16S rDNA Amplicon Sequencing Research Articles

359. INTRATUMORAL MICROBIOME IMPACTS IMMUNE INFILTRATES IN TUMOR MICROENVIRONMENT AND PREDICTS PROGNOSIS IN ESOPHAGEAL SQUAMOUS CELL CARCINOMA PATIENTS

Abstract Emerging evidence suggests different intratumoral microbiome exists in different tumors and plays a crucial function in carcinogenesis. However, whether intratumoral microbiome in esophageal squamous cell carcinoma (ESCC) impacts clinical outcomes and its mechanism remains unclear. 16S rDNA amplicon sequencing was performed on surgical resected samples from 106 ESCC patients including 25 paired tumorous and non-tumorous tissues (NTs) to analyze intratumoral microbiome abundance and its composition. Multiplex fluorescent immunohistochemistry (mfIHC) staining was used to profile the phenotypes of immune infiltrates in tumor microenvironment (TME). Both Shannon index and operational taxonomic units (OTUs), representing alpha-diversity of microbiome, were found significantly lower in tumors comparing to the corresponding NTs. Multivariate analysis revealed that the intratumoral Shannon index, the relative abundance of Lactobacillus and the pTNM stage were independently associated with patients’ overall survival. Furthermore, the relative abundance of Lactobacillus and Shannon index were both positively correlated with the proportions of PD-L1+ epithelial cells (ECs) and tumor-associated macrophages (TAMs, and Shannon index was negatively correlated with the proportions of natural killer cells (NKs) and activated cytotoxic T cells (aCTLs) in TME, indicating an immunosuppressive microenvironment. These findings underscored for the first time that the high abundance of intratumoral Lactobacillus and microbiome alpha-diversity were associated with poor long-term survival of ESCC patients, which might be achieved by increasing PD-L1 + ECs and PD-L1 + TAMs and decreasing aCTLs and NKs in TME.

Effects of Broad-Spectrum Antibiotic (Florfenicol) on Resistance Genes and Bacterial Community Structure of Water and Sediments in an Aquatic Microcosm Model.

This study evaluates the effects of a broad-spectrum antibiotic (florfenicol) on antibiotic resistance genes (ARGs) and bacterial community structure in aquatic environments. We constructed an indoor aquatic microcosm model, adding different concentrations of florfenicol (0.1, 1, 10, 100 mg L−1), and water and sediment samples were collected after 0, 7, 30, and 60 days. qPCR and 16S rDNA amplicon sequencing were used to study the changes in the ARGs and bacterial community structure of the collected samples. The results show that the inclusion of florfenicol resulted in an increased abundance of the floR and optrA genes. Adding 100 mg L−1 florfenicol to the water increased the abundance of optrA gene copies with the maximum on the Day 7, and increased the abundance of floR gene copies with the maximum on Day 30. Adding 100 mg L−1 florfenicol to the sediment increased the abundance of floR and optrA genes by one order of magnitude on Day 60. Meanwhile, the average number of operational taxonomic units (OTUs) in the water samples was 257, and the average number of OTUs in sediment samples was 823. The bacterial community diversity and richness in sediments were higher than those in water. The difference between the maximal and minimal values of the Shannon diversity index in the water and sediment samples was 4.36 and 1.95, respectively. The effect of florfenicol on the bacterial community structure in water was much higher than that in sediment. At 30 days, the diversity index and richness index of the florfenicol treatment groups with 1 and 10 mg L−1 concentrations began to increase; at 60 days, the diversity and richness indices of the 100 mg L−1 florfenicol treatment group began to increase. The samples at the same sampling time in the sediments clustered closer together. The results of this study provide a scientific basis for guiding the rational use of florfenicol in aquaculture, maintaining a healthy and stable microecological environment in aquaculture, and provide theoretical data for environmental ecological risk assessment and safety management caused by microbial resistance under the abuse of florfenicol.

