Integrated OMICs approach reveals energy metabolism pathway is vital for Salmonella Pullorum survival within the egg white.

Eggs, an important part of a healthy daily diet, can protect chicken embryo development due to the shell barrier and various antibacterial components within the egg white. Our previous study demonstrated that Salmonella Pullorum, highly adapted to chickens, can survive in the egg white and, therefore, be passed to newly hatched chicks. However, the survival strategy of Salmonella Pullorum in antibacterial conditions remains unknown. The overall transcripts in the egg white showed a large-scale shift compared to LB broth. The expression of common response genes and pathways, such as those involved in iron uptake, biotin biosynthesis, and virulence, was significantly changed, consistent with the other transovarial transmission serovar Enteritidis. Notably, membrane stress response, amino acid metabolism, and carbohydrate metabolism were specifically affected. Additional upregulated functionally relevant genes (JI728_13095, JI728_13100, JI728_17960, JI728_10085, JI728_15605, and nhaA) as mutants confirmed the susceptible phenotype. Furthermore, fim deletion resulted in an increased survival capacity in the egg white, consistent with the downregulated expression. The second-round RNA-Seq analysis of the Δfim mutant in the egg white revealed significantly upregulated genes compared with the wild type in the egg white responsible for energy metabolism located on the hyc and hyp operons regulated by FhlA, indicating the Δfim mutant cannot receive enough oxygen and switched to fermentative growth due to its inability to attach to the albumen surface. Together, this study provides a first estimate of the global transcriptional response of Salmonella Pullorum under antibacterial egg white and highlights the new potential role of fim deletion in optimizing energy metabolism pathways that may assist vertical transmission. IMPORTANCE: Pullorum disease, causing serious embryo death and chick mortality, results in substantial economic losses worldwide due to transovarial transmission. Egg-borne outbreaks are frequently reported in many countries. The present study has filled the knowledge gap regarding how the specific chicken-adapted pathogen Salmonella Pullorum behaves within the challenging environment of egg white. The deletion of the fim fimbrial system can increase survival in the albumen, possibly by reprogramming metabolism-related gene products, which reveals a new adaptive strategy of pathogens. Moreover, the comparison, including previous research on Salmonella Enteritidis, capable of vertical transmission, aims to provide diversified data sets in the field and further help to implement reasonable and effective measures to improve both food safety and animal health.

Salmonellosis outbreak archive in China: data collection and assembly.

Infectious disease outbreaks transcend the medical and public health realms, triggering widespread panic and impeding socio-economic development. Considering that self-limiting diarrhoea of sporadic cases is usually underreported, the Salmonella outbreak (SO) study offers a unique opportunity for source tracing, spatiotemporal correlation, and outbreak prediction. To summarize the pattern of SO and estimate observational epidemiological indicators, 1,134 qualitative reports screened from 1949 to 2023 were included in the systematic review dataset, which contained a 506-study meta-analysis dataset. In addition to the dataset comprising over 50 columns with a total of 46,494 entries eligible for inclusion in systematic reviews or input into prediction models, we also provide initial literature collection datasets and datasets containing socio-economic and climate information for relevant regions. This study has a broad impact on advancing knowledge regarding epidemic trends and prevention priorities in diverse salmonellosis outbreaks and guiding rational policy-making or predictive modeling to mitigate the infringement upon the right to life imposed by significant epidemics.

Microbiological hazards in infant and toddler food in China: A comprehensive study between 2004 and 2022.

Infant and toddler food (ITF), including powdered infant and follow-up formula (PIFF) and complementary food (CF), provides the majority of early-life nutrients for young children. As infants and toddlers are more vulnerable to foodborne diseases, the safety concern of ITF is the ultimate priority. However, nationwide surveillance for the presence of hazards, specifically microbiological hazards, in the Chinese ITF is partially known, posing a significant knowledge gap for risk ranking. Most importantly, the related regional surveys were largely published in Chinese, making the data unavailable for global sharing. To bridge these gaps, we screened 5,306 publications and conducted a comprehensive meta-analysis for microbiological hazards using 129 qualified studies. The four most reported microbiological hazards in ITF were Bacillus cereus (13.4 %), Cronobacter (4.8 %), Staphylococcus aureus (1.3 %), and Salmonella (1.1 %). B. cereus is a risk factor in ITF, specifically in PIFF, cereals, and ready-to-eat food. The prevalence of B. cereus was high in Northern and Southern China, while the prevalence of Cronobacter was high in Central China. Cronobacter is a microbiological hazard, specifically in PIFF, with a prevalence of 3.0 %. Interestingly, the prevalence dynamics of Cronobacter and B. cereus in ITF were rising and stable, respectively, whereas the prevalence of S. aureus and Salmonella decreased over time. Together, our analysis will promote the global sharing of these critical findings and may guide future policy making.

