Relative Abundance Of Operational Taxonomic Units Research Articles

Wound healing of experimental equine skin wounds and concurrent microbiota in wound dressings following topical propylene glycol gel treatment.

Topical wound treatments rely on carrier formulations with little to no biological impact. The potential for a common vehicle, a propylene glycol (PG) gel, to affect wound healing measures including microbiota is not known. Microbiome characterization, based on next generation sequencing methods is typically performed on tissue or directly obtained wound fluid samples. The utility for primary wound dressings to characterize equine wound microbiota in the context of topical treatments is currently unknown. This investigation reports the topical effect of an 80% PG based gel on wound healing and microbiota in wound dressings. Experiments were performed in six mature horses utilizing a surgical, distal limb wound model, histology of sequential wound biopsies, photographic wound measurements and microbiota profiling via 16s rRNA sequencing of wound dressing samples. Experimental wounds were surveyed for 42 days and either treated (Day 7, 14, 21 and 28; at 0.03 ml/cm2) or unexposed to the PG gel. Wound surface area, relative and absolute microbial abundances, diversity indices and histologic parameters were analyzed in the context of the experimental group (treatment; control) using qualitative or quantitative methods depending on data characteristics. Compared to controls, treatment slowed the wound healing rate (17.17 ± 4.27 vs. 18.56 ± 6.3 mm2/day), delayed the temporal decline of polymorphonucleated cells in wound beds and operational taxonomic units (OTU) in wound dressings and lowered alpha-diversity indices for microbiota in primary wound dressing. Relative abundances of OTUs were in line with those previously reported for equine wounds. Clinical outcomes 42 days post wounding were considered similar irrespective of PG gel exposure. Results highlight the potential for vehicle exposure to alter relevant wound outcome measures, imposing the need for stringent experimental control measures. Primary wound dressings may represent an alternate sample source for characterization of the wound microbiome alleviating the need for additional interventions. Further studies are warranted to contrast the microbiome in wound dressings against that present on wound surfaces to conclude on the validity of this approach.

Microbiomic profiles of bile in patients with benign and malignant pancreaticobiliary disease.

The prognostic and pathophysiologic significance of the biliary microbiota in pancreaticobiliary malignancies is little understood. Our goal was to find malignancy-related microbiomic fingerprints in bile samples taken from patients with benign and malignant pancreaticobiliary diseases. Bile specimens were collected from consenting patients during routine endoscopic retrograde cholangiopancreatography. We used PowerViral RNA/DNA Isolation kit to extract DNA from bile specimens. The Illumina 16S Metagenomic Sequencing Library Preparation guide was used to amplify the bacterial 16S rRNA gene and create libraries. QIIME (Quantitative Insights Into Microbial Ecology), Bioconductor phyloseq, microbiomeSeq, and mixMC packages were used for post-sequencing analysis. Of 46 enrolled patients, 32 patients had pancreatic cancers, 6 had cholangiocarcinoma and 1 had gallbladder cancer. Rest of the patients had benign diseases including gallstones, and acute and chronic pancreatitis. We used multivariate approach in mixMC to classify Operational Taxonomic Units (OTUs). Doing this, we found a predominance of genera Dickeya (p = 0.00008), [Eubacterium] hallii group (p = 0.0004), Bacteroides (p = 0.0006), Faecalibacterium (p = 0.006), Escherichia-Shigella (p = 0.008), and Ruminococcus 1 (p = 0.008) in bile samples from pancreaticobiliary cancers as compared to benign diseases. Additionally, bile samples from patients with pancreatic cancer exhibited a predominance of genus Rothia (p = 0.008) as compared to those with cholangiocarcinoma, whereas bile samples from patients with cholangiocarcinoma exhibited a predominance of genera Akkermansia (p = 0.031) and Achromobacter (p = 0.031) as compared to those with pancreatic cancers. Both benign and malignant pancreaticobiliary diseases have distinct microbiomic fingerprints. The relative abundance of OTUs in bile samples varies between patients with benign and malignant pancreaticobiliary diseases, as well as between cholangiocarcinoma and pancreatic cancer. Our data suggest that either these OTUs play a role in carcinogenesis or that benign disease-specific microenvironmental changes differ from cancer-specific microenvironmental changes, resulting to a clear separation of OTU clusters. We need more research to confirm and expand on our findings.

