Rumen Sampling Methods Bias Bacterial Communities Observed

AbstractThe rumen is a complex ecosystem that plays a critical role in our efforts to improve feed efficiency of cattle and reduce their environmental impacts. Sequencing of the 16S rRNA gene provides a powerful tool to survey shifts in the microbial community in response to feed additives and dietary changes. Oral stomach tubing a cow for a rumen sample is a rapid, cost-effective alternative to rumen cannulation for acquiring rumen samples. In this study, we determined how sampling method, as well as type of sample collected (liquid vs solid), bias the microbial populations observed. The abundance of major archaeal populations was not different at the family level in samples acquired via rumen cannula or stomach tube. Liquid samples were enriched for the order WCHB1-41 (phylum Kiritimatiellaeota) as well as the family Prevotellaceae and had significantly lower abundance of Lachnospiraceae compared with grab samples from the rumen cannula. Solid samples most closely resembled the grab samples; therefore, inclusion of particulate matter is important for an accurate representation of the rumen microbes. Stomach tube samples were the most variable and were most representative of the liquid phase. In comparison with a grab sample, stomach tube samples had significantly lower abundance of Lachnospiraceae, Fibrobacter and Treponema. Fecal samples did not reflect the community composition of the rumen, as fecal samples had significantly higher relative abundance of Ruminococcaceae and significantly lower relative abundance of Lachnospiraceae compared with samples from the rumen.

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Relative and quantitative rhizosphere microbiome profiling result in distinct abundance patterns

10.1101/2021.02.19.431941 ◽

2021 ◽

Author(s):

Hamed Azarbad ◽

Julien Tremblay ◽

Luke D. Bainard ◽

Etienne Yergeau

Keyword(s):

Water Stress ◽

Relative Abundance ◽

Its Region ◽

Cost Effective ◽

Amplicon Sequencing ◽

Rrna Gene ◽

Real Time Quantitative Pcr ◽

Rhizosphere Microbiome ◽

History Of ◽

Almost All

AbstractNext-generation sequencing is recognized as one of the most popular and cost-effective way of characterizing microbiome in multiple samples. However, most of the currently available amplicon sequencing approaches are inherently limited, as they are often presented based on the relative abundance of microbial taxa, which may not fully represent actual microbiome profiles. Here, we combined amplicon sequencing (16S rRNA gene for bacteria and ITS region for fungi) with real-time quantitative PCR (qPCR) to characterize the rhizosphere microbiome of wheat. We show that the increase in relative abundance of major microbial phyla does not necessarily result in an increase in abundance. One striking observation when comparing relative and quantitative abundances was a substantial increase in the abundance of almost all phyla associated with the rhizosphere of plants grown in soil with no history of water stress as compared with the rhizosphere of plants growing in soil with a history of water stress, which was in contradiction with the trends observed in the relative abundance data. Our results suggest that the estimated absolute abundance approach gives a different perspective than the relative abundance approach, providing complementary information that helps to better understand the rhizosphere microbiome.

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Effect of Selected Stilbenoids on Human Fecal Microbiota

Molecules ◽

10.3390/molecules24040744 ◽

2019 ◽

Vol 24 (4) ◽

pp. 744 ◽

Cited By ~ 6

Author(s):

Jose Jaimes ◽

Veronika Jarosova ◽

Ondrej Vesely ◽

Chahrazed Mekadim ◽

Jakub Mrazek ◽

...

