16S rRNA Amplicon Sequencing Research Articles

Investigating the Effects of Elevation on Microbial Communities and Soil Properties at Fanjing Mountain, China

Elevation is one of the most influential factors affecting soil characteristics and microbial communities in forest ecosystems. Nevertheless, there is no consensus on how soil characteristics, soil microbials, and their relationships response to the elevation of the mountain ecosystem. We investigated the soil physicochemical characteristics, the activity of soil enzymes, and the microbial community at elevational sites from 600 to 2400 m above sea level (asl) in the western slopes of the Fanjing Mountain ecosystem, China. The soil microbial communities were determined by high throughput 16S rRNA and ITS amplicon sequencing. The results demonstrated that soil total nitrogen (TN) showed a slight decrease, whereas total phosphorus (TP) and total potassium (TK) gradually tended to increase with increasing elevation. The large macroaggregates (>2 mm) accounted for the largest proportion of the aggregate fraction (66.23%–76.13%) in the 0–10 cm soil layer with elevation. The average values of the soil electrical conductivity (EC), soil organic carbon (SOC), and cation exchange capacity (CEC) concentration in the 0–60 cm layer undulated with increasing elevation, and the highest values were observed at 1500–1800 m asl and 1800–2100 m asl, respectively. The activities of soil urease, sucrase, acid phosphatase, and catalase clearly differed (p < 0.05) with increasing elevation, and the minimum values were found at 2100–2400 m asl. Interesting, with increasing soil depth, the values of these factors tended to decrease, indicating surface aggregation. In addition, the soil microbial (bacterial and fungal) community diversity exhibited a single-peak pattern with elevation. Our results also revealed that the soil bacterial and fungal communities varied significantly at different elevation sites. The bacterial communities were dominated by the phyla Acidobacteria, Pseudomonadota, and Chloroflexi, and the phyla Basidiomycota and Ascomycota dominated the fungal communities. The Pearson and redundancy analyses revealed that the SOC, TP, four soil enzymes, and soil aggregates were significant factors influencing the soil microbial community. In conclusion, soil properties and enzyme activities jointly explained the elevational pattern of the soil microbial community in the Fanjing Mountain. The results of this study provide insights into the influence of elevation on soil characteristics, microbial communities, and their relationships in the Fanjing Mountain ecosystem.

Wastewater-associated plastispheres: A hidden habitat for microbial pathogens?

Wastewater treatment plants (WWTPs) receive wastewater from various sources. Despite wastewater treatment aiming to remove contaminants, microplastics persist. Plastic surfaces are quickly colonized by microbial biofilm ("plastispheres"). Plastisphere communities are suggested to promote the spread and survival of potential human pathogens, suggesting that the transfer of plastispheres from wastewater to the environment could pose a risk to human and environmental health. The study aimed to identify pathogens in wastewater plastispheres, specifically food-borne pathogens, in addition to characterizing the taxonomic diversity and composition of the wastewater plastispheres. Plastispheres that accumulated on polypropylene (PP), polyvinyl chloride (PVC), and high-density polyethylene propylene (HDPE) surfaces exposed to raw and treated wastewater were analyzed via cultivation methods, quantitative reverse transcription PCR (RT‒qPCR) and 16S rRNA amplicon sequencing. RT‒qPCR revealed the presence of potential foodborne pathogenic bacteria and viruses, such as Listeria monocytogenes, Escherichia coli, norovirus, and adenovirus. Viable isolates of the emerging pathogenic species Klebsiella pneumoniae and Acinetobacter spp. were identified in the plastispheres from raw and treated wastewater, indicating that potential pathogenic bacteria might survive in the plastispheres during the wastewater treatment. These findings underscore the potential of plastispheres to harbor and disseminate pathogenic species, posing challenges to water reuse initiatives. The taxonomic diversity and composition of the plastispheres, as explored through 16S rRNA amplicon sequencing, were significantly influenced by the wastewater environment and the duration of time the plastic spent in the wastewater. In contrast, the specific plastic material did not influence the bacterial composition, while the bacterial diversity was affected. Without efficient wastewater treatment and proper plastic waste management, wastewater could act as a source of transferring plastic-associated pathogens into the food chain and possibly pose a threat to human health. Continued research and innovation are essential to improve the removal of microplastics and associated pathogenic microorganisms in wastewater.

Endophytic Bacillus velezensis GsB01 controls Gleditsia sinensis wilt by secreting antifungal metabolites and modulates symbiotic microbiota within trees.