Seasonal dynamics survey and association analysis of microbiota communities, antibiotic resistance genes distribution, and biotoxicities characterization in landfill-leachate

ObjectiveTo reflect the potential and intrinsic association among microbiota structure, antibiotic resistance genes distribution and biological toxicity of landfill-leachate according to seasonal change, and accurately assess the potential threat of leachate to the surrounding environment. MethodsOn the basis of the leachate water quality monitoring data from January to December 2020, principal component analysis (PCA) was used to identify the main pollutants in the leachate; Vibrio fischeri luminescence inhibition test was used to detect the comprehensive biotoxicity of the leachate; 16S rDNA amplicon sequencing was used for leachate microbiota. q-PCR was used to detect the class 1 integron (intI1), and eight antibiotic resistance genes (sul1, sul2, tetA, tetB, tetM, tetQ, mefA, and mexF); Canonical correspondence (CCA) analysis was carried out for the association analysis. ResultThe biotoxicity of leachate in the second quarter was the highest. The dominant phylum of leachate microbiota from 1st quarters to 4th quarters was Proteobacteria (94.97 %, 85.43 %, 88.20 %, and 84.11 %), and the dominant genera were Thiomonas (60.41 %, 26.83 %, 25.66 %, and 30.51 %), Pseudomonas (5.89 %, 1.86 %, 0.68 %, and 4.72 %), Desulfurella (8.52 %, 0.57 %, 3.81 %, and 8.25 %), and Acidithiobacillus (4.71 %, 0.69 %, 0.87 %, and 5.91 %); Nitrospirillum was negatively correlated with chemical oxygen demand (COD) (R=−0.561, P = 0.008) and five-day biochemical oxygen demand (BOD5) (R=−0.591, P = 0.005); Limnohabitans was positively correlated with pH (R=0.444, P = 0.044). Four AR genes (sul1, sul2，tetM, and tetQ) were detected in all the samples, while the second quarter had the highest concentration of sul1(6.31 ± 0.49 lg copies/ng DNA), tetM (3.01 ± 1.38 lg copies/ng DNA) and tetQ (3.64 ± 0.90 lg copies/ng DNA). ConclusionAs the mature landfill, the quality of this leachate met the pollution control standards for domestic waste landfills. Thiomycetes, Pseudomonas, Desulfurization, and Thiopterus acidophyllum constitute the dominant microbiota. However, leachate in the second quarter had more serious contamination, the higher biotoxicity, higher concentration of AR genes, together with higher microbiota richness and diversity, which deserved more attention for the potential threat to the surrounding environment.

Strong Linkage Between Symbiotic Bacterial Community and Host Age and Morph in a Hemipteran Social Insect.

The relationships between symbionts and insects are complex, and symbionts usually have diverse ecological and evolutionary effects on their hosts. The phloem sap-sucking aphids are good models to study the interactions between insects and symbiotic microorganisms. Although aphids usually exhibit remarkable life cycle complexity, most previous studies on symbiotic diversity sampled only apterous viviparous adult females or very few morphs. In this study, high-throughput 16S rDNA amplicon sequencing was used to assess the symbiotic bacterial communities of eleven morphs or developmental stages of the social aphid Pseudoregma bambucicola. We found there were significant differences in bacterial composition in response to different morphs and developmental stages, and for the first time, we revealed male aphids hosted very different symbiotic composition featured with low abundance of dominant symbionts but high diversity of total symbionts. The relative abundance of Pectobacterium showed relatively stable across different types of samples, while that of Wolbachia fluctuated greatly, indicating the former may have a consistent function in this species and the latter may provide specific function for certain morphs or developmental stages. Our study presents new evidence of complexity of symbiotic associations and indicates strong linkage between symbiotic bacterial community and host age and morph.

Fecal Microbiota Transplantation Combined with a Low FODMAP Diet for the Treatment of Irritable Bowel Syndrome with Predominant Diarrhea.

Background Fecal microbiota transplantation (FMT) has been found to be effective in irritable bowel syndrome with predominant diarrhea (IBS-D). We conducted this study to determine the impact of a low FODMAP diet (LFD) on the gut microbiota and the efficacy of FMT in the treatment of IBS-D. Methods A retrospective analysis of a single-arm open-label prospective study was conducted to investigate the impact of FMT alone (n = 40) and FMT+LFD (n = 40) in refractory IBS-D. The IBS-quality of life (QOL), IBS-severity scoring system (SSS), gastrointestinal symptom rating scale (GSRS), Hamilton anxiety scale (HAMA), and Hamilton depression scale (HAMD) were used to evaluate the efficacy, and partial 16S rDNA amplicon sequencing was used to profile the microbiota. Results The response rates were higher in the FMT+LFD group than in the FMT group (1 mo, 3 mo, 6 mo: 70.0% vs. 55.0%, 67.5% vs. 57.5%, 62.5% vs. 27.5%, respectively). The FMT+LFD group showed significantly better improvement in IBS-QOL at 1, 3, and 6 months; IBS-SSS at 6 months; and GSRS at 1 month compared to FMT alone. Changes in HAMA and HAMD were similar in the two groups. The LFD significantly upregulated the FMT-induced microbial diversity (OTUs: 666 vs. 574, Adonis: P = 0.02) and significantly strengthened the upregulation of Bacteroides, Alistipes, and Ruminococcaceae_UCG-002 and the downregulation of Bifidobacterium. Conclusion An LFD enhanced the efficacy of FMT, increased the gut microbial diversity after FMT, and strengthened the inhibitory effect of FMT on conditional pathogens.