Effects of antibiotics on the development and intestinal bacterial diversity of black soldier fly larvae

The effects of four antibiotics (metronidazole (M) levofloxacin (L), sodium ampicillin (A), and streptomycin sulphate (S)) and their pair-wise combinations at three doses on the development and intestinal bacterial diversity of the black soldier fly (BSF;Hermetia illucens) larvae were studied. At a low dose M and L were able to inhibit larval growth. At a high dose, all antibiotics were shown to inhibit larval growth. However, the pair-wise combinational use of the antibiotics did not effectively enhance the inhibitory effect. The gut bacterial diversity of the normal control (NC) was significantly higher than the antibiotic-treated groups with 737 operational taxonomic units (OTUs) from the larval guts of NC, compared to 305 and 227 from ML and AS. The number of anaerobic bacteria in ML was significantly lower than in NC and AS, with the relative abundance of OTUs from larval guts of ML being only about 0.01, compared to 0.4 for NC and 0.15 for AS. These results indicated that antibiotics at the experimental concentration did not affect the palatability of food for insects, but they would affect the diversity of food and intestinal microorganisms of BSF larvae, and the inhibitory effect of antibiotics on growth and development of BSF larvae displayed in this study was a complex effect.

Microbiomic profiles of bile in patients with benign and malignant pancreaticobiliary disease.

417 Background: In pancreaticobiliary (PB) cancers, there is a paucity of data on predictive and pathophysiologic role of the biliary microbiome. We analyzed bile collected from patients with benign and malignant PB diseases to identify microbiomic signatures associated with malignancy. Methods: We collected bile samples from consenting patients during routine endoscopic retrograde cholangiopancreatography at the Cleveland Clinic, approved by the Institutional Review Board. DNA was extracted from bile specimens using PowerViral RNA/DNA Isolation kit. Bacterial 16S rRNA gene amplification and library construction were performed according to the 16S Metagenomic Sequencing Library Preparation guide from Illumina. Post-sequencing analysis was done using QIIME (Quantitative Insights Into Microbial Ecology), Bioconductor phyloseq, microbiomeSeq and mixMC packages. Results: Of 46 enrolled patients, 32 had PB cancers including pancreatic (N = 25), cholangiocarcinoma (N = 6), and gallbladder (N = 1). The rest (N = 14) had benign PB diseases including acute and chronic pancreatitis, and gallstones. Using multivariate approach in mixMC to classify Operational Taxonomic Units (OTUs), we found a predominance of genera Dicekeya (p = 0.0002), [ Eubacterium] hallii group (p = 0.0007) , Bacteroides (p = 0.00099) , Faecalibacterium (p = 0.007) , Facklamia (p = 0.013) , Peptococcus (p = 0.013) , Bergeyella (p = 0.0024) , Lachnospira (p = 0.026) , and Lactobacillus (p = 0.025) in bile samples from PB cancers as compared to benign PB diseases. Furthermore, bile samples from patients with pancreatic cancer showed an increased abundance of genera Enterobacter, Parabacteroides, Atopobium, Alloprevotella, Prevotella 7, Acinetobacter, Bergeyella, Clostridium sensu stricto, Lactobacillus, and Rothia; and a decreased abundance of genera Tannerella, Peptococcus, Colinsella, Capnocytophaga, Achromobacter, Ruminococcus 2, Bacteroides, Alistipes, Barnesiella,, Lachnoclostridium, Lautropia, Akkermansia, and Christensenellaceae R-7 group as compared to bile samples from patients with cholangiocarcinoma. Conclusions: Distinct microbiome signatures are associated with benign and malignant PB diseases. There is a difference in the relative abundance of OTUs in bile samples between patients with benign PB diseases vs PB cancers, and between pancreatic cancer vs cholangiocarcinoma. Our findings raise the possibility that either these OTUs have a role in carcinogenesis, or that changes in the microenvironment of benign PB diseases differ from PB cancers leading to distinct separation of the OTU clusters. Further studies to explore and validate our findings are needed.

