Metagenomic analysis of Raphidiopsis raciborskii microbiome: beyond the individual

Raphidiopsis raciborskii is a toxic, invasive bacteria with a defined biogeographic pattern attributed to the generation of ecotypes subjected to local environmental filters and to phenotypic plasticity. The interactions taking place between the cyanobacterium and the other bacteria inhabiting the external polysaccharide-rich matrix surrounding the cells, or phycosphere, may be ecotype-specific and would have different influence on the carbon and nutrient cycling in the ecosystem. Here, we describe the bacterial community or microbiome (assessed by 16S rRNA metagenomics) associated to two R. raciborskii strains that have been described as different ecotypes: the saxitoxin-producer MVCC19 and the non-toxic LB2897. Our results showed that both ecotypes share 50% of their microbiomes and differ in their dominant taxa. The taxon having the highest abundance in the microbiome of MVCC19 was Neorhizobium (22.5% relative abundance), while the dominant taxon in LB2897 was the Planctomycetes SM1A02 (26.2% relative abundance). These groups exhibit different metabolic capabilities regarding nitrogen acquisition (symbiotic nitrogen-fixing in Neorhizobium vs. anammox in SM1A02), suggesting the existence of ecotype-specific microbiomes that play a relevant role in cyanobacterial niche-adaptation. In addition, as saxitoxin and analogues are nitrogen-rich (7 atoms per molecule), we hypothesise that saxitoxin-producing R. raciborskii benefits from external sources of nitrogen provided by the microbiome bacteria. Based on these findings, we propose that the mechanisms involved in the assembly of the cyanobacterial microbiome community are ecotype-dependent.

Strain-Specific Effects of Bifidobacterium longum on Hypercholesterolemic Rats and Potential Mechanisms

Hypercholesterolemia is an independent risk factor of cardiovascular disease, which is among the major causes of death worldwide. The aim of this study was to explore whether Bifidobacterium longum strains exerted intra-species differences in cholesterol-lowering effects in hypercholesterolemic rats and to investigate the potential mechanisms. SD rats underwent gavage with each B. longum strain (CCFM 1077, I3, J3 and B3) daily for 28 days. B. longum CCFM 1077 exerted the most potent cholesterol-lowering effect, followed by B. longum I3 and B3, whereas B. longum B3 had no effect in alleviating hypercholesterolemia. Divergent alleviation of different B. longum strains on hypercholesterolemia can be attributed to the differences in bile salt deconjugation ability and cholesterol assimilation ability in vitro. By 16S rRNA metagenomics analysis, the relative abundance of beneficial genus increased in the B. longum CCFM 1077 treatment group. The expression of key genes involved in cholesterol metabolism were also altered after the B. longum CCFM 1077 treatment. In conclusion, B. longum exhibits strain-specific effects in the alleviation of hypercholesterolemia, mainly due to differences in bacterial characteristics, bile salt deconjugation ability, cholesterol assimilation ability, expressions of key genes involved in cholesterol metabolism and alterations of gut microbiota.

Dental black plaque: metagenomic characterization and comparative analysis with white-plaque

Extrinsic black dental staining is an external dental discoloration of bacterial origin, considered a special form of dental plaque. Currently, there is no definitive therapeutic option for eliminating black stain. This study employed 16S rRNA metagenomics to analyze black stain and white-plaque samples from 27 adult volunteers. Study objectives were to: describe the microbial diversity of adult black stain samples; characterize their taxonomic profile; compare the microbiomes of black stain versus white-plaque from adult volunteers and propose a functional map of the black stain microbiome using PICRUSt2. The black stain microbiome was poorer in species diversity as compared to white-plaque. The five most abundant genera in black stain were Capnocytophaga, Leptotrichia, Fusobacterium, Corynebacterium and Streptococcus. Functional analysis of microbial species revealed conserved and consistent clustering of functional pathways within and between black stain and white-plaque microbiomes. We describe enrichment of heme biosynthetic pathways in black stain. Our results suggest that the dysbiosis in black stain resembles “orally healthy” communities. The increased abundance of heme biosynthetic pathways suggests that heme-dependent iron sequestration and subsequent metabolism are key for black stain formation. Further research should decipher the regulation of heme biosynthetic genes and characterize the temporal sequence leading to colonization and dysbiosis.