Dietary Efficacy Evaluation by Applying a Prediction Model Using Clinical Fecal Microbiome Data of Colorectal Disease to a Controlled Animal Model from an Obesity Perspective.

Obesity associated with a Western diet such as a high-fat diet (HFD) is a known risk factor for inflammatory bowel disease (IBD) and colorectal cancer (CRC). In this study, we aimed to develop fecal microbiome data-based deep learning algorithms for the risk assessment of colorectal diseases. The effects of a HFD and a candidate food (Nypa fruticans, NF) on IBD and CRC risk reduction were also evaluated. Fecal microbiome data were obtained from 109 IBD patients, 111 CRC patients, and 395 healthy control (HC) subjects by 16S rDNA amplicon sequencing. IBD and CRC risk assessment prediction models were then constructed by deep learning algorithms. Dietary effects were evaluated based on fecal microbiome data from rats fed on a regular chow diet (RCD), HFD, and HFD plus ethanol extracts or water extracts of NF. There were significant differences in taxa when IBD and CRC were compared with HC. The diagnostic performance (area under curve, AUC) of the deep learning algorithm was 0.84 for IBD and 0.80 for CRC prediction. Based on the rat fecal microbiome data, IBD and CRC risks were increased in HFD-fed rats versus RCD-fed rats. Interestingly, in the HFD-induced obesity model, the IBD and CRC risk scores were significantly lowered by the administration of ethanol extracts of NF, but not by the administration of water extracts of NF. In conclusion, changes in the fecal microbiome of obesity by Western diet could be important risk factors for the development of IBD and CRC. The risk prediction model developed in this study could be used to evaluate dietary efficacy.

The Gut Bacteria Dysbiosis Contributes to Chronic Graft-Versus-Host Disease Associated With a Treg/Th1 Ratio Imbalance.

IntroductionDysbiosis of gut bacteria has been discovered in a large number of autoimmune diseases. However, the influence of the gut bacteria in the mice model of chronic sclerodermatous graft-versus-host disease (Scl-GVHD), a disease that resembles an autoimmune disease characterized by chronic inflammation of multiple organs, such as skin, remains elusive. Here, we explore the role of gut bacteria in an Scl-cGVHD mice model.MethodsWe established a mouse model of Scl-cGVHD, collected fecal flora, analyzed the composition, and diversity of intestinal flora using 16S rDNA amplicon sequencing, and detected the proportion of Treg and Th1 cells in splenocytes of Scl-cGVHD mice. To verify the immunoregulatory effect of Scl-cGVHD intestinal flora, we prepared bacterial extracts, co-cultured with splenocytes in vitro, and used flow cytometry to detect T cell differentiation and cytokine secretion.ResultsBy examining T-cell differentiation in splenocytes of cGVHD mice, we found that Treg cells were significantly reduced (15.27 ± 0.23 vs. 12.23 ± 0.47, p = 0.0045) and Th1 cells were increased (1.54 ± 0.18 vs. 6.68 ± 0.80, p = 0.0034) in cGVHD mice. Significant differences were observed in the composition and diversity of the gut bacteria in mice with Scl-cGVHD versus without GVHD. Analysis of mice fecal bacteria samples (n = 10, 5 Scl-cGVHD and 5 Non-GVHD) showed significant separation [R = 0.732, p = 0.015, non-parametric analysis (ANOSIM)] in Scl-cGVHD and non-GVHD mice. The abundance of the family and genus Ruminococcaceae bacteria decreased and the family Lachnospiraceae and limited to the species Lachnospiraceae_bacterium_DW17 increased in Scl-cGVHD mice. In vitro results of the cellular level study suggest that the bacteria extracts of gut microbiota from Scl-cGVHD mice modulated the splenic T cells toward differentiation into CD4+IFN-γ+ Th1 cells (14.37 ± 0.32 vs. 10.40 ± 2.19, p = 0.036), and the percentage of CD4+CD25+Foxp3+ Tregs decreased (6.36 ± 0.39 vs. 8.66 ± 0.07, p = 0.001) compared with the non-GVHD mice. In addition, the secretion of proinflammatory interferon- γ (IFN-γ) cytokine in the supplement of cellular culture was increased (4,898.58 ± 235.82 vs. 4,347.87 ± 220.02 pg/ml, p = 0.042) in the mice model of the Scl-cGVHD group, but anti-inflammatory interleukin (IL)-10 decreased (7,636.57 ± 608.05 vs. 9,563.56 ± 603.34 pg/ml, p = 0.018).ConclusionOur data showed the different composition and diversity of gut bacteria in the Scl-cGVHD mice. The dysbiosis of gut bacteria may regulate the differentiation ratio of Treg and Th1 cells, which was associated with Scl-cGVHD.

