Heat-killed cell preparation of Corynebacterium glutamicum stimulates the immune activity and improves survival of mice against enterohemorrhagic Escherichia coli

Fermentation by Corynebacterium glutamicum is used by various industries to produce L-Glutamate, and the heat-killed cell preparation of this bacterium (HCCG) is a by-product of the fermentation process. In present study, we evaluated the immunostimulating and survival effects against enterohemorrhagic Escherichia coli (STEC) infection of HCCG. HCCG significantly stimulated in vitro IgA and interleukin-12 p70 production in murine Peyer’s patch cells and peritoneal macrophages, respectively. Oral administration of 10 mg/kg body weight (BW) of HCCG for seven consecutive days stimulated IgA concentration in murine cecal digesta. Mice were orally administered HCCG for 17 consecutive days (d0–d17), and challenged with STEC on d4 to d6. Survival of mice tended to improve by 100 mg/kg BW of HCCG administration compared with those in control group. In conclusion, HCCG supplementation was found to prevent STEC infection in mice, and thus it may have the potential to stimulate the immune status of mammals.     

Heat-killed Lactobacillus helveticus strain MCC1848 confers resilience to anxiety or depression-like symptoms caused by subchronic social defeat stress in mice

The gut microbiota is involved in the pathogenesis of stress-related disorders. Probiotics can benefit the central nervous system via the microbiota–gut–brain axis, which raises the possibility that probiotics are effective in managing depression. In the present study, we examined the effects of heat-killed Lactobacillus helveticus strain MCC1848 in subchronic and mild social defeat stress (sCSDS) model mice (a widely used animal model of depression). MCC1848 supplementation significantly enhanced the interaction time in the social interaction test and sucrose preference ratio in the sucrose preference test, suggesting that MCC1848 improved anxiety- or depressive-like behaviors in sCSDS mice. The gene expression profile analysis of the nucleus accumbens, which plays an important role in stress resilience, indicated that MCC1848 ameliorated sCSDS-induced gene expression alterations in signal transduction or nervous system development. These findings suggest that MCC1848 supplementation is useful as a preventive strategy for chronic-stress-induced depression.     

Effects of Moritella viscosa antigens on pro-inflammatory gene expression in an Atlantic salmon (Salmo salar Linnaeus) cell line (SHK-1)

Moritella viscosa is the causative agent of winter ulcer disease in salmonids reared in North-Atlantic countries. In this study the effects of selected M. viscosa antigens on cytotoxicity and pro-inflammatory gene expression in an Atlantic salmon (Salmo salar Linnaeus) macrophage-like cell line (SHK-1) were examined. SHK-1 cells were stimulated with live and heat-killed bacterial cells, extracellular products (ECP) and an extracellular vibriolysin, termed MvP1. Following incubation, cytotoxicity and expression levels of interleukin-1β (IL-1β) and interleukin-8 (IL-8) were examined at different time points. Both live M. viscosa cells and ECP were cytotoxic, but neither heat-killed cells, nor the MvP1 peptidase caused cell death. Expression levels of both IL-1β and IL-8 increased significantly after stimulation with live cells, but heat-killed cells only caused increased IL-8 expression. ECP did not affect IL-1β expression, but did stimulate IL-8 expression. The isolated MvP1 peptidase stimulated both IL-1β and IL-8 expression at the highest concentration tested. This study reveals a difference in the induction of pro-inflammatory gene expression in salmon SHK-1 cells between live and heat-killed M. viscosa cells, and also that an unknown secreted factor is the main stimulant of IL-β and IL-8 expression.     

