Weiyun Zhu

Amino acid metabolism in intestinal bacteria: links between gut ecology and host health.

Bacteria in the gastrointestinal (GI) tract play an important role in the metabolism of dietary substances in the gut and extraintestinal tissues. Amino acids (AA) should be taken into consideration in the development of new strategies to enhance efficiency of nutrient utilization because they are not only major components in the diet and building blocks for protein but also regulate energy and protein homeostasis in organisms. The diversity of the AA-fermenting bacteria and their metabolic redundancy make them easier to survive and interact with their neighboring species or eukaryotic host during transition along GI tract. The outcomes of the interactions have important impacts on gut health and whole-body homeostasis. The AA-derived molecules produced by intestinal bacteria affect host health by regulating either host immunity and cell function or microbial composition and metabolism. Emerging evidence shows that dietary factors, such as protein, non-digestible carbohydrates, probiotics, synbiotics and phytochemicals, modulate AA utilization by gut microorganisms. Interdisciplinary research involving nutritionists and microbiologists is expected to rapidly expand knowledge about crucial roles for AA in gut ecology and host health.

Characterising the bacterial microbiota across the gastrointestinal tracts of dairy cattle: membership and potential function

The bacterial community composition and function in the gastrointestinal tracts (GITs) of dairy cattle is very important, since it can influence milk production and host health. However, our understanding of bacterial communities in the GITs of dairy cattle is still very limited. This study analysed bacterial communities in ten distinct GIT sites (the digesta and mucosa of the rumen, reticulum, omasum, abomasum, duodenum, jejunum, ileum, cecum, colon and rectum) in six dairy cattle. The study observed 542 genera belonging to 23 phyla distributed throughout the cattle GITs, with the Firmicutes, Bacteroidetes and Proteobacteria predominating. In addition, data revealed significant spatial heterogeneity in composition, diversity and species abundance distributions of GIT microbiota. Furthermore, the study inferred significant differences in the predicted metagenomic profiles among GIT regions. In particular, the relative abundances of the genes involved in carbohydrate metabolism were overrepresented in the digesta samples of forestomaches, and the genes related to amino acid metabolism were mainly enriched in the mucosal samples. In general, this study provides the first deep insights into the composition of GIT microbiota in dairy cattle, and it may serve as a foundation for future studies in this area.

Impact of subacute ruminal acidosis (SARA) adaptation on rumen microbiota in dairy cattle using pyrosequencing

The objective of this study was to evaluate the changes in bacterial populations in the rumen of dairy cattle following adaptation to subacute ruminal acidosis (SARA) using 16S rRNA gene pyrosequencing. Rumen contents were collected from four cattle adapted to either a 40% (control diet, COD) or 70% (SARA induction diet, SAID) concentrate feeds. DNA was extracted from each of the samples. Bacterial 16S rRNA genes of ruminal DNA extracts were PCR amplified with 2 bar coded primer sets and sequenced by 454 pyrosequencing. At a high taxonomic level, the percentage of Proteobacteria and Bacteroidetes were reduced by SAID feeding, whereas Firmicutes and Actinobacteria were more abundant in the SAID than in the COD group. At the genus level, as compared with the COD group, the abundances of Prevotella, Treponema, Anaeroplasma, Papillibacter, Acinetobacter and unclassified populations including unclassified Lentisphaerae, and unclassified bacteria were lower (P < 0.05), while the percentages of Ruminococcus, Atopobium, unclassified Clostridiales and Bifidobacterium were increased (P < 0.05) in the SAID group. Feeding of SAID reduced (P < 0.001) the diversity of the rumen microbial community. Taken together, our findings provide a comprehensive picture of current knowledge of the community structure of the rumen bacterial ecosystem during SARA, and enhance our understanding about the ruminal microbial ecology that may be useful in the prevention of ruminal acidosis.

Changes in abundance of Lactobacillus spp. and Streptococcus suis in the stomach, jejunum and ileum of piglets after weaning

This present study investigated the changes in bacterial community composition, with an emphasis on Lactobacillus spp. and Streptococcus suis populations as potentially beneficial and harmful groups, in the stomach, jejunum and ileum of piglets after weaning (21 days postpartum) by 16S rRNA gene-based methods. Denaturing gradient gel electrophoresis analysis showed that, after weaning, predominant bands related to Lactobacillus spp. disappeared and were replaced by potential pathogenic species, such as Peptostreptococcus anaerobius, Moraxella cuniculi, S. suis and Porphyromonas catoniae. Real-time PCR revealed that the abundances of lactobacilli and Lactobacillus sobrius as a proportion of total bacterial abundance were significantly lower in the stomach, jejunum and ileum of weaned piglets than in 21-day-old piglets. A specific and sensitive real-time PCR assay was developed for quantification of the important pathogen S. suis within gastrointestinal microbiota. The assay showed that S. suis predominated in the stomach samples of weaned piglets with population levels up to 10(7) copies g(-1) digesta, while it was not detected in the stomach before weaning. Streptococcus suis was not dominant in the jejunum and ileum digesta before weaning, but became dominant after weaning, with population levels up to 10(7) copies g(-1) digesta. The results demonstrated for the first time the postweaning dominance of the potentially harmful S. suis in piglet intestine. The results also suggest that the defensive barrier of the stomach can be impaired as S. suis became dominant while the proportion of Lactobacillus populations decreased after weaning, which may further result in an increase of S. suis abundance in the intestine.