Keyword(s):

Gut Microbiota ◽

Relative Abundance ◽

Statistical Tests ◽

16S Rrna Gene Sequencing ◽

Fecal Microbiota ◽

Rrna Gene ◽

Microbial Composition ◽

Genus Clostridium ◽

The Family ◽

Rrna Gene Sequencing

Dietary phenolics or polyphenols are mostly metabolized by the human gut microbiota. These metabolites appear to confer the beneficial health effects attributed to phenolics. Microbial composition affects the type of metabolites produced. Reciprocally, phenolics modulate microbial composition. Understanding this relationship could be used to positively impact health by phenolic supplementation and thus create favorable colonic conditions. This study explored the effect of six stilbenoids (batatasin III, oxyresveratrol, piceatannol, pinostilbene, resveratrol, thunalbene) on the gut microbiota composition. Stilbenoids were anaerobically fermented with fecal bacteria from four donors, samples were collected at 0 and 24 h, and effects on the microbiota were assessed by 16S rRNA gene sequencing. Statistical tests identified affected microbes at three taxonomic levels. Observed microbial composition modulation by stilbenoids included a decrease in the Firmicutes to Bacteroidetes ratio, a decrease in the relative abundance of strains from the genus Clostridium, and effects on the family Lachnospiraceae. A frequently observed effect was a further decrease of the relative abundance when compared to the control. An opposite effect to the control was observed for Faecalibacterium prausnitzii, whose relative abundance increased. Observed effects were more frequently attributed to resveratrol and piceatannol, followed by thunalbene and batatasin III.

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Impacts of Aging and Blackcurrant Supplementation on the Gut Microbiome Profile of Female Mice (P20-031-19)

Current Developments in Nutrition ◽

10.1093/cdn/nzz040.p20-031-19 ◽

2019 ◽

Vol 3 (Supplement_1) ◽

Author(s):

Sang Gil Lee ◽

Cao Lei ◽

Melissa Melough ◽

Junichi Sakaki ◽

Kendra Maas ◽

...

Keyword(s):

Relative Abundance ◽

Gut Microbiome ◽

Health Benefits ◽

Alpha Diversity ◽

Young Female ◽

Rrna Gene ◽

Fecal Samples ◽

Female Mice ◽

Aged Mice ◽

Young Mice

Abstract Objectives Blackcurrant, an anthocyanin-rich berry, has multiple health benefits. The purpose of this study was to examine the impacts of blackcurrant supplementation and aging on gut bacterial communities in female mice. Methods Three-month and 18-month old female mice were provided standard chow diets with or without anthocyanin-rich blackcurrant extract (BC) (1% w/w) for four months. Upon study completion, fecal samples were collected directly from the animals’ colons. Microbiome DNA was extracted from the fecal samples and the V3-V4 regions of their 16S rRNA gene were amplified and sequenced using Results Taxonomic analysis showed a significantly decrease in alpha diversity in aged female mice, compared to young counterparts. BC consumption did not alter the alpha diversity in either young or aged mice compared to control diets. For beta diversity, we observed the clustering was associated with age but not diet. The phylogenic abundance analysis showed that the relative abundance of several phyla, including Firmicutes, Bacteroidetes, Cyanobacteria, Proteobacteria, and Tenericutes was higher in aged compared to young mice. Among them, the abundance of Firmicutes was downregulated by BC in the young but not the aged mice. The abundance of Bacteroidetes was increased by BC in both the young and the aged groups. Noticeably, Verrucomicrobia was the only phylum whose relative abundance was upregulated in the aged female mice compared to the young mice. Meanwhile, its relative abundance in the aged group was suppressed by BC. Interestingly, Desulfovibrio, which is the most representative sulfate-reducing genus, was detectable only in young female mice, and BC increased its relative abundance. Conclusions Our results characterized the gut microbiome compositions in young and aged female mice, and indicated that the gut microbiome of young and aged female mice responded differently to four month BC administration. Through additional research, the microbial alterations observed in this study should be further investigated to inform our understanding of the effect of BC on the gut microbiome, the possible health benefits related to these changes, and the differing effects of BC supplementation across populations. Funding Sources This study was supported by the USDA NIFA Seed Grant (#2016-67018-24492) and the University of Connecticut Foundation Esperance Funds to Dr. Ock K. Chun. We thank the National Institute on Aging for providing aged mice for the project and Just the Berries Ltd. for providing the blackcurrant extract.