Compound Bacillus alleviates diarrhea by regulating gut microbes, metabolites, and inflammatory responses in pet cats.

BACKGROUND: Pet cats frequently have diarrhea in their daily life. Bacillus has a protective role that has crucial beneficial functions on intestinal homeostasis. The aim of this research was to investigate the effects of the compound Bacillus on the prevention of diarrhea, microbiota and metabolism in pet cats. A total of 20 pet cats (1-2 years old, 3.91 ± 0.92 kg) were randomly divided into two groups and fed with a basal diet (Control group), or a basal diet supplemented with 3 × 109 CFU/kg compound Bacillus (Probiotics group). The experiment lasted 33 days. RESULTS: Results showed that the compound Bacillus significantly reduced the rate of soft stools and diarrhea in pet cats compared with the control group (P < 0.05, n = 10). Meanwhile, compared with the control group, the probiotics group significantly decreased the content of IL-1ß and IL-6 and significantly increased IL-10 (P < 0.05, n = 6) in the serum. In addition, feeding probiotics significantly increased the abundance of p_Patescibacter and g_Plectosphaerella, decreased the abundance of p_Firmicutes, p_Gemmatimonadetes, g_Ruminococcaceae_UCG-005, g_Ascochytahe and g_Saccharomyces in the feces of the pet cats (P < 0.05, n = 6). And it also can significantly increase the content of total SCFAs, acetic acid and butyric acid in the feces (P < 0.05, n = 6). The fecal and serum metabolomics analyses revealed that most fecal and serum compounds were involved in metabolism, particularly in chemical structure transformation maps and amino acid metabolism. Also, eugenitol and methyl sulfate were the most significantly increased serum metabolites, and log2FC were 38.73 and 37.12, respectively. Pearson's correlation analysis showed that changes in serum metabolism and fecal microbiota were closely related to immune factors. There was also a strong correlation between serum metabolites and microbiota composition. CONCLUSIONS: The results of this research highlight the potential of the compound Bacillus as a dietary supplement to alleviate diarrhea in pet cats.

Lactiplantibacillus plantarum Lac16 Attenuates Enterohemorrhagic Escherichia coli O157:H7 Infection by Inhibiting Virulence Traits and Improving Intestinal Epithelial Barrier Function.

Large-scale use of antimicrobials in agriculture and medicine contributes to antibiotic residues in raw foods, the spread of antimicrobial resistance (AMR) and drug pollution, which seriously threatens human health and imposes significant economic burdens on society, suggesting the need for novel therapeutic options that prevent or control zoonoses. In this study, four probiotics were selected to assess their capability to alleviate pathogen-induced damage. Results showed that a simulated gastrointestinal juice and bile tolerated L. plantarum Lac16 with high lactic acid secretion can significantly inhibit the growth of multiple zoonotic pathogens. Lac16 also significantly inhibited the biofilm formation and mRNA expression of virulence traits (genes related to virulence, toxins, flagella biogenesis and motility, antibiotic resistance, biofilm formation and AI-2 quorum sensing) of enterohemorrhagic E. coli O157:H7 (EHEC). Furthermore, Lac16 and Lac26 significantly protected C. elegans against zoonotic pathogen-induced (EHEC, S. typhimurium, C. perfringens) deaths. Moreover, Lac16 significantly promoted epithelial repair and ameliorated lipopolysaccharide (LPS)-induced intestinal epithelial apoptosis and barrier dysfunction by activating the Wnt/ß-catenin signaling pathway, and markedly reduced LPS-induced inflammatory responses by inhibiting the TLR4/MyD88 signaling pathway. The present results indicate that Lac16 attenuates enterohemorrhagic E. coli infection-induced damage by inhibiting key virulence traits of E. coli, promoting epithelial repair and improving intestinal epithelial barrier function, which may be mediated by the activated Wnt/ß-catenin signaling pathway and the inhibited TLR4/MyD88 signaling pathway of the intestinal epithelium.

Bacillus amyloliquefaciens SC06 attenuated high-fat diet induced anxiety-like behavior and social withdrawal of male mice by improving antioxidant capacity, intestinal barrier function and modulating intestinal dysbiosis.