Identifying effective biological control agents against fungal pathogens and determining their mechanisms of action are important in the control of plant diseases. In this study, we isolated an endophytic bacterial strain, GsB01, from the branches of asymptomatic Gleditsia sinensis. Multi-locus sequence analysis identified the strain as Bacillus velezensis. GsB01 exhibited significant antifungal activity against Thyronectria austroamericana, the causative agent of G. sinensis wilt. Liquid chromatography-mass spectrometry identified four consistently present antimicrobial compounds in GsB01 metabolite fractions with high antifungal activity: macrolactin A, bacillaene A, surfactin, and iturin. GsB01's active metabolite fractions altered the metabolic profiles of T. austroamericana, disrupting seven pathways, including arginine biosynthesis, nucleotide metabolism, purine metabolism, and the pentose phosphate pathway. Furthermore, absolute quantitative polymerase chain reaction analysis suggested that GsB01 may increase the abundance of endophytic bacteria in G. sinensis. The 16S rRNA amplicon sequencing revealed changes in the endophytic landscape in stems and roots following GsB01 introduction, particularly with significant variation in the dominant bacterial genera within the stems. The study highlights GsB01's potential against plant wilt and suggests that its antifungal activity is achieved by secreting antifungal metabolites. The study also recorded changes in the symbiotic microbiota within trees that had been infected with a pathogenic fungus and subsequently treated with an endophytic antagonistic bacterial strain. © 2024 Society of Chemical Industry.

Seasonality influences skin bacterial community structure and anti-Bd function in two anuran species

Microorganisms on amphibian skin reduce disease susceptibility and play an important role in pathogen defense. We hypothesized that anuran skin bacterial communities would change in response to seasonal variation and host species. To test this hypothesis, we used 16S rRNA amplicon sequencing to identify cutaneous bacterial communities of two frogs from the Qinling Mountains of China, Pelophylax nigromaculatus and Nanorana quadranus. We matched the amplicon sequence variants (ASVs) of microbes exhibiting protective effects against the pathogen Batrachochytrium dendrobatidis (Bd), using a database containing over 1900 16S rRNA gene sequences from amphibian skin bacteria. The results showed that seasonal variation had a stronger effect than host species on the structure (alpha-diversity, beta-diversity, species composition and abundance, and biomarkers) and anti-Bd function of cutaneous bacterial communities. These communities were highly dynamic but varied similarly between hosts. Their structural similarities were more consistent at the phylum level, but markedly less so at finer taxonomic levels. The highest relative abundance of anti-Bd reads was observed in P. nigromaculatus during summer, but anti-Bd reads were present in both frog species during different seasons. Therefore, the protective function of cutaneous microbial communities appears to be continuous despite between-species differences in anti-Bd ASV abundance. This observation does not directly explain why Bd infections have not been recorded in the region, butprovides important insight on anuran pathogen defense mechanisms. Our findings also suggest that specific seasons can be periods of high infection risk, with major implications for research on amphibian ecology and conservation.

Fecal and Sputum Microbiota and Treatment Response in Patients with Mycobacterium abscessus Pulmonary Disease.

The microbiota is a potential source of biomarkers for clinical outcomes in chronic respiratory conditions, but its role in Mycobacterium abscessus pulmonary disease (PD) remains unexplored. We aimed to identify microbial signatures in fecal and sputum microbiotas associated with treatment response in patients with M. abscessus PD. We conducted a cohort study prospectively enrolling patients undergoing antibiotic treatment for M. abscessus PD. Fecal and sputum samples were collected before, and 2 weeks and 6 months after, treatment initiation. 16S rRNA amplicon sequencing approach was used to characterize the microbiotas. After categorizing patients as early treatment responders and non-responders based on sputum culture results at 2 weeks, we elucidated the differences in their diversity and composition of microbiotas. Among 32 patients enrolled, 27 patients (median 66 years, 85.2% women, 48.1% with subspecies abscessus) were included for microbiota analysis. Fifteen patients (55.6%) achieved negative conversion at 2 weeks, which was maintained in 14 (93.3%) after 6 months. Alpha-diversity of the fecal microbiota after 2 weeks of treatment decreased significantly in responders, but not in non-responders (P=0.029). Increased abundance of Eubacterium hallii group in baseline fecal microbiota was indicative of unresponsiveness to antibiotics, whereas an increase in Enterococcus in fecal microbiota at week 2 was linked with favorable response. Increased abundance of Burkholderia-Caballeronia-Paraburkholderia and Porphyromonas in baseline, and decreased abundance of Rothia in week 2 sputum microbiota were also associated with good treatment response. In M. abscessus PD, changes in microbial diversity and compositional signatures vary with treatment response.

SALINITY-Induced Changes in Diversity, Stability, and Functional Profiles of Microbial Communities in Different Saline Lakes in Arid Areas

Saline lakes, characterized by high salinity and limited nutrient availability, provide an ideal environment for studying extreme halophiles and their biogeochemical processes. The present study examined prokaryotic microbial communities and their ecological functions in lentic sediments (with the salinity gradient and time series) using 16S rRNA amplicon sequencing and a metagenomic approach. Our findings revealed a negative correlation between microbial diversity and salinity. The notable predominance of Archaea in high-salinity lakes signified a considerable alteration in the composition of the microbial community. The results indicate that elevated salinity promotes homogeneous selection pressures, causing substantial alterations in microbial diversity and community structure, and simultaneously hindering interactions among microorganisms. This results in a notable decrease in the complexity of microbial ecological networks, ultimately influencing the overall ecological functional responses of microbial communities such as carbon fixation, sulfur, and nitrogen metabolism. Overall, our findings reveal salinity drives a notable predominance of Archaea, selects for species adapted to extreme conditions, and decreases microbial community complexity within saline lake ecosystems.