A high-grain diet causes massive disruption of ruminal epithelial tight junctions in goats

Alterations in rumen epithelial tight junctions (TJs) at the tissue and molecular levels during high-grain (HG) diet feeding are unknown. Here, 10 male goats were randomly assigned to either a hay diet (0% grain; n = 5) or HG diet group (65% grain; n = 5) to characterize the changes in ruminal epithelial structure and TJ protein expression and localization using scanning and transmission electron microscopy, quantitative real-time PCR, Western blot analysis, and immunofluorescence. After 7 wk of feeding, ruminal free LPS in HG group increased significantly (P < 0.001) compared with the hay group, and free LPS in the peripheral blood was detectable with concentrations of 0.8 ± 0.20 EU/ml, while not detectable in the control, suggesting a leakage of LPS into the blood in the HG group. Correspondingly, the HG-fed goats showed profound alterations in ruminal epithelial structure and TJ proteins, depicted by marked epithelial cellular damage and intercellular junction erosion, down-regulation of TJ proteins claudin-4, occludin, and zonula occludens-1 mRNA and protein expression, as well as redistribution of claudin-1, claudin-4, and occludin. Furthermore, these changes in TJ proteins in the HG group were coupled with the upregulation of mRNA levels for the cytokines TNF-α and IFN-γ in the ruminal epithelia. These results demonstrated for the first time that the HG diet feeding caused disruption of ruminal epithelial TJs that was associated with a local inflammatory response in the rumen epithelium. These findings may provide new insights into understanding the role of TJ proteins in the ruminal epithelial immune homeostasis of ruminants.

Microbiome-metabolome analysis reveals unhealthy alterations in the composition and metabolism of ruminal microbiota with increasing dietary grain in a goat model.

Currently, knowledge about the impact of high-grain (HG) feeding on rumen microbiota and metabolome is limited. In this study, a combination of the 454 pyrosequencing strategy and the mass spectrometry-based metabolomics technique was applied to investigate the effects of increased dietary grain (0%, 25% and 50% maize grain) on changes in whole ruminal microbiota and their metabolites using goat as a ruminant model. We observed a significant influence of HG feeding in shaping the ruminal bacterial community structure, diversity and composition, with an overall dominance of bacteria of the phylum Firmicutes along with a low abundance of Bacteriodetes in the HG group. High-grain feeding increased the number of ciliate and methanogens, and decreased the density of anaerobic fungi and the richness of the archaeal community. The metabolomics analysis revealed that HG feeding increased the levels of several toxic, inflammatory and unnatural compounds, including endotoxin, tryptamine, tyramine, histamine and phenylacetate. Correlation analysis on the combined datasets revealed some potential relationships between ruminal metabolites and certain microbial species. Information about these relationships may prove useful in either direct (therapeutic) or indirect (dietary) interventions for ruminal disorders due to microbial compositional shifts, such as ruminal acidosis.

Diversity, abundance and novel 16S rRNA gene sequences of methanogens in rumen liquid, solid and epithelium fractions of Jinnan cattle

Three methanogen 16S rRNA gene clone libraries were constructed from liquid (LM), solid (SM) and epithelium (EM) fractions taken from the rumen of Jinnan cattle in China. After the amplification by PCR using methanogen-specific primers Met86F and Met1340R, equal quantities of PCR products from the same fractions from each of the four cattle were mixed together and used to construct the three libraries. Sequence analysis showed that the 268 LM clones were divided into 35 phylotypes with 18 sequences of phylotypes affiliated with the genus Methanobrevibacter (84.3% of clones). The 135 SM clones were divided into 19 phylotypes with 11 phylotypes affiliated with the genus Methanobrevibacter (77.8%). The 267 EM clones were divided into 33 phylotypes with 15 phylotypes affiliated with the genus Methanobrevibacter (77.2%). Clones closely related to Methanomicrobium mobile and Methanobrevibacter wolinii were only found in the LM library, and those to Methanobrevibacter ruminantium and Methanobrevibacter gottschalkii only in the SM library. LM library comprised 12.4% unidentified euryarchaeal clones, SM library 23.7% and EM library 25.5%, respectively. Five phylotypes (accession number: EF055528 and EF055531-EF055534) did not belong to the Euryarchaeota sequences we had known. One possible new genus (represented by phylotype E17, accession number EF055528) belonging to Methanobacteriaceae was identified from EM library. Quantitative real-time PCR for the first time revealed that epithelium fraction had significantly higher density of methanogens, with methanogenic mcrA gene copies (9.95 log 10 (copies per gram of wet weight)) than solid (9.26, P < 0.01) and the liquid (8.44, P < 0.001). The three clone libraries also appeared different in Shannon index (EM library 2.12, LM library 2.05 and SM library 1.73). Our results showed that there were apparent differences in the methanogenic diversity and abundance in the three different fractions within the rumen of Jinnan cattle, with Methanobrevibacter species predominant in all the three libraries and with epithelium fraction having more unknown species and higher density of methanogens.