Anxiety-like behavior and social withdrawal induced by obesity and oxidative stress are significant health concerns in contemporary society. Our previously study found that Bacillus amyloliquefaciens SC06 (SC06) decreased the body weight of high-fat diet (HFD)-fed male mice and protected porcine intestinal epithelial cells against oxidative stress. The present study further investigated the effect of SC06 on HFD-induced obesity, anxiety-like behavior and social withdrawal of male mice and explored its mechanism. Results showed that SC06 significantly decreased HFD-induced obesity as evidenced by the decreased body weight, weight of liver and epididymal fat. Meanwhile, SC06 attenuated the anxiety-like behavior of HFD-fed male mice as illustrated by the more exploration time in both the open arms of elevated plus maze and the central area of open field and the reversed their social withdrawal tested in the three-chamber social choice task. SC06 also reduced reactive oxygen species (ROS) concentration and normalized the mitochondrial morphology in the hippocampus. SC06 reduced the systemic inflammation and increased the expression of intestinal tight junctions (ZO-1 and Claudin1). Furthermore, SC06 also altered the microbial diversity and composition, and decreased Firmicutes to Bacteroidetes ratio of HFD-fed male mice. These findings suggest SC06 attenuate HFD-induced anxiety-like behavior and social withdrawal of male mice by attenuating hippocampal oxidation stress, systemic inflammation, dysbiosis and improving intestinal barrier function.

Bacillus amyloliquefaciens SC06 in the diet improves egg quality of hens by altering intestinal microbiota and the effect is diminished by antimicrobial peptide.

This experiment investigated the effects of Bacillus amyloliquefaciens SC06 (BaSC06) and its combination with antimicrobial peptide (AMP) on the laying performance, egg quality, intestinal physical barrier, antioxidative status and immunity of commercial Jingbai strain laying hens. The results showed that BaSC06 significantly improved laying performance and egg quality of laying hens. However, there was a tendency to increase laying performance and decrease egg quality for the addition of AMP compared to the BaSC06 group. Also, both BaSC06 and its combination with AMP treatment increased length of microvilli and the content of tight junction protein in jejunum, and BaSC06 combination with AMP treatment is better than BaSC06 treatment alone. Compared to control, most of the serum antioxidant enzyme activities were significantly increased in the BaSC06+AMP group, the BaSC06 group only increased the activity of GSH-Px. Short-chain fatty acid analysis showed that BSC06 significantly increased the content of butyric, isobutyric and isovaleric acid in the cecum. However, the content of most of the short-chain fatty acids was even lower than that of the control group after the addition of AMP. Microbiota analysis showed that BaSC06 increased the absolute abundance of the butyrate-producing gut bacteria Ruminococaaoeae UCG-005, while the addition of AMP reduced the number of microorganisms detected and weakened the effect of BaSC06. BaSC06 acts as an anti-inflammatory agent by regulating the gut microbiota, and AMP further attenuates the immune response by reducing the number of gut microbes based on improved intestinal microbiota composition.

Dietary supplementation of Macleaya cordata extract and Bacillus in combination improve laying performance by regulating reproductive hormones, intestinal microbiota and barrier function of laying hens.

BACKGROUND: This study aimed to investigate whether the combination of Macleaya cordata extract (MCE) and Bacillus could improve the laying performance and health of laying hens better. METHODS: A total of 360 29-week-old Jingbai laying hens were randomly divided into 4 treatments: control group (basal diet), MCE group (basal diet + MCE), Probiotics Bacillus Compound (PBC) group (basal diet + compound Bacillus), MCE + PBC group (basal diet + MCE + compound Bacillus). The feeding experiment lasted for 42 d. RESULTS: The results showed that the laying rate and the average daily egg mass in the MCE + PBC group were significantly higher than those in the control group (P < 0.05) and better than the MCE and PBC group. Combination of MCE and Bacillus significantly increased the content of follicle-stimulating hormone (FSH) in the serum and up-regulated the expression of related hormone receptor gene (estrogen receptor-ß, FSHR and luteinizing hormone/choriogonadotropin receptor) in the ovary of laying hens (P < 0.05). In the MCE + PBC group, the mRNA expressions of zonula occluden-1, Occludin and mucin-2 in jejunum was increased and the intestinal epithelial barrier detected by transmission electron microscopy was enhanced compared with the control group (P < 0.05). In addition, compared with the control group, combination of MCE and Bacillus significantly increased the total antioxidant capacity and catalase activity (P < 0.05), and down-regulated the mRNA expressions of inflammation-related genes (interleukin-1ß and tumor necrosis factor-α) as well as apoptosis-related genes (Caspase 3, Caspase 8 and P53) (P < 0.05). The concentration of acetic acid and butyric acid in the cecum content of laying hens in the MCE + PBC group was significantly increased compared with the control group (P < 0.05). CONCLUSIONS: Collectively, dietary supplementation of 600 µg/kg MCE and 5 × 108 CFU/kg compound Bacillus can improve laying performance by improving microbiota to enhance antioxidant capacity and intestinal barrier, regulate reproductive hormones and the concentration of cecal short-chain fatty acids of laying hens, and the combined effect of MCE and Bacillus is better than that of single supplementation.

Modified Montmorillonite Improved Growth Performance of Broilers by Modulating Intestinal Microbiota and Enhancing Intestinal Barriers, Anti-Inflammatory Response, and Antioxidative Capacity.