The Role of Intestinal Microbiota in Development of Irinotecan Toxicity and in Toxicity Reduction through Dietary Fibres in Rats

CPT-11 is a drug used as chemotherapy for colorectal cancer. CPT-11 causes toxic side-effects in patients. CPT-11 toxicity has been attributed to the activity of intestinal microbiota, however, intestinal microbiota may also have protective effects in CP!-11 chemotherapy. This study aimed to elucidate mechanisms through which microbiota and dietary fibres could modify host health. Rats bearing a Ward colon carcinoma were treated with a two-cycle CPT-11/5-fluorouracil therapy recapitulating clinical therapy of colorectal cancer. Animals were fed with a semi-purified diet or a semi-purified diet was supplemented with non-digestible carbohydrates (isomalto-oligosaccharides, resistant starch, fructo-oligosaccharides, or inulin) in 3 independent experiments. Changes in intestinal microbiota, bacteria translocating to mesenteric lymphnodes, cecal GUD activity, and cecal SCFA production, and the intestinal concentration of CPT-11 and its metabolites were analysed. Non-digestible carbohydrates significantly influenced feed intake, body weight and other indicators of animal health. The identification of translocating bacteria and their quantification in cecal microbiota indicated that overgrowth of the intestine by opportunistic pathogens was not a major contributor to CPT-11 toxicity. Remarkably, fecal GUD activity positively correlated to body weight and feed intake but negatively correlated to cecal SN-38 concentrations and IL1-β. The reduction in CPT-11 toxicity by non-digestible carbohydrates did not correlate to stimulation of specific bacterial taxa. However, cecal butyrate concentrations and feed intake were highly correlated. The protective role of intestinal butyrate production was substantiated by a positive correlation of the host expression of MCT1 (monocarboxylate transporter 1) with body weight as well as a positive correlation of the abundance of bacterial butyryl-CoA gene with cecal butyrate concentrations. These correlations support the interpretation that the influence of dietary fibre on CPT-11 toxicity is partially mediated by an increased cecal production of butyrate.     

The significance of the diversity and composition of the cecal microbiota of the Tibetan swine

The Tibetan swine (TIS) is a non-ruminant herbivore with high disease resistance. Also, it has the ability to digest plants with high fiber content. However, it is not known whether any relationship exist between these characteristics of the TIS and its cecal microbiota. Thus, this study aims to investigate the cecal microbiota of the adult TIS using high-throughput sequencing techniques in order to explore possible relationships between these unique characteristics of the TIS (high disease resistance and ability to digest high fiber plants) and its cecal microbiota. PIC pigs (lean type) were chosen as controls. The results show that 75,069 valid sequences of the 16S rRNA gene at V4-V5 region were obtained in the cecal content of TIS. They were composed of 15 phyla, 70 genera and divided into 660 Operational Taxonomic Units (OTUs). Bacteroidetes and Firmicutes were the predominant phyla in both breeds, but TIS had more Bacteroidetes than Firmicutes. Also, 42.4% of the cecal bacteria were found to be unclassified and uncultured. Many cellulolytic bacteria were also found in the two breeds. TIS (88.10%) had much higher abundance in the core bacterial communities than PIC pigs (81.29%), and the proportion of Bacteroides and Spirochaetes that can effectively degrade cellulose were 6.01 and 6.40% higher than PIC pigs, respectively, while Proteobacteria that are closely related to gastrointestinal diseases were 1.61% lower than PIC pigs. Thus, the disease resistance of the TIS and its ability to digest plants with high fiber content may be related to high abundance of core bacterial communities as well as the large number of unknown and unclassified bacteria.     

Study of oral microbiota diversity among groups of families originally from different countries

The diversity of oral microbiota is affected by diets habits, gender, age, ethnic group, and environment. The acquisition of oral microbiota and the role of family on oral microbiota development is poorly understood. This study aims to characterize and compare the oral bacterial microbiota among families using 16S rRNA gene sequencing. This work was conducted in Jeddah city from 2020 to 2021, in which four families composed of 20 members of different ethnicity and lifestyle were recruited. After the collection of saliva samples, the DNA was extracted and processed for 16S rRNA gene metagenomics sequencing. Among 378 OUTs generated, 39 (10.3%) were unique in group A, 13 (3.4%) unique in group B, and 11 (2.9%) were unique in groups C and D. We observed a significant variation at the level of top abundance phylum (14), families (23), genera (24), and species (22) of bacteria among family members. Within family groups, different bacterial species were reported to be more dominant among certain family members than the other; Prevotella melaninogenica, Prevotella histicola and Haemophilus parainfluenzae, Veillonella atypica, Porphyromonas pasteri and Haemophilus pittmaniae were more dominant in parents of some families than the other family member. In summary, this study highlights the precise and perceptible association of oral microbial between family members. Our findings documented the clustering of certain bacterial species in family groups, supporting the role of community in the development of oral microbiota.     