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Bacteroidales Diversity in Ring-Billed Gulls (Laurus delawarensis) Residing at Lake Michigan Beaches

Applied and Environmental Microbiology ◽

10.1128/aem.02261-08 ◽

2009 ◽

Vol 75 (6) ◽

pp. 1525-1533 ◽

Cited By ~ 28

Author(s):

Sonja N. Jeter ◽

Colleen M. McDermott ◽

Patricia A. Bower ◽

Julie L. Kinzelman ◽

Melinda J. Bootsma ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Lake Michigan ◽

Rrna Gene ◽

Fecal Indicator ◽

Fecal Samples ◽

Operational Taxonomic Units ◽

The North ◽

The Family ◽

Sample Representation

ABSTRACT This study investigated the occurrence and diversity of Bacteroidales fecal bacteria in gulls residing in the Great Lakes region. Members of this bacterial order have been widely employed as human and bovine host-specific markers of fecal pollution; however, few studies have focused on gulls, which can be a major source of fecal indicator bacteria and pathogens at beaches. We found a low but consistent occurrence of Bacteroidales in gulls at five beaches in three different counties spanning the Wisconsin shoreline of Lake Michigan. The percentages of gulls positive for Bacteroidales were 4 to 8% at beaches in the southern part of the state and 8 to 50% at beaches in the north. Sequencing of 931 clones from seven gull Bacteroidales 16S rRNA gene libraries revealed a large amount of diversity in both individual and pooled gull fecal samples. Two libraries constructed from pooled gull fecal samples (n = 5 and n = 6) did not have a greater richness of sequences than individual samples, suggesting that even within a single gull diversity is high and an extensive sequencing effort is needed to characterize the populations. Estimates of the numbers of operational taxonomic units (OTUs) for the libraries obtained using different similarity levels revealed a large amount of microdiveristy with a limited number of OTUs at the 95% similarity level. Gull sequences were clustered by the beach from which they were collected, suggesting that there were geographic effects on the distribution of Bacteriodales. More than 53% of the 16S rRNA gene sequences from gulls at the southern beaches were associated with the family Porphyromonadaceae, primarily the genus Parabacteroides, whereas sequences from gulls at the northern beaches were comprised of Bacteroidaceae and Prevotellaceae sequences. Comparison of gull sequences with sequences from goose, canine, raccoon, and sewage sources revealed distinct clusters of closely related gull sequences; however, these sequences were widely dispersed across a dendrogram that included all other sources, including previously characterized gull Bacteroidales from other studies, suggesting that geographic influence or simply sample representation plays a greater role in the observed population structure than strictly the host gut environment.

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The effect of essential oils of Zataria multiflora and Mentha spicata on the in vitro rumen fermentation, and growth and deaminative activity of amino acid-fermenting bacteria isolated from Mehraban sheep

Animal Production Science ◽

10.1071/an12244 ◽

2014 ◽

Vol 54 (3) ◽

pp. 299 ◽

Cited By ~ 2

Author(s):

M. Taghavi-Nezhad ◽

D. Alipour ◽

M. D. Flythe ◽

P. Zamani ◽

G. Khodakaramian

Keyword(s):