This study aims to explore the effects of modified montmorillonite (MMT, copper loading) on the growth performance, gut microbiota, intestinal barrier, antioxidative capacity and immune function of broilers. Yellow-feathered broilers were randomly divided into control (CTR), modified montmorillonite (MMT), and antibiotic (ANTI) groups. Results revealed that MMT supplementation increased the BW and ADG and decreased the F/R during the 63-day experiment period. 16S rRNA sequencing showed that MMT modulated the cecal microbiota composition of broilers by increasing the relative abundance of two phyla (Firmicutes and Bacteroidetes) and two genera (Bacteroides and Faecalibacterium) and decreasing the abundance of genus Olsenella. MMT also improved the intestinal epithelial barrier indicated by the up-regulated mRNA expression of claudin-1, occludin, and ZO-1 and the increased length of microvilli in jejunum and the decreased levels of DAO and D-LA in serum. In addition, MMT enhanced the immune function indicated by the increased levels of immunoglobulins, the decreased levels of MPO and NO, the down-regulated mRNA expression of IL-1ß, IL-6, and TNF-α, and the up-regulated mRNA expression of IL-4 and IL-10. Moreover, MMT down-regulated the expression of jejunal TLRs/MAPK/NF-κB signaling pathway-related genes (TLR2, TLR4, Myd88, TRAF6, NF-κB, and iNOS) and related proteins (TRAF6, p38, ERK, NF-κB, and iNOS). In addition, MMT increased the antioxidant enzyme activities and the expression of Nrf2/HO-1 signaling pathway-related genes and thereby decreased the apoptosis-related genes expression. Spearman's correlation analysis revealed that Bacteroides, Faecalibacterium, and Olsenella were related to the inflammatory index (MPO and NO), oxidative stress (T-AOC, T-SOD, and CAT) and intestinal integrity (D-LA and DAO). Taken together, MMT supplementation improved the growth performance of broilers by modulating intestinal microbiota, enhancing the intestinal barrier function, and improving inflammatory response, which might be mediated by inhibiting the TLRs/MAPK/NF-κB signaling pathway, and antioxidative capacity mediated by the Nrf2/HO-1 signaling pathway.

Lactobacillus plantarum-derived postbiotics prevent Salmonella-induced neurological dysfunctions by modulating gut-brain axis in mice.

Postbiotics are the inactive bacteria and/or metabolites of beneficial microbes which have been recently found to be as effective as their live probiotic. This study aimed to evaluate the benefits of Lactobacillus plantarum (LP)-derived postbiotics on ameliorating Salmonella-induced neurological dysfunctions. Mice were pretreated with LP postbiotics (heat-killed bacteria or the metabolites) or active bacteria, and then challenged with Salmonella enterica Typhimurium (ST). Results showed that LP postbiotics, particularly the metabolites, effectively prevented ST infection in mice, as evidenced by the inhibited weight loss, bacterial translocation, and tissue damages. The LP postbiotics markedly suppressed brain injuries and neuroinflammation (the decreased interleukin (IL)-1ß and IL-6, and the increased IL-4 and IL-10). Behavior tests indicated that LP postbiotics, especially the metabolites, protected mice from ST-induced anxiety and depressive-like behaviors and cognitive impairment. A significant modulation of neuroactive molecules (5-hydroxytryptamine, gamma-aminobutyric acid, brain-derived neurotrophic factor, dopamine, acetylcholine, and neuropeptide Y) was also found by LP postbiotic pretreatment. Microbiome analysis revealed that LP postbiotics optimized the cecal microbial composition by increasing Helicobacter, Lactobacillus and Dubosiella, and decreasing Mucispirillum, norank_f_Oscillospiraceae, and Eubacterium_siraeum_group. Moreover, LP postbiotics inhibited the reduction of short-chain fatty acids caused by ST infection. Pearson's correlation assays further confirmed the strong relationship of LP postbiotics-mediated benefits and gut microbiota. This study highlights the effectiveness of postbiotics and provide a promising strategy for preventing infection-induced brain disorders by targeting gut-brain axis.

Bacillus amyloliquefaciens SC06 alleviates the obesity of ob/ob mice and improves their intestinal microbiota and bile acid metabolism.