16S ribosomal RNA-based methods to monitor changes in the hindgut bacterial community of piglets after oral administration of Lactobacillus sobrius S1.

16S ribosomal RNA (rRNA) gene based PCR/denaturing gradient gel electrophoresis (DGGE) and real-time PCR were used to monitor the changes in the composition of microbiota in the hindgut of piglets after oral administration of Lactobacillus sobrius S1. Six litters of neonatal piglets were divided randomly into control group and treatment group. At 7, 9, and 11 days of age, piglets in the treatment group orally received a preparation of L. sobrius S1. At 7, 14, 21(weaning), 24, and 35 days of age, one piglet from each litter was sacrificed and digesta samples of hindgut were collected. DGGE analysis of 16S rRNA gene V6-V8 region for all bacteria showed that several populations present in the hindgut of piglets, represented by far-migrating bands, disappeared after weaning. Most of these bands corresponded to Lactobacillus spp. as revealed by sequence analysis. Quantitative real-time PCR specific for lactobacilli further demonstrated that the number of lactobacilli population tended to decrease after the piglets were weaned. Drastic changes of L. amylovorus and L. sobrius in total Lactobacillus populations were also observed in the colon of piglets around weaning, as monitored by 16S rRNA gene V2-V3 region based Lactobacillus-specific PCR-DGGE. Species-specific real-time PCR also revealed that the population of L. sobrius declined apparently in the colon of piglets after weaning. No remarkable changes in the overall microbial community in the hindgut were found between control and treatment groups. However, comparison of DGGE profiles between the two groups revealed a specific band related to Clostridium disporicum that was found in treatment group on day 14. On day 35, a specific band appeared only in the control group, representing a population most closely related to Streptococcus suis (99%). Real-time PCR showed that L. sobrius 16S rRNA gene copies in treatment group were relatively higher than in the control group (10(8.45) vs. 10(6.83)) on day 35, but no significant difference was observed between the two groups.

Effect of increasing levels of bioflavonoids in broiler feed on plasma anti-oxidative potential, lipid metabolites, and fatty acid composition of meat

This study was conducted to investigate the supplemental effects of purified bioflavonoids (genistein and hesperidin), as potential alternatives to plant/herbs or synthetic antioxidants, individually and in combination for fatty acid profile, lipid metabolites, and antioxidant status of broilers. Three hundred sixty 1-d-old broilers were divided into 6 treatment groups: control (basal diet), G5 (5 mg of genistein per kg of feed), and H20 (20 mg hesperidin per kg of feed), whereas the other 3 groups were supplemented with a mixture of genistein and hesperidin (20% genistein + 80% hesperidin) having a dosage of 5 mg•kg(-1) (GH5), 10 mg•kg(-1) (GH10), and 20 mg•kg(-1) (GH20), respectively. Broilers were slaughtered at 42 d, and breast muscle, liver, and blood samples were collected. A dose-dependent increase (P < 0.05) was observed for plasma antioxidant parameters, including total antioxidant capacity, malondialdehyde production, and total superoxide dismutase activity. Cholesterol and triglyceride contents were found to decrease (P < 0.05) in serum and breast muscle. The proportion of total polyunsaturated fatty acids and the ratio of n-6 to n-3 fatty acids and polyunsaturated fatty acids to saturated fatty acids in breast muscles was significantly improved (P < 0.05) by increasing levels of dietary bioflavonoids. The current results implied that dietary bioflavonoids genistein and hesperidin could positively improve the fatty acid and lipid metabolite profile of broiler breast meat in a dose-dependent fashion. Thus, bioflavonoids could be a feasible alternative of antioxidant plants/herbs and synthetic feed additives for the production of healthier chicken meat.

Gut Microbiota: The Brain Peacekeeper

Gut microbiota regulates intestinal and extraintestinal homeostasis. Accumulating evidence suggests that the gut microbiota may also regulate brain function and behavior. Results from animal models indicate that disturbances in the composition and functionality of some microbiota members are associated with neurophysiological disorders, strengthening the idea of a microbiota–gut–brain axis and the role of microbiota as a “peacekeeper” in the brain health. Here, we review recent discoveries on the role of the gut microbiota in central nervous system-related diseases. We also discuss the emerging concept of the bidirectional regulation by the circadian rhythm and gut microbiota, and the potential role of the epigenetic regulation in neuronal cell function. Microbiome studies are also highlighted as crucial in the development of targeted therapies for neurodevelopmental disorders.