Putative Microbial Population Shifts Attributable to Nasal Administration of Streptococcus salivarius 24SMBc and Streptococcus oralis 89a

Changes in bacterial composition of nasal microbiota may alter the host’s susceptibility to several infectious and allergic diseases such as chronic rhinosinusitis and allergic rhinitis. The aim of this study was to evaluate the effects of 1-week administration of a probiotic product, composed by a combination of Streptococcus salivarius 24SMBc and Streptococcus oralis 89a, on the nostril microbiota. Differences in the nasal microbiota composition were investigated by using a next-generation sequencing approach. A strong and significant decrease in Staphylococcus aureus abundance was detected immediately after the bacterial administration. Moreover, comparing the microbial networks of nostril microbiota before and 1 month after the end of treatment, we detected an increase in the total number of both bacterial nodes and microbial correlations, with particular regard to the beneficial ones. Furthermore, a less abundance of microbial genera commonly associated to potential harmful bacteria has been observed. These results suggest a potential ability of S. salivarius 24SMBc and S. oralis 89a to regulate and reorganize the nasal microbiota composition, possibly favoring those microorganisms that may be able to limit the overgrowth of potential pathogens.

     

High-throughput sequence-based analysis of the intestinal microbiota of weanling pigs fed genetically modified MON810 maize expressing Bacillus thuringiensis Cry1Ab (Bt maize) for 31 days

The objective of this study was to investigate if feeding genetically modified (GM) MON810 maize expressing the Bacillus thuringiensis insecticidal protein (Bt maize) had any effects on the porcine intestinal microbiota. Eighteen pigs were weaned at ~28 days and, following a 6-day acclimatization period, were assigned to diets containing either GM (Bt MON810) maize or non-GM isogenic parent line maize for 31 days (n = 9/treatment). Effects on the porcine intestinal microbiota were assessed through culture-dependent and -independent approaches. Fecal, cecal, and ileal counts of total anaerobes, Enterobacteriaceae, and Lactobacillus were not significantly different between pigs fed the isogenic or Bt maize-based diets. Furthermore, high-throughput 16S rRNA gene sequencing revealed few differences in the compositions of the cecal microbiotas. The only differences were that pigs fed the Bt maize diet had higher cecal abundance of Enterococcaceae (0.06 versus 0%; P < 0.05), Erysipelotrichaceae (1.28 versus 1.17%; P < 0.05), and Bifidobacterium (0.04 versus 0%; P < 0.05) and lower abundance of Blautia (0.23 versus 0.40%; P < 0.05) than pigs fed the isogenic maize diet. A lower enzyme-resistant starch content in the Bt maize, which is most likely a result of normal variation and not due to the genetic modification, may account for some of the differences observed within the cecal microbiotas. These results indicate that Bt maize is well tolerated by the porcine intestinal microbiota and provide additional data for safety assessment of Bt maize. Furthermore, these data can potentially be extrapolated to humans, considering the suitability of pigs as a human model.     

Infant gut microbiota is protective against cow's milk allergy in mice despite immature ileal T-cell response

Faecal microbiota of healthy infant displays a large abundance of Bifidobacterium spp. and Bacteroides spp. Although some studies have reported an association between these two genera and allergy, these findings remain a subject of debate. Using a gnotobiotic mouse model of cow's milk allergy, we investigated the impact of an infant gut microbiota – mainly composed of Bifidobacterium and Bacteroides spp. – on immune activation and allergic manifestations. The transplanted microbiota failed to restore an ileal T-cell response similar to the one observed in conventional mice. This may be due to the low bacterial translocation into Peyer's patches in gnotobiotic mice. The allergic response was then monitored in germ-free, gnotobiotic, and conventional mice after repeated oral sensitization with whey proteins and cholera toxin. Colonized mice displayed a lower drop of rectal temperature upon oral challenge with b-lactoglobulin, lower plasma mMCP-1, and lower anti-BLG IgG1 than germ-free mice. The foxp3 gene was highly expressed in the ileum of both colonized mice that were protected against allergy. This study is the first demonstration that a transplanted healthy infant microbiota mainly composed of Bifidobacterium and Bacteroides had a protective impact on sensitization and food allergy in mice despite altered T-cell response in the ileum.     