Essential Oils ◽

Rumen Fermentation ◽

Gas Production ◽

Feed Additives ◽

Rrna Gene ◽

Ammonia Production ◽

Mentha Spicata ◽

Zataria Multiflora ◽

Rumen Microbes

Gas (CO2 and CH4) and ammonia production in the rumen represent major sources of lost carbon and nitrogen, respectively. The essential oils of some plants have been shown to decrease gas and ammonia production by selectively inhibiting rumen microbes. Particularly, those of Zataria multiflora (ZEO; thymol 21%, carvacrol 32%) and Mentha spicata (SEO; carvone 55%) were evaluated in vitro as ruminant-feed additives. The experiments employed mixed rumen microbes and a hyper-ammonia-producing bacterium (HAP) isolated from the rumen of a Mehraban sheep. Both ZEO and SEO decreased in vitro fibre digestibility and also gas production by mixed rumen microbes that were fermenting a typical growing-lamb diet. ZEO decreased ammonia concentration in mixed culture of rumen microbes, but SEO exerted the opposite effect. A bacterial isolate (MT8) was obtained from the rumen of a Mehraban sheep, and the 16S rRNA gene sequence indicated that it was most closely related to Clostridium bifermentans. Isolate MT8 exhibited rapid ammonia production when peptides were the growth substrate, which indicated that MT8 was a HAP. Both oils inhibited the growth and ammonia production of isolate MT8. However, ZEO decreased ammonia production at lower doses, and to a greater degree, than did SEO. These results indicated that both essential oils could potentially be used to modulate rumen fermentation. The detrimental effects on fibre digestion could be problematic in high-forage diets, and this requires further investigation. Isolate MT8 is the first described HAP from the Mehraban sheep rumen. Results on ammonia production by isolate MT8 and mixed rumen microbes indicate differential mode of action of each oil on this parameter.

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Impact of a probiotic, inulin, or their combination on the piglets’ microbiota at different intestinal locations

Beneficial Microbes ◽

10.3920/bm2014.0030 ◽

2015 ◽

Vol 6 (4) ◽

pp. 473-483 ◽

Cited By ~ 10

Author(s):

V.A. Sattler ◽

K. Bayer ◽

G. Schatzmayr ◽

A.G. Haslberger ◽

V. Klose

Keyword(s):

16S Rrna ◽

Gut Microbiota ◽

16S Rrna Gene ◽

Real Time ◽

Relative Abundance ◽

Real Time Pcr ◽

Denaturing Gradient Gel Electrophoresis ◽

Feed Additives ◽

Rrna Gene ◽

16S Rrna Gene Pyrosequencing

Natural feed additives are used to maintain health and to promote performance of pigs without antibiotics. Effects of a probiotic, inulin, and their combination (synbiotic), on the microbial diversity and composition at different intestinal locations were analysed using denaturing gradient gel electrophoresis (DGGE), real-time PCR, and 16S rRNA gene pyrosequencing. Bacterial diversity assessed by DGGE and/or pyrosequencing was increased by inulin in all three gut locations and by the synbiotic in the caecum and colon. In contrast, the probiotic did only affect the microbiota diversity in the ileum. Shifts in the DGGE microbiota profiles of the caecum and colon were detected for the pro- and synbiotic fed animals, whereas inulin profiles were more similar to the ones of the control. 16S rRNA gene pyrosequencing revealed that all three additives could reduce Escherichia species in each gut location, indicating a potential beneficial effect on the gut microbiota. An increase of relative abundance of Clostridiaceae in the large intestine was found in the inulin group and of Enterococcaceae in the ileum of probiotic fed pigs. Furthermore, real-time PCR results showed that the probiotic and synbiotic increased bifidobacterial numbers in the ileum, which was supported by sequencing results. The probiotic and inulin, to different extents, changed the diversity, relative abundance of phylotypes, and community profiles of the porcine microbiota. However, alterations of the bacterial community were not uniformly between gut locations, demonstrating that functionality of feed additives is site specific. Therefore, gut sampling from various locations is crucial when investigations aim to identify the composition of a healthy gut microbiota after its manipulation through feed additives.

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Age-Based Variations in the Gut Microbiome of the Shennongjia (Hubei) Golden Snub-Nosed Monkey (Rhinopithecus roxellana hubeiensis)

BioMed Research International ◽

10.1155/2021/6667715 ◽

2021 ◽

Vol 2021 ◽

pp. 1-17

Author(s):

Lijuan Yao ◽

Xiang Li ◽

Zutao Zhou ◽

Deshi Shi ◽

Zili Li ◽

...