Comparison of Productivity and Fecal Microbiotas of Sows in Commercial Farms

Sow productivity, that is, the number of weaned piglets per sow per year, depends on their health status. The gut microbiota is considered a crucial factor in the health of pigs and may affect sow productivity. In the present study, we aimed to investigate the relationship between productivity and the fecal microbiotas of sows in different farms. Feces of sows were collected from 18 farms (10 samples/farm). A total of 90 fecal samples of high-reproductive performance farms were labeled as group H, and 90 fecal samples from low-reproductive performance farms were labeled as group L. Fecal microbiotas were analyzed by 16S rRNA metagenomics, and the organic acids and putrefactive metabolites of the microbiotas were measured. β-diversity was significantly different between groups H and L (p < 0.01), and the relative abundances of 43 bacterial genera, including short-chain fatty acid-producing and fiber-degrading bacteria such as Ruminococcus, Fibrobacter and Butyricicoccus, significantly differed between groups (p < 0.05). In addition, the concentrations of acetate, propionate and n-butyrate were significantly higher in group H than in group L (p < 0.05). In conclusion, sow productivity in farms was likely associated with the compositions of the fecal microbiotas.

Moderate to Severe Ulcerative Colitis Results in Differential Metabolism of Cranberry Polyphenols by the Colon Microbiome Ex Vivo

Objectives The microbiome plays a major role in polyphenol metabolism, producing metabolites that are bioavailable and potentially more bioactive than the polyphenol compounds from which they are derived. A consequence of ulcerative colitis (UC) is a ‘dysbiotic’ microbiome characterized by decreased species diversity, which may impact an individual's capability to metabolize polyphenols into bioavailable metabolites. The objective was to compare the microbial metabolism of cranberry polyphenols between fecal donors that were healthy or had moderate to severe UC to determine if dysbiosis effects polyphenol metabolism. Methods Fecal samples were donated by volunteers with or without moderate to severe UC (n = 9). Fecal samples were cultured anaerobically within 2 hours of defecation in the presence of A and B type proanthocyanidins, flavonols and anthocyanins extracted from cranberry powder. Aliquots were taken at 5 different time points over a 48-hour period and the resulting polyphenol metabolites were quantified via LC-ESI-MS/MS using SRM and authentic standards. 16S rRNA metagenomics analysis was also utilized to assess changes in the composition of the fecal microbiota. Results After 6 hours, healthy individuals produced 2.50, 20.1 and 5.25-fold higher (P &lt; 0.05) concentrations of 3-(3-hydroxyphenyl)propionic acid, 3,4-dihydroxyphenylacetic acid and 3’,4’-dihydroxyphenyl-gamma-valerolactone, respectively. Healthy individuals continued to have significantly higher concentrations of these metabolites over the 48-hour period. UC microbiomes did not produce 3-hydroxyphenylacetic until after 48 hours, suggesting that the microbiome of those with UC may have significantly lower de-hydroxylase activity. Healthy microbiomes contained higher (P &lt; 0.05) proportions of Ruminococcaceae, which could explain their ability to produce higher concentrations of cranberry polyphenol metabolites. Conclusions The microbiome of individuals with UC produces significantly lower cranberry polyphenol metabolites than healthy microbiomes. These results suggest that efficiency of polyphenol metabolism is dependent on health status of the colon and polyphenols could be potential biomarkers of colon health. Funding Sources Ocean Spray Cranberries, Inc, Middleborough, MA.

Comparative Analysis of Fecal Microbiota in Grasscutter (Thryonomys swinderianus) and Other Herbivorous Livestock in Ghana

The grasscutter (also known as the greater cane rat; Thryonomys swinderianus) is a large rodent native to West Africa that is currently under domestication process for meat production. However, little is known about the physiology of this species. In the present study, aiming to provide information about gut microbiota of the grasscutter and better understand its physiology, we investigated the intestinal microbiota of grasscutters and compared it with that of other livestock (cattle, goat, rabbit, and sheep) using 16S rRNA metagenomics analysis. Similar to the other herbivorous animals, bacteria classified as Bacteroidales, Clostridiales, Ruminococcaceae, and Lachnospiraceae were abundant in the microbiome of grasscutters. However, Prevotella and Treponema bacteria, which have fiber fermentation ability, were especially abundant in grasscutters, where the relative abundance of these genera was higher than that in the other animals. The presence of these genera might confer grasscutters the ability to easily breakdown dietary fibers. Diets for grasscutters should be made from ingredients not consumed by humans to avoid competition for resources and the ability to digest fibers may allow the use of fiber-rich feed materials not used by humans. Our findings serve as reference and support future studies on changes in the gut microbiota of the grasscutter as domestication progresses in order to establish appropriate feeding methods and captivity conditions.