Short-term responses to ocean acidification: effects on relative abundance of eukaryotic plankton from the tropical Timor Sea

Anthropogenic carbon dioxide (CO2) emissions drive climate change and pose one of the major challenges of our century. The effects of increased CO2in the form of ocean acidification (OA) on the communities of marine planktonic eukaryotes in tropical regions such as the Timor Sea are barely understood. Here, we show the effects of high CO2(mean ± SDpCO2= 1823 ± 161 μatm and pHT= 7.46 ± 0.05) versusin situCO2(504 ± 42 µatm, 7.95 ± 0.04) seawater on the community composition of marine planktonic eukaryotes after 3 and 48 h of treatment exposure in a shipboard microcosm experiment. Illumina sequencing of the V9 hypervariable region of 18S rRNA (gene) was used to study the eukaryotic community composition. Increased CO2significantly suppressed the relative abundances of different eukaryotic operational taxonomic units (OTUs), including important primary producers, although the chlorophyllaconcentration remained constant. OA effects on eukaryotes were consistent between total (DNA-based) and active (cDNA-based) taxa after 48 h, e.g. for the diatomsTrieres chinensisandStephanopyxis turris.Effects of OA on the relative abundances of OTUs were often species- or even ecotype-specific, and the incubation selectively allowed for detection of the OA-sensitive OTUs that benefitted the most from incubation in a closed bottle, as containment effects on the community structure were evident after 48 h. Many OTUs were adversely affected by sudden decreases of seawater pH, suggesting high sensitivity to OA at the base of the tropical marine biodiversity and difficult-to-predict outcomes for food-web functioning in the future ocean.

Revealing Cryptic Changes of Cyanobacterial Community Structure in Two Eutrophic Lakes Using eDNA Sequencing.

Harmful cyanobacterial blooms pose a risk to human health worldwide. To enhance understanding on the bloom-forming mechanism, the spatiotemporal changes in cyanobacterial diversity and composition in two eutrophic lakes (Erhai Lake and Lushui Reservoir) of China were investigated from 2010 to 2011 by high-throughput sequencing of environmental DNA. For each sample, 118 to 260 cpcBA-IGS operational taxonomic units (OTUs) were obtained. Fifty-two abundant OTUs were identified, which made up 95.2% of the total sequences and were clustered into nine cyanobacterial groups. Although the cyanobacterial communities of both lakes were mainly dominated by Microcystis, Erhai Lake had a higher cyanobacterial diversity. The abundance of mixed Nostocales species was lower than that of Microcystis, whereas Phormidium and Synechococcus were opportunistically dominant. The correlation between the occurrence frequency and relative abundance of OTUs was poorly fitted by the Sloan neutral model. Deterministic processes such as phosphorus availability were shown to have significant effects on the cyanobacterial community structure in Erhai Lake. In summary, the Microcystis-dominated cyanobacterial community was mainly affected by the deterministic process. Opportunistically dominant species have the potential to replace Microcystis and form blooms in eutrophic lakes, indicating the necessity to monitor these species for drinking water safety.

DNA metabarcoding effectively quantifies diatom responses to nutrients in streams