Keyword(s):

Gut Microbiota ◽

Relative Abundance ◽

Gut Microbiome ◽

Developmental Stages ◽

Rrna Gene ◽

Rhinopithecus Roxellana ◽

Fecal Samples ◽

Group 3 ◽

Group 2 ◽

Diverse Groups

The gut microbiota represents a source of genetic and metabolic diversity of a complex polymicrobial ecosystem within its host. To investigate age-based variations of the gut microbiota among Shennongjia golden snub-nosed monkeys (Rhinopithecus roxellana hubeiensis), we characterized the microbial species in fecal samples from 18 Shennongjia golden snub-nosed monkeys evenly pooled into 3 aged groups (Group 1, 1-3 years; Group 2, 5-8 years; Group 3, above 12 years) in Shennongjia, Hubei Province, China. Genomic DNA was extracted from fecal samples, and the 16S rRNA gene V4 region was sequenced using the Illumina high-throughput MiSeq platform PE250. A total of 28 microbial phyla were identified in the gut microbiome of these monkeys with the ten most abundant phyla (i.e., Firmicutes, Bacteroidetes, Verrucomicrobia, Spirochaetes, Tenericutes, Proteobacteria, Planctomycetes, Fibrobacteres, Cyanobacteria, and Euryarchaeota). A total of 1,469 (of 16 phyla and 166 genera), 1,381 (of 16 phyla and 157 genera), and 1,931 (of 19 phyla and 190 genera) operational taxonomic units (OTUs) were revealed in Groups 1, 2, and 3, respectively, with Group 3 containing the most diverse groups of OTUs as revealed by the species relative abundance clustering analysis. These results suggest that the gut microbiota in these monkeys maintain a dynamic status, starting from the early developmental stages of life with the species relative abundance increasing with age. This is the first study to comprehensively characterize the gut microbiota and provide valuable information for monitoring the health and nutritional needs of this endangered primate at different ages.

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Fecal microbiome profiles of neonatal dairy calves with varying severities of gastrointestinal disease

PLoS ONE ◽

10.1371/journal.pone.0262317 ◽

2022 ◽

Vol 17 (1) ◽

pp. e0262317

Author(s):

Giovana S. Slanzon ◽

Benjamin J. Ridenhour ◽

Dale A. Moore ◽

William M. Sischo ◽

Lindsay M. Parrish ◽

...

Keyword(s):

Health Status ◽

Relative Abundance ◽

Gastrointestinal Disease ◽

Bifidobacterium Longum ◽

Dairy Calves ◽

Rrna Gene ◽

Probiotic Properties ◽

Fecal Samples ◽

Fecal Microbiome ◽

Microbiome Composition

Gastrointestinal disease (GI) is the most common illness in pre-weaned dairy calves. Studies have associated the fecal microbiome composition with health status, but it remains unclear how the microbiome changes across different levels of GI disease and breeds. Our objective was to associate the clinical symptoms of GI disease with the fecal microbiome. Fecal samples were collected from calves (n = 167) of different breeds (Holstein, Jersey, Jersey-cross and beef-cross) from 4–21 d of age. Daily clinical evaluations assessed health status. Calves with loose or watery feces were diagnosed with diarrhea and classified as bright-sick (BS) or depressed-sick (DS) according to behavior. Calves with normal or semiformed feces and no clinical illness were classified as healthy (H). One hundred and three fecal samples were obtained from consistently healthy calves and 64 samples were from calves with diarrhea (n = 39 BS; n = 25 DS). The V3-V4 region of 16S rRNA gene was sequenced and analyzed. Differences were identified by a linear-mixed effects model with a negative binomial error. DS and Jersey calves had a higher relative abundance of Streptococcus gallolyticus relative to H Holstein calves. In addition, DS calves had a lower relative abundance of Bifidobacterium longum and an enrichment of Escherichia coli. Species of the genus Lactobacillus, such as an unclassified Lactobacillus, Lactobacillus reuteri, and Lactobacillus salivarius were enriched in calves with GI disease. Moreover, we created a model to predict GI disease based on the fecal microbiome composition. The presence of Eggerthella lenta, Bifidobacterium longum, and Collinsella aerofaciens were associated with a healthy clinical outcome. Although lactobacilli are often associated with beneficial probiotic properties, the presence of E. coli and Lactobacillus species had the highest coefficients positively associated with GI disease prediction. Our results indicate that there are differences in the fecal microbiome of calves associated with GI disease severity and breed specificities.