Trimethylamine-N-Oxide Postprandial Response in Plasma and Urine Is Lower After Fermented Compared to Non-Fermented Dairy Consumption in Healthy Adults

Trimethylamine-N-oxide (TMAO) can be produced by the gut microbiota from dietary substrates and is associated with cardiovascular disease. While dairy products contain TMAO precursors, the effect of fermented dairy on TMAO metabolism remains unclear. We used plasma and urine samples collected for two randomised cross-over studies to evaluate the effects of fermented dairy consumption on TMAO metabolism. In Study 1, thirteen healthy young men tested a yogurt and an acidified milk during postprandial tests and a two-week daily intervention. In Study 2, ten healthy adults tested milk and cheese during postprandial tests. TMAO and five related metabolites were measured in plasma and urine by LC-MS/MS and NMR. Faecal microbiota composition was assessed in Study 1 (16S rRNA metagenomics sequencing). Fermented milk products were associated with lower postprandial TMAO responses than non-fermented milks in urine (Study 1, p = 0.01; Study 2, p = 0.02) and in plasma, comparing yogurt and acidified milk (Study 1, p = 0.04). Daily consumption of dairy products did not differentially affect fasting TMAO metabolites. Significant correlations were observed between microbiota taxa and circulating or urinary TMAO concentrations. Fermentation of dairy products appear, at least transiently, to affect associations between dairy products and circulating TMAO levels.

Exploring the microbial diversity in oil-contaminated mangrove sediments using 16S rRNA metagenomics

With an area of 390 hectares, the mangrove forests of Nayband Bay are the widest mangrove communities above 27 degrees latitude in the Persian Gulf. They are the last dense, extensive structures of these ecosystems found in the north-west Indian Ocean. Growing industrial activities near the Nayband Bay and the consequent marine pollution has resulted in damage and threatened different marine habitats in recent years. To date, a comprehensive description of the microbial life in the mangrove ecosystem that has been exposed to oil contamination is lacking. This information could significantly contribute to a better overview of the function and resilience of the ecosystem. This work represents a first effort to better understand the Nayband Bay mangrove microbiology by applying 16S rRNA metagenomics. A total of 65,408 readings from the V3–V4 16S rRNA gene regions were obtained from 24 sediment samples, each measuring 440 bp. Most sequences belonged to members of the Proteobacteria phylum (mainly γ-Proteobacteria); however, members of the Bacteroidetes phylum (mainly Flavobacteriia) were also well represented in the samples. We discovered that the community of this ecosystem strongly exhibit typical structures of oil-contaminated marine environments. This is likely due to the growing industrial activity in the area and its consequent marine polluting effects. The use of practicable and applicable bioremediation protocols for habitat restoration in this valuable area is needed.

Rapid analysis of bacterial composition in prosthetic joint infection by 16S rRNA metagenomic sequencing

ObjectivesProsthetic joint infection (PJI) is the most common cause of arthroplasty failure. However, infection is often difficult to detect by conventional bacterial cultures, for which false-negative rates are 23% to 35%. In contrast, 16S rRNA metagenomics has been shown to quantitatively detect unculturable, unsuspected, and unviable pathogens. In this study, we investigated the use of 16S rRNA metagenomics for detection of bacterial pathogens in synovial fluid (SF) from patients with hip or knee PJI.MethodsWe analyzed the bacterial composition of 22 SF samples collected from 11 patients with PJIs (first- and second-stage surgery). The V3 and V4 region of bacteria was assessed by comparing the taxonomic distribution of the 16S rDNA amplicons with microbiome sequencing analysis. We also compared the results of bacterial detection from different methods including 16S metagenomics, traditional cultures, and targeted Sanger sequencing.ResultsPolymicrobial infections were not only detected, but also characterized at different timepoints corresponding to first- and second-stage exchange arthroplasty. Similar taxonomic distributions were obtained by matching sequence data against SILVA, Greengenes, and The National Center for Biotechnology Information (NCBI). All bacteria isolated from the traditional culture could be further identified by 16S metagenomics and targeted Sanger sequencing.ConclusionThe data highlight 16S rRNA metagenomics as a suitable and promising method to detect and identify infecting bacteria, most of which may be uncultivable. Importantly, the method dramatically reduces turnaround time to two days rather than approximately one week for conventional cultures. Cite this article: M-F. Chen, C-H. Chang, C. Chiang-Ni, P-H. Hsieh, H-N. Shih, S. W. N. Ueng, Y. Chang. Rapid analysis of bacterial composition in prosthetic joint infection by 16S rRNA metagenomic sequencing. Bone Joint Res 2019;8:367–377. DOI: 10.1302/2046-3758.88.BJR-2019-0003.R2.