Effects of Dietary Inclusion of β-Hydroxy-β-Methylbutyrate on Growth Performance,Fat Deposition,Bile Acid Metabolism,and Gut Microbiota Function in High-Fat and High-Cholesterol Diet-Challenged Layer Chickens

Excessive lipid deposition in layer chickens due to inappropriate feeding adversely affects egg production; however, nutritional manipulation methods to deal with this issue are still limited. β-hydroxy-β-methylbutyrate (HMB), a metabolite of L-leucine, was recently reported as a lipid-lowering nutrient in mice and pigs, although its role in layers had not been investigated. Here, we employed high-fat and high-cholesterol diet (HFHCD)−challenged growing layers as an obese model to explore HMB function in the regulation of lipid metabolism and the potential mechanisms involved. We found that dietary supplementation with (0.05% or 0.10%) HMB significantly reduced HFHCD−induced bodyweight growth in layers, mainly due to reduction in abdominal fat deposition. Mechanistically, HMB supplementation enhanced hepatic bile acid synthesis from cholesterol through elevating expression of Cyp7a1, a gene coding a key enzyme in bile acid synthesis. Furthermore, 16S rRNA gene sequencing revealed that HMB supplementation remodeled the diversity and composition of the layers’ cecal microbiota, and the abundance of Bacteroidetes at the phylum level were especially affected. Correlation analysis further indicated a strong negative association between Bacteroidetes abundance and lipid metabolism−related parameters. Taken together, these data suggest that dietary HMB supplementation could improve abdominal fat deposition in layers, probably through modulating hepatic bile acid synthesis and gut microbiota function.     

Effects of two probiotic Lactobacillus strains on jejunal and cecal microbiota of broiler chicken under acute heat stress condition as revealed by molecular analysis of 16S rRNA genes

We examined the effects of probiotic Lactobacillus strains of Lactobacillus agilis JCM 1048 and Lactobacillus salivarius subsp. salicinius JCM 1230 on jejunal and cecal microbiota of broiler chicken under heat stress condition using terminal restriction fragment length polymorphism (T-RFLP) analysis. The jejunal bacterial community was limited to a few bacterial groups, mostly Lactobacillus spp. A relatively abundant and higher prevalence of Lactobacillus spp. were observed in the jejunal and cecal microbiota of the probiotic chickens compared with those of the control chickens under heat stress condition. In general, the probiotic strains did not significantly affect the abundance of L. agilis and L. salivarius in chicken intestine but clearly contributed to increasing their prevalence in the probiotic chickens. The probiotic Lactobacillus strains enriched the diversity of Lactobacillus flora in chicken jejunum and cecum by increasing the abundance and prevalence of Lactobacillus spp. inhabiting the intestine. The richness of Lactobacillus species tended to be similar among the jejunal and cecal microbiota. The bacterial community of cecum was complex and age-dependent. The major components of the cecal microbiota were clostridia and lactobacilli. The Clostridium subcluster XIVa was the most predominant group in chicken cecum. Probiotic Lactobacillus strains restored the microbial balance and maintained the natural stability of indigenous bacterial microbiota following heat stress-induced changes.     