Dietary interventions with probiotics have been widely reported to be effective in regulating obesity, and the intestinal microbiota is considered to be an important environmental factor. However, few reports focus on the interactions of microbiota-metabolites-phenotypic variables in ob/ob mice, and they have not been characterized in great detail. In this study, we investigated the effects of Bacillus amyloliquefaciens SC06 on obesity, the intestinal microbiota and the bile acid metabolism of ob/ob mice using biochemical testing, histochemical staining, high-throughput sequencing of the 16S rRNA gene, LC-MS/MS analysis and qRT-PCR. The results showed that SC06 ameliorated the fat mass percentage, hepatic steatosis and liver lipid metabolism disorders and reshaped the gut microbiota and metabolites in male ob/ob mice, specifically deceasing f_S24-7, p_TM7, s_Alistipes massiliensis, f_Rikenellaceae, f_Prevotellaceae, f_Lactobacillaceae, g_Alistipes, g_Flexispira, g_Lactobacillus, g_Odoribacter, g_AF12 and g_Prevotella and increasing f_Bacteroidaceae, g_Bacteroides and f_Desulfovibrionaceae. Meanwhile, SC06 treatment groups had lower ibuprofen and higher glycodeoxycholic acid and 7-dehydrocholesterol. Correlation analysis further clarified the relationships between compositional changes in the microbiota and alterations in the metabolites and phenotypes of ob/ob mice. Moreover, SC06 downregulated bile acid synthesis, export and re-absorption in the liver and increased ileum re-absorption into the blood in ob/ob mice, which may be mediated by the FXR-SHP/FGF15 signaling pathway. These results suggest that Bacillus amyloliquefaciens SC06 can ameliorate obesity in male ob/ob mice by reshaping the intestinal microbial composition, changing metabolites and regulating bile acid metabolism via the FXR signaling pathway.

Probiotic Bacillus Alleviates Oxidative Stress-Induced Liver Injury by Modulating Gut-Liver Axis in a Rat Model.

Emerging evidence suggests a key role of gut microbiota in maintaining liver functions through modulating the gut-liver axis. In this study, we investigated whether microbiota alteration mediated by probiotic Bacillus was involved in alleviating oxidative stress- induced liver injury. Sprague-Dawley rats were orally administered Bacillus SC06 or SC08 for a 24-day period and thereafter intraperitoneally injected diquat (DQ) to induce oxidative stress. Results showed that Bacillus, particularly SC06 significantly inhibited hepatic injuries, as evidenced by the alleviated damaged liver structure, the decreased levels of ALT, AST, ALP and LDH, and the suppressed mitochondrial dysfunction. SC06 pretreatment markedly enhanced the liver antioxidant capacity by decreasing MDA and p47, and increasing T-AOC, SOD and HO-1.16S rRNA sequencing analysis revealed that DQ significantly changed the diversities and composition of gut microbiota, whereas Bacillus pretreatments could attenuate gut dysbiosis. Pearson's correlation analysis showed that AST and MDA exerted a positive correlation with the opportunistic pathogenic genera and species (Escherichia and Shigella), and negatively correlated with the potential probiotics (Lactobacillus), while SOD exerted a reverse trend. The microbial metagenomic analysis demonstrated that Bacillus, particularly SC06 markedly suppress the metabolic pathways such as carbohydrate metabolism, lipid metabolism, amino acid metabolism and metabolism of cofactors and vitamins. Furthermore, SC06 decreased the gene abundance of the pathways mediating bacterial replication, secretion and pathogenicity. Taken together, Bacillus SC06 alleviates oxidative stress-induced liver injuries via optimizing the composition, metabolic pathways and pathogenic replication and secretion of gut microbiota. These findings elucidate the mechanisms of probiotics in alleviating oxidative stress and provide a promising strategy for preventing liver diseases by targeting gut microbiota.

Effects of glucose oxidase and its combination with B. amyloliquefaciens SC06 on intestinal microbiota, immune response and antioxidative capacity in broilers.

Glucose oxidase (GOD) is an aerobic dehydrogenase, which catalyses the oxidation of ß-D-glucose to gluconic acid and hydrogen peroxide. This study aimed to investigate the effects of dietary glucose oxidase and its combined effects with Bacillus amyloliquefaciens SC06 (BaSC06) on the intestinal microbiota, immune function and antioxidant capacity of broilers. One-day-old male Lingnan yellow-feathered broilers (n = 720) were randomly assigned to four treatment groups: Control group (basal diet), Anti group (basal diet supplemented with 200 mg/kg enramycin), GOD group (basal diet supplemented with 75 U/kg GOD), and combination of GOD and BaSC06 (GB) group (GOD diet (75 U/kg) supplemented with 1 × 108 colony-forming units BaSC06/kg feed), with six replicates per group and 30 birds per replicate. The experiment was conducted over 52 days. The results indicated a significant decrease in α-diversity (Observed species, Chao1, PD_whole_tree and Shannon) with GOD treatment, compared with the control group. GB treatment also significantly decreased the Shannon index of cecal microbiota. GOD treatment significantly decreased the α-diversity, whereas GB treatment significantly increased these indices except for the Chao1 index, compared with the Anti group. Compared with the control group, the relative abundance of Bacteroides in the GOD and GB groups was significantly increased, whereas a decrease in Firmicutes was observed. Compared with the Anti group, GOD treatment significantly increased the relative abundances of Bacteroides and Lactobacillales, while GB treatment significantly increased Lactobacillales and decreased Proteobacteria levels. In addition, GOD treatment significantly decreased interleukin-10 and interferon-γ levels, compared with the control group. In contrast, GB treatment significantly downregulated interferon-γ levels and upregulated secretory immunoglobulin A, transforming growth factor-ß and interleukin-2 expression in the jejunal mucosa. GOD treatment significantly decreased transforming growth factor-ß and interleukin-10 levels, whereas GB treatment markedly increased interferon-γ expression in the jejunal mucosa compared with the Anti group. Furthermore, GB treatment significantly increased the total antioxidant capability levels and the total superoxide dismutase (T-SOD) and catalase (CAT) activities compared with the control group. Meanwhile, GOD treatment significantly increased glutathione peroxidase (GSH-Px) activity in the jejunal mucosa. Total superoxide dismutase, GSH-Px and CAT activities in the Anti group were higher than in the GOD and GB groups. The malondialdehyde levels in the control group were the highest among all groups. In conclusion, our results indicated that supplementation with GOD alone and its combination with BaSC06 in diet could increase antioxidant capacity, immune function and improve the intestinal microbiota composition of broilers. Combination treatment with GOD with BaSC06 exerted stronger effects than GOD treatment only.