Protective Effect of Shengmaiyin in Myocardial Hypertrophy-Induced Rats: A Genomic Analysis by 16S rDNA.

Background The gut-cardiac axis theory provides new insights into the complex mechanisms of cardiac hypertrophy and provides new therapeutic targets. Cardiac hypertrophy is a risk factor for heart failure. Shengmaiyin (SMY) is a traditional Chinese medicine formula with clear effects in the treatment and prevention of cardiac hypertrophy, but the mechanism by which it improves cardiac hypertrophy is still unclear. Therefore, this study aimed to investigate the protective effect and mechanism of SMY on isoproterenol (ISO)-induced myocardial hypertrophy in rats. Methods First, various pharmacodynamic methods were used to evaluate the therapeutic effect of SMY on ISO-induced myocardial hypertrophy in rats. Then, 16S rDNA amplicon sequencing technology was used to study the effect of SMY on the intestinal flora of rats with myocardial hypertrophy. Finally, the mechanism underlying the effect of SMY on cardiac hypertrophy was predicted by bioinformatics network analysis and verified by Western blotting. Results SMY increased ejection fraction (EF%) and left ventricular fractional shortening (FS%), ameliorated myocardial cell injury and fibrosis, regulated blood lipids and energy metabolism, and decreased cardiac hypertrophy marker gene expression. The gut microbiota of ISO-induced myocardial hypertrophy rats were significantly changed, while SMY effectively ameliorated the dysbiosis of the intestinal flora in rats with myocardial hypertrophy, especially Prevotella 9, Lactobacillus, and Clostridium. Mechanistic studies have shown that the anticardiac hypertrophy effect of SMY is related to the inhibition of the expression of HIF1α/PPAR signalling pathway-related proteins. Conclusion SMY significantly improves cardiac function, relieves myocardial cell fibrosis and necrosis, resists cardiac hypertrophy, improves blood lipid metabolism and energy metabolism, regulates intestinal microbial disturbance, and protects the heart.

Genotype and organ effect on the occupancy of phyllosphere prokaryotes in different rice landraces.

Bacteria community provides essential ecological services to rice plants. The bacterial diversity of rice varies across host plant genotype and organs. This study employed 16S rDNA amplicon sequencing to characterise the bacterial community associated with three rice landraces using leaf blade and stem samples. The prokaryotic community found in these rice landraces comprised of two kingdoms, 12 phyla, 25 classes, 40 orders, 80 families, and 118 genera. Proteobacteria (53.9%) was the most abundant phylum. The most abundant genus was an undefined genus under Cyanobacteria (33.0%). Homogeneity of prokaryotic community was observed across the three rice landraces, which may suggest a high similarity in biological and genetical properties of the rice landraces. The difference in prokaryotic composition between leaf blade and stem was depicted based on principal coordinate analysis. This study observed that the prokaryotic inhabitants in rice plants is predominantly determined by rice plant organs.

Year-Long Microbial Succession on Microplastics in Wastewater: Chaotic Dynamics Outweigh Preferential Growth.

