endometritis, but it remains unknown how neutrophils induce inflammation and tissue damage. Neutrophil extracellular traps (NETs) clear invading pathogens during infection but induce pyroptosis, leading to inflammation and tissue damage. Thus, our objective was to investigate whether NETs participate in bovine endometrial epithelial cell (BEEC) pyroptosis during endometritis. To confirm this, NETs and caspase-1/4; apoptosis-associated speck-like protein containing a caspase-recruitment domain(ASC); nod-like receptor protein-3 (NLRP3); and gasdermin D N-terminal (GSDMD-N), TNF-a, IL-1ß, IL-6, and IL-18 in endometrial tissue were detected. Pathological section and vaginal discharge smears were performed to visually determine endometritis in the uterus. BEECs were stimulated with NETs to induce pyroptosis, which was treated with DNase I against pyroptosis. Caspase-1/4, ASC, NLRP3, GSDMD-N, TNF-a, IL-1ß, IL-6, and IL-18 in BEECs were analyzed in endometrial tissue. The results showed that NET formation, as well as pyroptosis-related proteins and proinflammatory, cytokines were elevated in the endometrial tissue of cows with endometritis. Pathological sections and vaginal discharge smears showed increased neutrophils and plasma cells in the uterus, as well as tissue congestion. In BEECs, NETs increased the level of pyroptosis-related proteins and proinflammatory cytokines and were diminished by DNase I. In summary NETs participate BEEC pyroptosis during endometritis in dairy cows.

Recombineering using RecET-like recombinases from Xenorhabdus and its application in mining of natural products.

Xenorhabdus can produce a large number of secondary metabolites with insecticidal, bacteriostatic, and antitumor activities. Efficient gene editing tools will undoubtedly facilitate the functional genomics research and bioprospecting in Xenorhabdus. In this study, BlastP analysis using the amino acid sequences of Redαß or RecET recombinases as queries resulted in the identification of an operon (XBJ1_operon 0213) containing RecET-like recombinases encoding genes from the genome of Xenorhabdus bovienii strain SS-2004. Three proteins encoded by this operon was indispensable for full activity of recombineering, namely XBJ1-1173 (RecE-like protein), XBJ1-1172 (RecT-like protein), and XBJ1-1171 (single-strand annealing protein). Using this newly developed recombineering system, a gene cluster responsible for biosynthesis of a novel secondary metabolite (Min16) was identified from X. stockiae HN_xs01 strain. Min16 which exhibited antibacterial and cytotoxic activities was determined to be a cyclopeptide composed of Acyl-Phe-Thr-Phe-Pro-Pro-Leu-Val by using high-resolution mass spectrometry and nuclear magnetic resonance analysis, and was designated as changshamycin. This host-specific recombineering system was proven to be effective for gene editing in Xenorhabdus, allowing for efficient discovery of novel natural products with attractive bioactivities. KEY POINTS: • Screening and identification of efficient gene editing tools from Xenorhabdus • Optimization of the Xenorhabdus electroporation parameters • Discovery of a novel cyclopeptide compound with multiple biological activities.

Lactobacillus rhamnosus CY12 Enhances Intestinal Barrier Function by Regulating Tight Junction Protein Expression, Oxidative Stress, and Inflammation Response in Lipopolysaccharide-Induced Caco-2 Cells.

The intestinal barrier is vital for preventing inflammatory bowel disease (IBD). The objectives of this study were to assess whether the Lactobacillus rhamnosus CY12 could alleviate oxidative stress, inflammation, and the disruption of tight junction (TJ) barrier functions induced by lipopolysaccharide (LPS), and therefore to explore the potential underlying molecular mechanisms. Our results showed that LPS-induced Cancer coli-2 (Caco-2) cells significantly increased the levels of reactive oxygen species (ROS), lactate dehydrogenase, inflammatory cytokines interleukin-1ß, interleukin-6, interleukin-8, and tumor necrosis factor-α (IL-1ß, IL-6, IL-8, and TNF-α), and the cell apoptosis rate while decreasing the levels of TJ proteins occludin, zonula occludens-1 (ZO-1), and claudin and antioxidant enzymes, such as catalase, superoxide dismutase, and glutathione peroxidase(CAT, SOD, and GSH-Px) (p < 0.05). However, Lactobacillus rhamnosus CY12 could relieve cytotoxicity, apoptosis, oxidative stress, and pro-inflammatory cytokine expressions, and also inhibit the Toll-like receptor 4/nuclear factor kappa-B(TLR4/NF-κB) signaling pathway. Furthermore, the gene expression of antioxidant enzymes, as well as the mRNA and protein expressions of TJ proteins, was improved. Particularly, the concentration of 108 cfu/mL significantly prevented the inflammatory injury induced by LPS in Caco-2 cells (p < 0.05). These findings support a potential application of Lactobacillus rhamnosus CY12 as a probiotic to prevent LPS-induced intestinal injury and treat intestinal barrier dysfunction.

Age-dependent variations in rumen bacterial community of Mongolian cattle from weaning to adulthood.