Protective Effects of Lactobacillus plantarum Lac16 on Clostridium perfringens Infection-Associated Injury in IPEC-J2 Cells.

Clostridium perfringens (C. perfringens) causes intestinal injury through overgrowth and the secretion of multiple toxins, leading to diarrhea and necrotic enteritis in animals, including pigs, chickens, and sheep. This study aimed to investigate the protective effects of Lactobacillus plantarum (L. plantarum) Lac16 on C. perfringens infection-associated injury in intestinal porcine epithelial cell line (IPEC-J2). The results showed that L. plantarum Lac16 significantly inhibited the growth of C. perfringens, which was accompanied by a decrease in pH levels. In addition, L. plantarum Lac16 significantly elevated the mRNA expression levels of host defense peptides (HDPs) in IPEC-J2 cells, decreased the adhesion of C. perfringens to IPEC-J2 cells, and attenuated C. perfringens-induced cellular cytotoxicity and intestinal barrier damage. Furthermore, L. plantarum Lac16 significantly suppressed C. perfringens-induced gene expressions of proinflammatory cytokines and pattern recognition receptors (PRRs) in IPEC-J2 cells. Moreover, L. plantarum Lac16 preincubation effectively inhibited the phosphorylation of p65 caused by C. perfringens infection. Collectively, probiotic L. plantarum Lac16 exerts protective effects against C. perfringens infection-associated injury in IPEC-J2 cells.

Dietary Supplementation With Lactobacillus plantarum Ameliorates Compromise of Growth Performance by Modulating Short-Chain Fatty Acids and Intestinal Dysbiosis in Broilers Under Clostridium perfringens Challenge.

Clostridium perfringens is an important zoonotic pathogen associated with food contamination and poisoning, gas gangrene, necrotizing enterocolitis or necrotic enteritis in humans and animals. Dysbacteriosis is supposedly associated with the development of C. perfringens infection induced necrotic enteritis, but the detailed relationship between intestinal health, microbiome, and C. perfringens infection-induced necrotic enteritis remains poorly understood. This research investigated the effect of probiotics on the growth performance and intestinal health of broilers, and the involved roles of intestinal microbiota and microbial metabolic functions under C. perfringens infection. Results showed that subclinical necrotic enteritis was successfully induced as evidenced by the significant lower body weight (BW), suppressed feed conversion ratio (FCR), decreased ileal villus height and mucosal barrier function, and increased ileal histopathological score and bursal weight index. Lactobacillus plantarum or Paenibacillus polymyxa significantly attenuated C. perfringens-induced compromise of growth performance (BW, FCR) and ileal mucosa damage as illustrated by the increased ileal villus height and villus/crypt ratio, the decreased ileal histopathological score and the enhanced ileal mucosal barrier function. L. plantarum also significantly alleviated C. perfringens-induced enlarged bursa of fabricius and the decreased levels of ileal total SCFAs, acetate, lactate, and butyrate. Furthermore, dietary L. plantarum improved C. perfringens infection-induced intestinal dysbiosis as evidenced by significantly enriched short-chain fatty acids-producing bacteria (Lachnospiraceae, Ruminococcaceae, Oscillospira, Faecalibacterium, Blautia), reduced drug-resistant bacteria (Bacteroides, Alistipes) and enteric pathogens (Escherichia coli, Bacteroides fragilis) and bacterial metabolic dysfunctions as illustrated by significantly increased bacterial fatty acid biosynthesis, decreased bacterial lipopolysaccharide biosynthesis, and antibiotic biosynthesis (streptomycin and vancomycin). Additionally, the BW and intestinal SCFAs were the principal factors affecting the bacterial communities and microbial metabolic functions. The above findings indicate that dietary with L. plantarum attenuates C. perfringens-induced compromise of growth performance and intestinal dysbiosis by increasing SCFAs and improving intestinal health in broilers.