Abundant and diverse fungal microbiota in the murine intestine

Enteric microbiota play a variety of roles in intestinal health and disease. While bacteria in the intestine have been broadly characterized, little is known about the abundance or diversity of enteric fungi. This study utilized a culture-independent method termed oligonucleotide fingerprinting of rRNA genes (OFRG) to describe the compositions of fungal and bacterial rRNA genes from small and large intestines (tissue and luminal contents) of restricted-flora and specific-pathogen-free mice. OFRG analysis identified rRNA genes from all four major fungal phyla: Ascomycota, Basidiomycota, Chytridiomycota, and Zygomycota. The largest assemblages of fungal rRNA sequences were related to the genera Acremonium, Monilinia, Fusarium, Cryptococcus/Filobasidium, Scleroderma, Catenomyces, Spizellomyces, Neocallimastix, Powellomyces, Entophlyctis, Mortierella, and Smittium and the order Mucorales. The majority of bacterial rRNA gene clones were affiliated with the taxa Bacteroidetes, Firmicutes, Acinetobacter, and Lactobacillus. Sequence-selective PCR analyses also detected several of these bacterial and fungal rRNA genes in the mouse chow. Fluorescence in situ hybridization analysis with a fungal small-subunit rRNA probe revealed morphologically diverse microorganisms resident in the mucus biofilm adjacent to the cecal and proximal colonic epithelium. Hybridizing organisms comprised about 2% of the DAPI (4',6-diamidino-2-phenylindole, dihydrochloride)-positive organisms in the mucus biofilm, but their abundance in fecal material may be much lower. These data indicate that diverse fungal taxa are present in the intestinal microbial community. Their abundance suggests that they may play significant roles in enteric microbial functions.     

Comparative study on gut microbiota in three Anura frogs from a mountain stream

Composition and diversity in gut microbiota are impacted by a wide variety of factors. The similarity of gut microbiota in related or sympatric species has been gaining recent traction. Here, 16S rRNA gene sequencing technology was employed to study the gut microbiota of three sympatric frog species, namely Odorrana tormota, O. graminea, and Amolops wuyiensis. In these three frog species, the most abundant phylum was Proteobacteria, followed by Bacteroidetes, Verrucomicrobia, and Firmicutes. The most abundant family was Burkholderiaceae in three species. The most dominant genera were Burkholderia, Caballeronia, and Paraburkholderia with the highest relative abundance in O. tormota, O. graminea, and A. wuyiensis, respectively. No differences were observed in alpha diversity indexes among the three frog species. However, bacterial similarity of gut microbiota was significantly different between O. tormota and A. wuyiensis and between O. graminea and A. wuyiensis. Metabolism‐related gene function was predominantly enriched in the gut microbiota of the three evaluated frog species. From these findings, that the relative abundance of the gut microbiota and predicted gene functions differed in three species, we conclude that there were significant differences in the gut microbiota of the three species. Similar alpha diversity and interspecific bacterial similarity in the gut might be related to bacterial transmission among the three Anura frogs evaluated in this study.     

Correlation between protection against sepsis by probiotic therapy and stimulation of a novel bacterial phylotype

Prophylactic probiotic therapy has shown beneficial effects in an experimental rat model for acute pancreatitis on the health status of the animals. Mechanisms by which probiotic therapy interferes with severity of acute pancreatitis and associated sepsis, however, are poorly understood. The aims of this study were to identify the probiotic-induced changes in the gut microbiota and to correlate these changes to disease outcome. Duodenum and ileum samples were obtained from healthy and diseased rats subjected to pancreatitis for 7 days and prophylactically treated with either a multispecies probiotic mixture or a placebo. Intestinal microbiota was characterized by terminal-restriction fragment length polymorphism (T-RFLP) analyses of PCR-amplified 16S rRNA gene fragments. These analyses showed that during acute pancreatitis the host-specific ileal microbiota was replaced by an "acute pancreatitis-associated microbiota." This replacement was not reversed by administration of the probiotic mixture. An increase, however, was observed in the relative abundance of a novel bacterial phylotype most closely related to Clostridium lituseburense and referred to as commensal rat ileum bacterium (CRIB). Specific primers targeting the CRIB 16S rRNA gene sequence were developed to detect this phylotype by quantitative PCR. An ileal abundance of CRIB 16S rRNA genes of more than 7.5% of the total bacterial 16S rRNA gene pool was correlated with reduced duodenal bacterial overgrowth, reduced bacterial translocation to remote organs, improved pancreas pathology, and reduced proinflammatory cytokine levels in plasma. Our current findings and future studies involving this uncharacterized bacterial phylotype will contribute to unraveling one of the potential mechanisms of probiotic therapy.     