BACKGROUND: Rumen microbes play an important role in ruminant energy supply and animal performance. Previous studies showed that the rumen microbiome of Mongolian cattle has adapted to degrade the rough forage to provide sufficient energy to tolerate the harsh desert ecological conditions. However, little is known about the succession of rumen microbes in different developmental stages of post-weaning Mongolian cattle. METHODS: Here, we examined the succession of the rumen microbial composition and structure of 15 post-weaning Mongolian cattle at three developmental stages i.e., 5 months (RM05), 18 months (RM18) and, 36 months (RM36) by using the 16S rRNA gene sequencing method. RESULTS: We did not find any age-dependent variations in the ruminal concentrations of any volatile fatty acid (VFA) of Mongolian cattle. The diversity of the rumen bacterial community was significantly lower in RM05 group, which reached to stability with age. Bacteroidetes and Firmicutes were the two dominant phyla among all age groups. Phylum Actinobacteria was significantly higher in RM05 group, phyla Spirochaetes, and Tenericutes were highly abundant in RM18 group, and phyla Proteobacteria and Epsilonbacteraeota were enriched in RM36 group. Genera Prevotella_1, Bacteroides, and Bifidobacterium were abundant in RM05 group. The short chain fatty acid (SCFA) producing bacteria Rikenellaceae_RC9_gut_group showed high abundance in RM18 group and fiber degrading genus Alloprevotella was highly abundant in RM36 group. Random forest analysis identified Alloprevotella, Ileibacterium, and Helicobacter as important age discriminatory genera. In particular, the genera Ruminococcaceae_UCG-005, Bacteroides, Saccharofermentans, and Fibrobacter in RM05, genera [Eubacterium] coprostanoligenes_group, Erysipelotrichaceae_UCG-004, Helicobacter, Saccharofermentans, Papillibacter, and Turicibacter in RM18, and genera Rikenellaceae_RC9_gut_group, Lachnospiraceae_AC2044_group, and Papillibacter in RM36 showed the top interactions values in the intra-group interaction network. CONCLUSIONS: The results showed that rumen microbiota of Mongolian cattle reached to stability and maturity with age after weaning. This study provides some theoretical evidence about the importance of functional specific rumen bacteria in different age groups. Further studies are needed to determine their actual roles and interactions with the host.

A novel tumor-targeting strain of Xenorhabdus stockiae exhibits potent biological activities.

Xenorhabdus are symbionts of soil entomopathogenic nematodes of the genus Steinernema presenting two distinct forms in their life cycle, and can produce a broad range of bioactive compounds. In this study, a novel Xenorhabdus stockiae strain HN_xs01 was isolated from a soil sample via an entrapment method using Galleria melonella nematodes. The supernatants of strain HN_xs01 exhibited antimicrobial properties against Gram-negative and Gram-positive bacteria, and insecticidal properties against Helicoverpa armigera larvae, and antitumor properties as well. Moreover, three linear rhabdopeptides (1, 2 and 3) were identified from strain HN_xs01 using nuclear magnetic resonance analysis, which exhibited significant cytotoxic activity against the human epithelial carcinoma cell line A431 and the human chronic myelogenous leukemia cell line K562. Some bacteria have been reported to colonize the tumor region, and we determined that HN_xs01 could grow in tumor xenografts in this study. HN_xs01 invaded and replicated in B16 melanoma cells grafted into C57BL/6 mice, resulting in tumor inhibition. Moreover, strain HN_xs01 not only merely aggregated in the tumor environment, but also prevented pulmonary metastasis. It caused fragmentation of vessels and cell apoptosis, which contributed to its antitumor effect. In conclusion, X. stockiae HN_xs01 is a novel tumor-targeting strain with potential applications in medicinal and agricultural fields.

Genomic analyses of wild argali, domestic sheep, and their hybrids provide insights into chromosome evolution, phenotypic variation, and germplasm innovation.

Understanding the genetic mechanisms of phenotypic variation in hybrids between domestic animals and their wild relatives may aid germplasm innovation. Here, we report the high-quality genome assemblies of a male Pamir argali (O ammon polii, 2n = 56), a female Tibetan sheep (O aries, 2n = 54), and a male hybrid of Pamir argali and domestic sheep, and the high-throughput sequencing of 425 ovine animals, including the hybrids of argali and domestic sheep. We detected genomic synteny between Chromosome 2 of sheep and two acrocentric chromosomes of argali. We revealed consistent satellite repeats around the chromosome breakpoints, which could have resulted in chromosome fusion. We observed many more hybrids with karyotype 2n = 54 than with 2n = 55, which could be explained by the selfish centromeres, the possible decreased rate of normal/balanced sperm, and the increased incidence of early pregnancy loss in the aneuploid ewes or rams. We identified genes and variants associated with important morphological and production traits (e.g., body weight, cannon circumference, hip height, and tail length) that show significant variations. We revealed a strong selective signature at the mutation (c.334C > A, p.G112W) in TBXT and confirmed its association with tail length among sheep populations of wide geographic and genetic origins. We produced an intercross population of 110 F2 offspring with varied number of vertebrae and validated the causal mutation by whole-genome association analysis. We verified its function using CRISPR-Cas9 genome editing. Our results provide insights into chromosomal speciation and phenotypic evolution and a foundation of genetic variants for the breeding of sheep and other animals.