Nutrient pollution from human activities remains a common problem facing stream ecosystems. Identifying ecological responses to phosphorus and nitrogen can inform decisions affecting the protection and management of streams and their watersheds. Diatoms are particularly useful because they are a highly diverse group of unicellular algae found in nearly all aquatic environments and are sensitive responders to increased nutrient concentrations. Here, we used DNA metabarcoding of stream diatoms as an approach to quantifying effects of total phosphorus (TP) and total nitrogen (TN). Threshold indicator taxa analysis (TITAN) identified operational taxonomic units (OTUs) that increased or decreased along TP and TN gradients along with nutrient concentrations at which assemblages had substantial changes in the occurrences and relative abundances of OTUs. Boosted regression trees showed that relative abundances of gene sequence reads for OTUs identified by TITAN as low P, high P, low N, or high N diatoms had strong relationships with nutrient concentrations, which provided support for potentially using these groups of diatoms as metrics in monitoring programs. Gradient forest analysis provided complementary information by characterizing multi‐taxa assemblage change using multiple predictors and results from random forest models for each OTU. Collectively, these analyses showed that notable changes in diatom assemblage structure and OTUs began around 20 µg TP/L, low P diatoms decreased substantially and community change points occurred from 75 to 150 µg/L, and high P diatoms became increasingly dominant from 150 to 300 µg/L. Diatoms also responded to TN with large decreases in low N diatoms occurring from 280 to 525 µg TN/L and a transition to dominance by high N diatoms from 525–850 µg/L. These diatom responses to TP and TN could be used to inform protection efforts (i.e., anti‐degradation) and management goals (i.e., nutrient reduction) in streams and watersheds. Our results add to the growing support for using diatom metabarcoding in monitoring programs.

The effect of the microalgae-bacteria microbiome on wastewater treatment and biomass production.

The use of microalgae for wastewater treatment has been proposed as a cost-effective method to produce biofuels while remediating waste streams. This study examined the microalgae biomass production rate, wastewater treatment efficiency, and prokaryotic organism microbiome associated with microalgae Chlorella sorokiniana cultivated on anaerobic digestate effluent. Final microalgae biomass concentrations from nine photobioreactors were highly variable and had values that ranged between 0.14 g/L and 0.90 g/L. Nutrient removal efficiencies for TN (total nitrogen), N-NH4 (ammonium nitrogen), and COD (chemical oxygen demand) ranged from 34% to 67%, 65% to 97%, and-60% to 14%, respectively. Analysis of individual OTUs (operational taxonomic units) from the microbial community revealed that microalgae biomass concentrations were significantly correlated with the relative abundance of OTUs in the genus Pusillimonas. Predictive metagenomic analyses identified additional correlations associated with biomass production and nutrient removal. These results suggest that the microbial community present during microalgae cultivation on wastewater can impact the performance of the system for biomass production and wastewater treatment.

Distinct Coastal Microbiome Populations Associated With Autochthonous- and Allochthonous-Like Dissolved Organic Matter.

Coastal zones are important transitional areas between the land and sea, where both terrestrial and phytoplankton supplied dissolved organic matter (DOM) are respired or transformed. As climate change is expected to increase river discharge and water temperatures, DOM from both allochthonous and autochthonous sources is projected to increase. As these transformations are largely regulated by bacteria, we analyzed microbial community structure data in relation to a 6-month long time-series dataset of DOM characteristics from Roskilde Fjord and adjacent streams, Denmark. The results showed that the microbial community composition in the outer estuary (closer to the sea) was largely associated with salinity and nutrients, while the inner estuary formed two clusters linked to either nutrients plus allochthonous DOM or autochthonous DOM characteristics. In contrast, the microbial community composition in the streams was found to be mainly associated with allochthonous DOM characteristics. A general pattern across the land-to-sea interface was that Betaproteobacteria were strongly associated with humic-like DOM [operational taxonomic units (OTUs) belonging to family Comamonadaceae], while distinct populations were instead associated with nutrients or abiotic variables such as temperature (Cyanobacteria genus Synechococcus) and salinity (Actinobacteria family Microbacteriaceae). Furthermore, there was a stark shift in the relative abundance of OTUs between stream and marine stations. This indicates that as DOM travels through the land-to-sea interface, different bacterial guilds continuously degrade it.

Biases in the metabarcoding of plant pathogens using rust fungi as a model system.