Bacillus amyloliquefaciens SC06 Induced AKT-FOXO Signaling Pathway-Mediated Autophagy to Alleviate Oxidative Stress in IPEC-J2 Cells.

Autophagy is a conserved proteolytic mechanism, which degrades and recycles damaged organs and proteins in cells to resist external stress. Probiotics could induce autophagy; however, its underlying molecular mechanisms remain elusive. Our previous study has found that BaSC06 could alleviate oxidative stress by inducing autophagy in rats. This research aimed to verify whether Bacillus amyloliquefaciens SC06 can induce autophagy to alleviate oxidative stress in IPEC-J2 cells, as well as explore its mechanisms. IPEC-J2 cells were first pretreated with 108 CFU/mL BaSC06, and then were induced to oxidative stress by the optimal dose of diquat. The results showed that BaSC06 significantly triggered autophagy, indicated by the up-regulation of LC3 and Beclin1 along with downregulation of p62 in IPEC-J2 cells. Further analysis revealed that BaSC06 inhibited the AKT-FOXO signaling pathway by inhibiting the expression of p-AKT and p-FOXO and inducing the expression of SIRT1, resulting in increasing the transcriptional activity of FOXO3 and gene expression of the ATG5-ATG12 complex to induce autophagy, which alleviated oxidative stress and apoptosis. Taken together, BaSC06 can induce AKT-FOXO-mediated autophagy to alleviate oxidative stress-induced apoptosis and cell damage, thus providing novel theoretical support for probiotics in the prevention and treatment of oxidative damage.

Probiotic Paenibacillus polymyxa 10 and Lactobacillus plantarum 16 enhance growth performance of broilers by improving the intestinal health.

With the ever-growing strict prohibitions on antibiotic growth promoters (AGP) in animal production, in-feed probiotics are becoming attractive alternatives to antibiotics in the poultry industry. To investigate the effects of Paenibacillus polymyxa 10 and Lactobacillus plantarum 16 on the growth performance and intestinal health of broilers, 540 male Cobb 500 broilers of 1 d old were randomly divided into 3 groups with 6 replicates per group and 30 chicks per replicate. Broilers were fed with either a basal diet or basal diets supplemented with 1 × 108 colony-forming units (CFU)/kg P. polymyxa 10 (BSC10) or L. plantarum 16 (Lac16) for 42 d. Results showed that Lac16 treatment improved (P < 0.05) the growth performance (body weight and feed conversion) of broilers at the starter phase, while BSC10 treatment slightly improved (P > 0.05) the growth performance of the starter phase broilers. The increased villus height (P < 0.05) at d 14, 21 and 42 and villus height to crypt depth ratio (P < 0.05) at d 14 and 21 were observed in the ileum of the 2 probiotic groups. Besides, transmission electron microscopy results showed that the 2 probiotics enhanced the intestinal epithelial barrier. Both probiotic treatments up-regulated (P < 0.05) the mRNA expression of fatty acid binding protein 1 (FABP1) and sodium-dependent glucose transporters-1 (SGLT-1) in the ileal mucosa of broilers at d 21. In addition, BSC10 and Lac16 treatments significantly (P < 0.05) increased the relative abundance of short-chain fatty acids-producing bacteria, such as Butyricicoccus pullicaecorum, Faecalibacterium prausnitzii, Lachnospira and Coprococcu, and significantly (P < 0.05) decreased the relative abundance of enteric pathogens (Escherichia coli, Bacteroides fragilis and Shigella sonnei). Furthermore, the 2 probiotic treatments also increased the positive connection among the intestinal microbes and the carbohydrate metabolism-related pathways of the intestinal bacteria (P < 0.05), with decreasing (P < 0.05) nucleotides biosynthesis-related pathways of the intestinal bacteria. Overall, these results suggest that the 2 probiotics, especially Lac16, have a potential beneficial effect on the growth performance and intestinal health of starter phase broilers.

Glycyrrhizin Attenuates Salmonella Typhimurium-Induced Tissue Injury, Inflammatory Response, and Intestinal Dysbiosis in C57BL/6 Mice.