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Cecal versus fecal microbiota in Ossabaw swine and implications for obesity

Physiological Genomics ◽

10.1152/physiolgenomics.00110.2017 ◽

2018 ◽

Vol 50 (5) ◽

pp. 355-368 ◽

Cited By ~ 9

Author(s):

Matthew R. Panasevich ◽

Umesh D. Wankhade ◽

Sree V. Chintapalli ◽

Kartik Shankar ◽

R. Scott Rector

Keyword(s):

Control Diet ◽

Critical Role ◽

Alpha Diversity ◽

Fecal Microbiota ◽

Rrna Gene ◽

Metabolic Capacity ◽

Fecal Samples ◽

Fecal Microbiome ◽

The Metabolic Syndrome ◽

High Fructose

The gut microbiome plays a critical role in the onset and progression of obesity and the metabolic syndrome. However, it is not well documented whether the cecal vs. the fecal microbiome is more relevant when assessing their contributions to these diseases. Here, we amplified the V4 region of the 16S rRNA gene from cecal and fecal samples of female Ossabaw swine fed a low-fat control diet (10.5% fat, n = 4) or Western diet (43.0% fat, 17.8% high fructose corn syrup, 2% cholesterol; n = 3) for 36 wk. Obesity significantly lowered alpha-diversity ( P < 0.05), and there was clear separation in beta-diversity between lean and obese pigs, as well as between cecal and fecal samples ( P < 0.05). Obesity dramatically increased ( P < 0.05) the Firmicutes:Bacteroidetes ratio in fecal samples, and Actinobacteria was higher ( P < 0.05) in fecal vs. cecal samples in obese pigs. Cyanobacteria, Proteobacteria, and Fusobacteria were increased ( P < 0.05), while Spirochaetes, Tenericutes, and Verrucomicrobia were decreased ( P < 0.05) in obese vs. lean pigs. Prevotellaceae was reduced ( P < 0.05) in obese fecal vs. cecal samples. Moreover, cecal samples in obese had greater ( P < 0.05) predicted metabolic capacity for glycan biosynthesis and metabolism and LPS biosynthesis compared with fecal. Obese pigs also had greater ( P < 0.05) capacity for carbohydrate metabolism, which was driven by obese fecal rather than cecal samples and was opposite in lean pigs ( P < 0.05). The observed differences in pro-inflammatory microbiota and their metabolic capacity in cecal vs. fecal samples of obese pigs provide new insight into evaluating the microbiome in the pathogenesis of obesity and metabolic disease.

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A multi-view model for relative and absolute microbial abundances

10.1101/761486 ◽

2019 ◽

Cited By ~ 4

Author(s):

Brian D. Williamson ◽

James P. Hughes ◽

Amy D. Willis

Keyword(s):

Relative Abundance ◽

Estimation Error ◽

Bacterial Species ◽

Critical Role ◽

Sensitivity Study ◽

Universal Primers ◽

Rrna Gene ◽

Absolute Abundance ◽

Abundance Data ◽

The Absolute

AbstractThe absolute abundance of bacterial taxa in human host-associated environments play a critical role in reproductive and gastrointestinal health. However, obtaining the absolute abundance of many bacterial species is typically prohibitively expensive. In contrast, relative abundance data for many species is comparatively cheap and easy to collect (e.g., with universal primers for the 16S rRNA gene). In this paper, we propose a method to jointly model relative abundance data for many taxa and absolute abundance data for a subset of taxa. Our method provides point and interval estimates for the absolute abundance of all taxa. Crucially, our proposal accounts for differences in the efficiency of taxon detection in the relative and absolute abundance data. We show that modeling taxon-specific efficiencies substantially reduces the estimation error for absolute abundance, and controls the coverage of interval estimators. We demonstrate the performance of our proposed method via a simulation study, a sensitivity study where we jackknife the taxa with observed absolute abundances, and a study of women with bacterial vaginosis.

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