Plant pathogens such as rust fungi (Pucciniales) are of global economic and ecological importance. This means there is a critical need to reliably and cost‐effectively detect, identify, and monitor these fungi at large scales. We investigated and analyzed the causes of differences between next‐generation sequencing (NGS) metabarcoding approaches and traditional DNA cloning in the detection and quantification of recognized species of rust fungi from environmental samples. We found significant differences between observed and expected numbers of shared rust fungal operational taxonomic units (OTUs) among different methods. However, there was no significant difference in relative abundance of OTUs that all methods were capable of detecting. Differences among the methods were mainly driven by the method's ability to detect specific OTUs, likely caused by mismatches with the NGS metabarcoding primers to some Puccinia species. Furthermore, detection ability did not seem to be influenced by differences in sequence lengths among methods, the most appropriate bioinformatic pipeline used for each method, or the ability to detect rare species. Our findings are important to future metabarcoding studies, because they highlight the main sources of difference among methods, and rule out several mechanisms that could drive these differences. Furthermore, strong congruity among three fundamentally different and independent methods demonstrates the promising potential of NGS metabarcoding for tracking important taxa such as rust fungi from within larger NGS metabarcoding communities. Our results support the use of NGS metabarcoding for the large‐scale detection and quantification of rust fungi, but not for confirming the absence of species.

Changes in the Total Fecal Bacterial Population in Individual Horses Maintained on a Restricted Diet Over 6 Weeks.

Twelve mature (aged 5–16 years) horses and ponies of mixed breed and type were fed restricted (1.25% BM Dry matter) quantities of one of two fiber based diets formulated to be iso-caloric. Diet 1 comprised of 0.8% body mass (BM) of chaff based complete feed plus 0.45% BM low energy grass hay (the same hay used for both diets). Diet 2 comprised 0.1% BM of a nutrient balancer plus 1.15% BM grass hay. Fecal samples were collected at week 10 and week 16. DNA was extracted and the V1-V2 regions of 16SrDNA were 454-pyrosequenced to investigate the bacterial microbiome of the horse. The two most abundant phyla found in both diets and sampling periods were the Firmicutes and Bacteroidetes. There was a clear reduction in Bacteroidetes with a concordant increase in Firmicutes over time. There was a limited degree of stability within the bacterial community of the hindgut of horses, with 65% of bacteria retained, over a 6 week period whilst on a uniform diet. The presence of a core community defined by being present in all samples (each animal/diet combination) included in the study and being present at 0.1% relative abundance (or greater) was identified. In total 65 operational taxonomic units (OTUs) were identified that fit the definition of core making up 21–28% of the total sequences recovered. As with total population the most abundant phyla were the Bacteroidetes followed by the Firmicutes, however there was no obvious shift in phyla due to period. Indeed, when the relative abundance of OTUs was examined across diets and periods there was no significant effect of diet or period alone or in combination on the relative abundance of the core OTUs.

Modeling human enteric dysbiosis and rotavirus immunity in gnotobiotic pigs.

BackgroundRotavirus vaccines have poor efficacy in infants from low- and middle-income countries. Gut microbiota is thought to influence the immune response to oral vaccines. Thus, we developed a gnotobiotic (Gn) pig model of enteric dysbiosis to study the effects of human gut microbiota (HGM) on immune responses to rotavirus vaccination, and the effects of rotavirus challenge on the HGM by colonizing Gn pigs with healthy HGM (HHGM) or unhealthy HGM (UHGM). The UHGM was from a Nicaraguan infant with a high enteropathy score (ES) and no seroconversion following administration of oral rotavirus vaccine, while the converse was characteristic of the HHGM. Pigs were vaccinated, a subset was challenged, and immune responses and gut microbiota were evaluated.ResultsSignificantly more rotavirus-specific IFN-γ producing T cells were in the ileum, spleen, and blood of HHGM than those in UHGM pigs after three vaccine doses, suggesting HHGM induces stronger cell-mediated immunity than UHGM. There were significant correlations between multiple Operational Taxonomic Units (OTUs) and frequencies of IFN-γ producing T cells at the time of challenge. There were significant positive correlations between Collinsella and CD8+ T cells in blood and ileum, as well as CD4+ T cells in blood, whereas significant negative correlations between Clostridium and Anaerococcus, and ileal CD8+ and CD4+ T cells. Differences in alpha diversity and relative abundances of OTUs were detected between the groups both before and after rotavirus challenge.ConclusionAlterations in microbiome diversity and composition along with correlations between certain microbial taxa and T cell responses warrant further investigation into the role of the gut microbiota and certain microbial species on enteric immunity. Our results support the use of HGM transplanted Gn pigs as a model of human dysbiosis during enteric infection, and oral vaccine responses.Electronic supplementary materialThe online version of this article (doi:10.1186/s13099-016-0136-y) contains supplementary material, which is available to authorized users.