Salmonellae are one of the most important foodborne pathogens, which threaten the health of humans and animals severely. Glycyrrhizin (GL) has been proven to exhibit anti-inflammatory and tissue-protective properties. Here, we investigated the effects of GL on tissue injury, inflammatory response, and intestinal dysbiosis in Salmonella Typhimurium-infected mice. Results showed that GL or gentamicin (GM) significantly (P < 0.05) alleviated ST-induced splenomegaly indicated by the decreased spleen index, injury of liver and jejunum indicated by the decreased hepatocytic apoptosis, and the increased jejunal villous height. GL significantly (P < 0.05) increased secretion of inflammatory cytokines (IFN-γ, IL-12p70, IL-6, and IL-10) in spleen and IL-12p40 mRNA expression in liver. Meanwhile, GL or GM pre-infection treatments significantly (P < 0.05) decreased ST-induced pro-inflammatory cytokine (IFN-γ, TNF-α, and IL-6) expression in both spleen and liver and increased (P < 0.05) anti-inflammatory cytokine IL-10 secretion in spleen. Furthermore, GL or GM pre-infection treatment also regulates the diversities and compositions of intestinal microbiota and decreased the negative connection among the intestinal microbes in ST-infected mice. The above findings indicate that GL alleviates ST-induced splenomegaly, hepatocytic apoptosis, injury of jejunum and liver, inflammatory response of liver and spleen, and intestinal dysbacteriosis in mice.

Effects of Bacillus amyloliquefaciens Instead of Antibiotics on Growth Performance, Intestinal Health, and Intestinal Microbiota of Broilers.

The aim of this study was to evaluate the dietary effects of Bacillus amyloliquefaciens SC06 (SC06) instead of antibiotics on the growth performance, intestinal health, and intestinal microbiota of broilers. A total of 360 30-day-old Lingnan yellow broilers were randomly allocated into two groups with six replicates per group (30 birds per replicate). The broilers were fed either a non-supplemented diet or a diet supplemented with 108 colony-forming units lyophilized SC06 per kilogram feed for 30 days. Results showed that SC06 supplementation had no effect on the growth performance compared with that of the control group. SC06 treatment significantly (P <0.05) increased the total antioxidant capacity (T-AOC), total superoxide dismutase (T-SOD) activity in the liver, and the activities of trypsin, α-amylase (AMS), and Na+K+-ATPase in the ileum, whereas it decreased (P < 0.05) lipase, gamma glutamyl transpeptidase (γ-GT), and maltase activities in the ileum. Meanwhile, SC06 treatment also improved the immune function indicated by the significantly (P < 0.05) increased anti-inflammatory cytokine [interleukin (IL)-10] level and the decreased (P < 0.05) pro-inflammatory cytokine [IL-6 and tumor necrosis factor (TNF)-α] levels in the ileum. Furthermore, we also found that SC06 enhanced the intestinal epithelial intercellular integrity (tight junction and adhesion belt) in the ileum. Microbial analysis showed that SC06 mainly increased the alpha diversity indices in the jejunum, ileum, and cecum. SC06 treatment also significantly (P < 0.05) increased the abundances of Bacteroidetes, Bacteroidales, Bacteroides, Fusobacteria, Clostridiaceae, and Veillonellaceae in the cecum and simultaneously decreased the abundances of Planococcaceae in the duodenum, Microbacteriaceae in the jejunum, and Lachnospiraceae, [Ruminococcus] and Ruminococcus in cecum. In conclusion, these results suggested that B. amyloliquefaciens instead of antibiotics showed a potential beneficial effect on the intestinal health of broilers.

Saccharomyces boulardii attenuates inflammatory response induced by Clostridium perfringens via TLR4/TLR15-MyD8 pathway in HD11 avian macrophages.

Macrophages are professional phagocytic cells that play a critical role in initiating immune responses by presenting antigen and phagocytic clearance. The macrophages can be targeted for immunomodulation by beneficial microbes, such as probiotics. The aim of this study is to investigate the protective effect of Saccharomyces boulardii against Clostridium perfringens infection in avian macrophage cell line HD11. In this study, HD11 macrophages were prestimulated with S. boulardii for 6 h and then infected with C. perfringens for 3 h. Results showed that S. boulardii enhanced phagocytosis and bactericidal capacity against C. perfringens by HD11 cells. The S. boulardii effectively promoted the mRNA expression of CD80, CD83, and CD197 cell-surface molecules in C. perfringens-infected HD11 cells. Moreover, we found that prestimulation with S. boulardii reduced the mRNA expression of CD40, toll-like receptor [TLR] 4, and TLR15 induced by C. perfringens and thereby downregulated the mRNA expression of myeloid differentiation primary response 88, TNF receptor associated factor 6, nuclear factor kappa-B p65 subunit, and c-Jun N-terminal kinase genes in HD11 cells. The upregulation of cytokines (interleukin [IL]-6, tumor necrosis factor alpha, and IL-10) and inducible nitric oxide synthase mRNA expression in C. perfringens-infected HD11 cells were noticeably inhibited by S. boulardii pretreatment. Conclusively, these results might provide a new insight into the role of S. boulardii in regulating avian immune defense against C. perfringens invasion and immune escape.