Microplastics are a globally-ubiquitous aquatic pollutant and have been heavily studied over the last decade. Of particular interest are the interactions between microplastics and microorganisms, especially the pursuit to discover a plastic-specific biome, the so-called plastisphere. To follow this up, a year-long microcosm experimental setup was deployed to expose five different microplastic types (and silica beads control) to activated aerobic wastewater in controlled conditions, with microbial communities being measured four times over the course of the year using 16S rDNA (bacterial) and ITS (fungal) amplicon sequencing. The biofilm community shows no evidence of a specific plastisphere, even after a year of incubation. Indeed, the microbial communities (particularly bacterial) show a clear trend of increasing dissimilarity between plastic types as time increases. Despite little evidence for a plastic-specific community, there was a slight grouping observed for polyolefins (PE and PP) in 6–12-month biofilms. Additionally, an OTU assigned to the genus Devosia was identified on many plastics, increasing over time while showing no growth on silicate (natural particle) controls, suggesting this could be either a slow-growing plastic-specific taxon or a symbiont to such. Both substrate-associated findings were only possible to observe in samples incubated for 6–12 months, which highlights the importance of studying long-term microbial community dynamics on plastic surfaces.

Tremella fuciformis polysaccharides alleviate induced atopic dermatitis in mice by regulating immune response and gut microbiota.

Atopic dermatitis (AD), characterized by severe pruritus, immune imbalance, and skin barrier dysfunction, has a high incidence worldwide. Recent evidence has shown that the modulation of gut microbiota is crucial for alleviating clinical symptoms of AD. Tremella fuciformis polysaccharides (TFPS) have been demonstrated to have a variety of biological activities such as immunomodulatory, anti-tumor, antioxidant, anti-inflammatory, neuroprotective, hypoglycemic and hypolipidemic effects. However, their effects on AD treatment have never been investigated. In this study, we compared the therapeutic effects of topical or oral administration of TFPS on AD in dinitrofluorobenzene (DNFB)-induced AD mice. Both topical application and oral administration of TFPS led to improvement on transdermal water loss, epidermal thickening, and ear edema in AD mice, but the oral administration showed significantly better efficacy than the topical application. The TFPS treatment increased the proportion of CD4 (+) CD25 (+) Foxp3 (+) regulatory T cells in mesenteric lymph nodes. Additionally, the non-targeted metabolomics and sequencing of 16S rDNA amplicons were performed, revealing metabolite modulation in feces and changed composition of gut microbiota in mice, which were induced for AD-like disorder and treated by oral administration of TFPS. Collectively, these data suggest that the oral administration of TFPS may constitute a novel effective therapy for AD, with underlying mechanisms associated with the regulation of immune response, and improvement of both metabolism and the composition of intestinal microbiota.

Trimethylamine-N-oxide (TMAO) mediates the crosstalk between the gut microbiota and hepatic vascular niche to alleviate liver fibrosis in nonalcoholic steatohepatitis.

Liver fibrosis is one main histological characteristic of nonalcoholic steatohepatitis (NASH), a disease paralleling a worldwide surge in metabolic syndromes with no approved therapies. The role of the gut microbiota in NASH pathogenesis has not been thoroughly illustrated, especially how the gut microbiota derives metabolites to influence the distal liver in NASH. Here, we performed 16S rDNA amplicon sequencing analysis of feces from a mouse NASH model induced by a Western diet and CCl4 injury and found genera under Streptococcaceae, Alcaligenaceae, Oscillibacter, and Pseudochrobactrum, which are related metabolites of TMAO. Injection of the gut microbial metabolite TMAO reduced the progression of liver fibrosis in the mouse NASH model. Further analysis revealed that the anti-fibrotic TMAO normalized gut microbiota diversity and preserved liver sinusoidal endothelial cell integrity by inhibiting endothelial beta 1-subunit of Na (+), K (+)-ATPase (ATP1B1) expression. Collectively, our findings suggest TMAO-mediated crosstalk between microbiota metabolites and hepatic vasculature, and perturbation of this crosstalk disrupts sinusoidal vasculature to promote liver fibrosis in NASH.