Bacterial Community Development in Experimental Gingivitis

Current knowledge of the microbial composition of dental plaque in early gingivitis is based largely on microscopy and cultural methods, which do not provide a comprehensive description of oral microbial communities. This study used 454-pyrosequencing of the V1–V3 region of 16S rRNA genes (approximately 500 bp), and bacterial culture, to characterize the composition of plaque during the transition from periodontal health to gingivitis. A total of 20 healthy volunteers abstained from oral hygiene for two weeks, allowing plaque to accumulate and gingivitis to develop. Plaque samples were analyzed at baseline, and after one and two weeks. In addition, plaque samples from 20 chronic periodontitis patients were analyzed for cross-sectional comparison to the experimental gingivitis cohort. All of the healthy volunteers developed gingivitis after two weeks. Pyrosequencing yielded a final total of 344 267 sequences after filtering, with a mean length of 354 bases, that were clustered into an average of 299 species-level Operational Taxonomic Units (OTUs) per sample. Principal coordinates analysis (PCoA) plots revealed significant shifts in the bacterial community structure of plaque as gingivitis was induced, and community diversity increased significantly after two weeks. Changes in the relative abundance of OTUs during the transition from health to gingivitis were correlated to bleeding on probing (BoP) scores and resulted in the identification of new health- and gingivitis-associated taxa. Comparison of the healthy volunteers to the periodontitis patients also confirmed the association of a number of putative periodontal pathogens with chronic periodontitis. Taxa associated with gingivitis included Fusobacterium nucleatum subsp. polymorphum, Lachnospiraceae [G-2] sp. HOT100, Lautropia sp. HOTA94, and Prevotella oulorum, whilst Rothia dentocariosa was associated with periodontal health. Further study of these taxa is warranted and may lead to new therapeutic approaches to prevent periodontal disease.

Long‐term nitrogen and phosphorus fertilization effects on N2 fixation rates and nifH gene community patterns in mangrove sediments

AbstractThe bioavailability of nutrients is important in controlling ecological processes and nitrogen cycling in oligotrophic mangrove forests, yet the variation of diazotrophic community structure and activity with nutrient availability in sediments remains largely unexplored. To investigate for the first time how nutrients in sediments affect spatial and temporal patterns of diazotrophic community structure and activity, the sedimentary environment of Twin Cays, Belize, was examined with respect to the effects of long‐term fertilization [treatments: control (Ctrl), nitrogen (N), and phosphorus (P)] on N2 fixation rates and nifH gene community structure. We found that N2 fixation rates were significantly higher at the P‐treatment, intermediate at the Ctrl‐treatment and lower in the N‐treatment (P: 4.2 ± 0.5, Crtl: 0.8 ± 0.1, N: 0.4 ± 0.1 nmol·N·g−1·h−1; P < 0.001) with spatial (Ctrl‐ and P‐treatments) and temporal (only P‐treatment) variability positively correlated with live root abundance (r2 = 0.473, P < 0.001) and concentration (r2 = 0.458, P < 0.0001). The community structure of diazotrophs showed larger spatial and temporal variability in the fertilized treatments than in the Ctrl‐treatment, with the relative abundance of OTUs (nifH operational taxonomic units) at the fertilized treatments inversely related to live root abundance. Overall, long‐term fertilization (with either N or P) affects not only nutrient levels in mangrove sediments directly, but also spatial and temporal patterns of both community structure and activity and likely plant‐microbe interactions as well. Our findings suggest that the maintenance of natural nutrient conditions in mangrove sediments is important to ensure the stability of microbial functional groups like diazotrophs.