Synthetic microbial consortia for the treatment of Clostridioides difficile infection in mice model

Clostridioides difficile infection (CDI) as of recent has become a great concern to the impact on human health due to its high hazardous risk and rate of recurrence. Live bacterial therapeutics is a promising method to treat or prevent CDI. Here, a synthetic microbial consortia (SMC) B10 was constructed using probiotic strains with antibacterial and anti-quorum sensing activities, and the therapeutic effect of SMC B10 against C. difficile infection was evaluated in vitro. Compared to the model group, the treatment of SMC B10 significantly increased the survival rate. The clinical signs of mice were significantly ameliorated, especially the cecum injury, while the secretion of pro-inflammatory associated cytokines such as IL-1α, IL-6, IL-17A and TNF-α was reduced, the expression of TLR4 was inhibited, which alleviated the inflammatory response, and the expression of the tight junction protein Claudin-1 was increased, ultimately promoting the recovery of host health. The treatment of B10 restored gut microbiota dysbiosis and led to a healthy intestinal microbiota structure, significantly improved alpha diversity, suppressing potentially harmful bacteria and restoring other core bacterial species. In conclusion, SMC B10 can effectively treat CDI through modulate gut microbiota and attenuate the inflammatory response.     

Polydatin alleviates bleomycin-induced pulmonary fibrosis and alters the gut microbiota in a mouse model

To investigate the effect and mechanism of polydatin on bleomycin (BLM)-induced pulmonary fibrosis in a mouse model. The lung fibrosis model was induced by BLM. The contents of TNF-α, LPS, IL-6 and IL-1β in lung tissue, intestine and serum were detected by ELISA. Gut microbiota diversity was detected by 16S rDNA sequencing; R language was used to analyse species composition, α-diversity, β-diversity, species differences and marker species. Mice were fed drinking water mixed with four antibiotics (ampicillin, neomycin, metronidazole, vancomycin; antibiotics, ABx) to build a mouse model of ABx-induced bacterial depletion; and faecal microbiota from different groups were transplanted into BLM-treated or untreated ABx mice. The histopathological changes and collagen I and α-SMA expression were determined. Polydatin effectively reduced the degree of fibrosis in a BLM-induced pulmonary fibrosis mouse model; BLM and/or polydatin affected the abundance of the dominant gut microbiota in mice. Moreover, faecal microbiota transplantation (FMT) from polydatin-treated BLM mice effectively alleviated lung fibrosis in BLM-treated ABx mice compared with FMT from BLM mice. Polydatin can reduce fibrosis and inflammation in a BLM-induced mouse pulmonary fibrosis model. The alteration of gut microbiota by polydatin may be involved in the therapeutic effect.     

Bifidobacteria Abundance-Featured Gut Microbiota Compositional Change in Patients with Behcet’s Disease

Gut microbiota compositional alteration may have an association with immune dysfunction in patients with Behcet’s disease (BD). We conducted a fecal metagenomic analysis of BD patients. We analyzed fecal microbiota obtained from 12 patients with BD and 12 normal individuals by sequencing of 16S ribosomal RNA gene. We compared the relative abundance of bacterial taxa. Direct comparison of the relative abundance of bacterial taxa demonstrated that the genera Bifidobacterium and Eggerthella increased significantly and the genera Megamonas and Prevotella decreased significantly in BD patients compared with normal individuals. A linear discriminant analysis of bacterial taxa showed that the phylum Actinobacteria, including Bifidobacterium, and the family Lactobacillaceae exhibited larger positive effect sizes than other bacteria in patients with BD. The phylum Firmicutes and the class Clostridia had large effect sizes in normal individuals. There was no significant difference in annotated species numbers (as numbers of operational taxonomic unit; OTU) and bacterial diversity of each sample (alpha diversity) between BD patients and normal individuals. We next assigned each sample to a position using three axes by principal coordinates analysis of the OTU table. The two groups had a significant distance as beta diversity in the 3-axis space. Fecal sIgA concentrations increased significantly in BD patients but did not correlate with any bacterial taxonomic abundance. These data suggest that the compositional changes of gut microbes may be one type of dysbiosis (unfavorable microbiota alteration) in patients with BD. The dysbiosis may have an association with the pathophysiology of BD.