High-fat-diet-induced gut microbiome changes in mice

Obesity is one of the most severe public health problems and may result in many other related diseases, such as heart disease, diabetes, and stroke. Living habits, particularly excessive caloric intake, are a vital contributor to increasing obesity incidence worldwide. Recent studies have revealed an intimate cross-talk between living habits and the gut microbiome. The gut microbiome has also been identified as a critical player in the cause of obesity. Here we used 16S rDNA amplicon sequencing to investigate changes in the gut microbiome composition of mice fed with a high-fat diet (HFD). The total number of OTUs, Chao index, and Shannon index of the gut microbiome showed an increase in the abundance of specific gut microbiome species. Alternatively, Simpson index indicated a decrease in gut microbiome diversity after HFD feeding. We also found that HFD leads to augments in Firmicutes:Bacteroidetes ratio mainly caused by increased Firmicutes. The total abundance of Bacteroidetes was not changed at the phylum level, while at the family level, both Rikenellaceae and Bacteroidaceae showed a significant increase after the HFD. Additionally, after HFD, we found an increase in the abundance of Proteobacteria related to inflammation and a significant decrease in the proportion of Verrucomicrobia. Our results show that HFD induces a broad gut microbiome change in mice.

Therapeutic effect of fecal microbiota transplantation on chronic unpredictable mild stress-induced depression.

Background and objectiveDepression is a complex neuropsychiatric disease with extensive morbidity. Its pathogenesis remains unclear, and it is associated with extremely low rates of cure and complete remission. It is vital to study the pathogenesis of depression to develop effective treatments. This study aimed to explore the therapeutic effects and mechanisms of fecal microbiota transplantation (FMT) for the treatment of depression in rats.MethodsThirty Sprague-Dawley (SD) rats were randomly divided into three groups: control, chronic unpredictable mild stress (CUMS) to model depression, and CUMS+FMT. For the CUMS and CUMS+FMT groups, after CUMS intervention (four weeks), the rats were given normal saline or FMT (once/week for three weeks), respectively. Behavior, colonic motility, 16S rDNA amplicon sequencing, and untargeted metabolomics on fecal samples were compared between the three rat groups. The following markers were analyzed: 5-hydroxytryptamine (5-HT), gamma-aminobutyric acid (GABA), glutamate (Glu), and brain-derived neurotrophic factor (BDNF) levels in the hippocampus; glucagon-like peptide 1 (GLP-1), lipopolysaccharide (LPS), and interleukin (IL)-6 levels in the serum; and GLP-1, GLP-1 receptor (GLP-1R), and serotonin 4 receptor (5-HT4R) levels in colonic tissues.ResultsFMT improved symptoms of depression and colonic motility in rats exposed to CUMS. The expression levels of 5-HT, GABA, BDNF, and other biochemical indices, significantly differed among the three groups. Meanwhile, the intestinal microbiota in the CUMS+FMT group was more similar to that of the control group with a total of 13 different fecal metabolites.ConclusionFMT exerted antidepressant effects on CUMS-induced depression in rats, and the mechanism involved various neurotransmitters, inflammatory factors, neurotrophic factors, and glucagon-like peptides.

Intestinal aberrant sphingolipid metabolism shaped-gut microbiome and bile acids metabolome in the development of hepatic steatosis.

Conjugated bile acids (CBAs) play major roles in hepatic gene regulation via nuclear S1P-inhibited histone deacetylase (HDACs). Gut microbiota modifies bile acid pool to generate CBAs and then CBAs returned to liver to regulate hepatic genes, fatty liver, and non-alcoholic fatty liver disease (NAFLD). However, it is not yet known how the gut microbiota was modified under the environment of inflammatory bowel disease (IBD). Here, we revealed that aberrant intestinal sphingosine kinases (SphKs), a major risk factor of IBD, modified gut microbiota by increasing the proportions of Firmicutes and Verrucomicrobia, which were associated with the increase in CBAs. When exposed to a high-fat diet (HFD), sphingosine kinases 2knockout (SphK2KO) mice developed more severity of intestinal inflammation and hepatic steatosis than their wild-type (WT) littermates. Due to knockdown of nuclear SphK2, Sphk2KO mice exhibited an increase in sphingosine kinases 1 (SphK1) and sphingosine-1-phosphate (S1P) in intestinal epithelial cells. Therefore, the microbiota was modified in the environment of the SphK1/S1P-induced IBD. 16S rDNA amplicon sequencing of cecal contents indicated an increase of Firmicutes and Verrucomicrobia. Ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) measured an increase in CBAs, including taurocholic acid (TCA), taurodeoxycholic acid (TDCA), and glycocholic acid (GCA), in cecal contents and liver tissues of Sphk2KO mice. These CBAs accumulated in the liver promoted hepatic steatosis through downregulating the acetylation of H3K9, H3K14, H3K18 and H3K27 due to the CBAs-S1PR2-nuclear SphK2-S1P signaling pathway was blocked in HFD-SphK2KO mice. In summary, intestinal aberrant sphingolipid metabolism developed hepatic steatosis through the increase in CBAs associated with an increase in Firmicutes and Verrucomicrobia.

Environmental Selection Shapes Bacterial Community Composition in Traditionally Fermented Maize-Based Foods from Benin, Tanzania and Zambia.

Natural (microbial) communities are complex ecosystems with many interactions and cross-dependencies. Among other factors, selection pressures from the environment are thought to drive the composition and functionality of microbial communities. Fermented foods, when processed using non-industrial methods, harbor such natural microbial communities. In non-alcoholic fermented foods the fermenting microbiota is commonly dominated by 4–10 species of bacteria, which make them suitable model systems to study ecosystem assembly and functioning. In this study, we assess the influence of the environment on the composition of microbial communities of traditional fermented products from Africa. We compare differences between microbial communities that are found in similar products but come from different countries, hypothesizing they experience different environmental selection pressures. We analyzed bacterial community composition in 36 samples of various cereal-based fermented foods from Benin, Tanzania and Zambia using 16S rDNA amplicon sequencing. The differential abundance analysis indicates that the bacterial communities of fermented foods from the three countries are dominated by mostly lactic acid bacteria belonging to the genera of Lactobacillus, Weisella and Curvibacter. The samples from Zambia contain the most dissimilar microbial communities in comparison with samples from Benin and Tanzania. We propose this is caused by the relatively low temperature in Zambia, suggesting that indeed environmental selection can shape community composition of fermenting microbes.

Protective effects of (R)-enantiomers but not (S)-enantiomers of β2-adrenergic receptor agonists against acute colitis: The role of β2AR

The outcomes of ulcerative colitis (UC) treatment remain unsatisfactory. Salbutamol is a β2-adrenergic receptor (β2AR) agonist that is frequently used to treat human airway diseases, and it is a chiral drug with (RS)-isomers. However, the effects of (RS)-enantiomers of this drug on acute ulcerative colitis remain unknown. The present work determined and compared the effects of different chiral β2AR agonists in acute colitis. Acute colitis was established in mice with 3% dextran sulfate sodium and the mice were orally administered different salbutamol isomers. Body weight loss, colon length, disease activity index (DAI), and colon histopathology were assessed. Inflammatory cytokine levels were detected by ELISA. Colonic biopsies were collected from colitis patients. 16S rDNA amplicon sequencing was carried out to assess the composition and relative abundance of the gut microbiome. The expression of M1 and M2 macrophage markers in the colon were assessed by immunofluorescence staining and Western blotting. The results revealed that (R)-salbutamol prevented body weight loss and colonic shortening, decreased the DAI and histopathological scores, and reduced splenomegaly and inflammatory cytokine levels significantly better than (RS)-salbutamol and (S)-salbutamol. (R)-salbutamol downregulated levels of inflammatory protein in LPS-induced human colon tissue specimens. Furthermore, (R)-salbutamol ameliorated gut dysbiosis and macrophage polarization in mice with colitis. The β2AR antagonist ICI-118551 reversed the effect of (R)-salbutamol in ameliorating acute colitis. Taken together, (R)-salbutamol ameliorated the mice with acute colitis, which can serve as a new candidate or lead compound for UC treatment.

Antidepressant Shugan Jieyu Capsule Alters Gut Microbiota and Intestinal Microbiome Function in Rats With Chronic Unpredictable Mild Stress -Induced Depression.

Shugan Jieyu Capsule (SG) has been widely used in China to treat mild to moderate depression. Hypericum perforatum L. (St John’s Wort, SJW) is the main ingredient of SG and has been used as herbal medicine to treat depression in western countries. However, it is known that SJW has low bioavailability and does not easily get through the blood-brain barrier. Therefore, how SG plays an antidepressant effect in the central nervous system (CNS) remains an urgent problem to be solved. Mounting research has described the relationship between antidepressants and intestinal microbiota to illuminate antidepressive mechanisms in the CNS. We aimed to investigate the effects of therapy with SG on the function of gut microbiota and intestinal microbiota in rats with chronic unpredictable mild stress (CUMS)-induced depression. The psychophysiological state and the hypothalamic-pituitary-adrenal axis function of rats are evaluated through behavioral experiments, corticosterone levels, serotonin levels, and adrenal index measurements. 16S rDNA amplicon sequencing is used to test the changes in gut microbiota and make functional predictions of genes. With treatment of SG, the depression-like behaviors of CUMS-induced rats were reversed; the corticosterone levels and the adrenal index decreased significantly; the level of serotonin increased significantly; and the alpha and beta diversity analysis of microbiota showed an increase in the richness and uniformity of the flora were increased. SG regulated the relative abundance of Actinobacteria, Erysipelotrichaceae, Bifidobacteriaceae, Atopobiaceae, Dubosiella, and Bifidobacterium; Linear discriminant analysis effect size analysis demonstrated that Lactobacillaceae (family level), Lactobacillus (genus level), Lactobacillales (order level), Bacilli (class level), and Lactobacillus-reuteri (species level) were biomarkers in the SG group samples, and also likely to modulate metabolic pathways, such as those involved in carbohydrate metabolism, amino acid metabolism, and signal transduction. These data clearly illustrated the effect of SG on gut microbiome, thus laying the foundation for uncovering more insights on the therapeutic function of the traditional Chinese antidepressants. The potential of SG on mechanisms of antidepression to alter gut microbiota and intestinal microbiome function exposed to CUMS can be explored.

Microbial water quality of the copepod Acartia tonsa in cultures for use as live feed

Live feed is important for rearing of marine larvae in the aquaculture industry and copepods have shown some superior qualities compared to Artemia and rotifers. Whereas the nutritional composition is one factor contributing to this, the mechanisms behind better performance are not fully known. Microbial conditions could be an important contributor. We characterized the microbiota associated with intensive rearing of the calanoid copepod Acartia tonsa by culture-dependent and independent methods. During three independent 14-days production cycles, qualitative and quantitative aspects of incoming water, algal cultures (Rhodomonas baltica), and copepod production tanks were measured using flow cytometry with RNA-staining, plate counts, 16S-rDNA amplicon sequencing and hemolytic activity. Assessment of the microbial quality was done based on r/K-theory, presence/absence of hemolytic activity and taxonomic characterization. None of the sample types showed detectable bacteria with hemolytic activity. The incoming water was dominated by opportunistic r-strategists, whereas the algae reservoir was dominated by K-strategist. The production tank had dominance of r-strategists. Abundance of genera with known pathogenic members were not detected in any samples, except the incoming water samples. However, the incoming water contributed with only 1–3% of the cells to the production tank. Because of this, the intensively reared A. tonsa cultures appear to have a good microbial water quality as live feed for marine larvae, and the microbial water quality may contribute to the good results obtained when using copepods during first feeding.

Toxic effects of long-term dual or single exposure to oxytetracycline and arsenic on Xenopus tropicalis living in duck wastewater

Direct discharge of aquaculture wastewater may have toxic effects, due to the presence of heavy metals, antibiotics, and even resistant pathogens, but little attention has been given. Here, tanks simulating a wild ecosystem were built to study the effects of long-term exposure to duck wastewater containing oxytetracycline (OTC) and/or arsenic (As) on the growth, physiological function, and gut microbiota evolution of Xenopus tropicalis. The results showed that duck wastewater had no apparent impact on X. tropicalis, but the impact increased significantly (P < 0.05) with exposure to OTC and/or As, especially the impact on body weight and growth rate. Biochemical indicators revealed varying degrees of oxidative stress damage, hepatotoxicity (inflammation, necrosis, and sinusoids), and collagen fibrosis of X. tropicalis in all treated groups after 72 days of exposure, which indirectly inhibited X. tropicalis growth. Moreover, 16S rDNA amplicon sequencing results showed that the gut microbiota structure and metabolic function were perturbed after chronic exposure, which might be the leading cause of growth inhibition. Interestingly, the abundance of intestinal resistance genes (RGs) increased with exposure time owing to the combined direct and indirect effects of stress factors in duck wastewater. Moreover, once the RGs were expressed, the resistance persisted for at least 24 days, especially that conferred by tetA. These results provide evidence of the toxic effects of DW containing OTC (0.1–4.0 mg/L) and/or As (0.3–3.5 µg/L) on amphibians and indicate that it is vital to limit the usage of heavy metals and antibiotics on farms to control the biotoxicity of wastewater.