Bacterial Diversity Along the Geothermal Gradients: Insights from the High-altitude Himalayan Hot Spring Habitats of Sikkim

Geothermal habitats present a unique opportunity to study microbial adaptation to varying temperature conditions. In such environments, distinct temperature gradients foster diverse microbial communities, each adapted to its optimal niche. However, the complex dynamics of bacterial populations in across these gradients high-altitude hot springs remain largely unexplored. We hypothesize that temperature is a primary driver of microbial diversity, and bacterial richness peaks at intermediate temperatures. To investigate this, we analysed bacterial diversity using 16S rRNA amplicon sequencing across three temperature regions: hot region of 56-65°C (hot spring), warm region of 35-37°C (path carrying hot spring water to the river), and cold region of 4-7°C (river basin). Our findings showed that Bacillota was the most abundant phylum (45.51%), followed by Pseudomonadota (32.81%) and Actinomycetota (7.2%). Bacillota and Chloroflexota flourished in the hot and warm regions, while Pseudomonadota thrived in cooler areas. Core microbiome analysis indicated that species richness was highest in the warm region, declining in both cold and hot regions. Interestingly, an anomaly was observed with Staphylococcus, which was more abundant in cases where ponds were used for bathing and recreation. In contrast, Clostridium was mostly found in cold regions, likely due to its viability in soil and ability to remain dormant as a spore-forming bacterium. The warm region showed the highest bacterial diversity, while richness decreased in both cold and hot regions. This highlights the temperature-dependent nature of microbial communities, with optimal diversity in moderate thermal conditions. The study offers new insights into microbial dynamics in high-altitude geothermal systems.

Bioactive fraction of Musa balbisiana seed mitigates D-galactose-induced brain aging via SIRT1/PGC-1α/FoxO3a activation and intestinal barrier dysfunction by modulating gut microbiota and core metabolites

Aging is an inevitable biological process, and emerging research has highlighted the potential of dietary and pharmacological interventions to decelerate the trajectory of age-related diseases and prolong the health span. This study evaluates the protective effects of Musa balbisiana seed on healthy aging using D-galactose-induced accelerated aging rats. The results suggested that the bioactive ethyl acetate fraction of Musa balbisiana seed extract (BF) exhibited protective effects against aging-induced oxidative stress by reducing oxidative DNA damage, advanced glycation end-product formation, and malondialdehyde levels while restoring antioxidant and glyoxalase enzyme activities. BF also ameliorated neurodegeneration by decreasing acetylcholinesterase enzyme activity and amyloid beta plaque formation. Histopathological analysis demonstrated the protective effects of BF against brain aging, liver disruption, renal damage, and intestinal barrier dysfunction. BF further restored intestinal permeability by upregulating the tight junctions (zonula occludens 1 and 2, claudin 1,2,3 and 4, and occludin) and mucin (mucin 2 and mucin 5ac) gene expression while downregulating the expression of inflammatory cytokines (IL-1β, IL-6, and TNF-α). BF significantly induced the phosphorylation of FoxO3a proteins and upregulated the gene expression of SIRT1, PGC-1α, and TFAM in the hippocampus. Next-generation sequencing (NGS) of 16s rRNA amplicons of fecal metagenomics DNA and metabolites profiling showed that BF intervention restructured the gut microbiota and altered core metabolites related to cholesterol metabolism. Overall, our findings demonstrated the multifaceted protective effects of Musa balbisiana seed against D-galactose-induced aging.

Gut microbiota and antibiotic resistance genes in endangered migratory Scaly-sided merganser (Mergus squamatus) in northeast China

Studying the diversity of gut microbiota and antibiotic resistance genes (ARGs) in endangered migratory birds, as well as their responses to different environments, can reflect the health status of migratory birds and the ecological risk of ARG transmission. The globally endangered Scaly-sided merganser (Mergus squamatus) is a typical migratory bird, yet the diversity of its intestinal microbiota and ARGs has not been reported. In this study, fecal samples were collected from two major riparian foraging areas utilized by M. squamatus in the Changbai Mountains of China: the Lushui River and the Manjiang River. The gut microbiota and ARG composition were characterized using 16S rRNA amplicon and metagenomic sequencing. The results indicated that the relative abundance of gut microbiota in the two habitats differed significantly, but the core flora was similar. Moreover, the relative abundance of pathogenic bacteria was high, suggesting that M. squamatus faces a high risk of disease. A total of 382 ARGs were identified, which were primarily resistant to multidrug, tetracycline, macrolides-lincosamides-streptogramins, and mupirocin. MsbA, abcA, rpoB2, and tetT were the most abundant shared ARGs between the two sites. The findings demonstrate that habitat influences the diversity of gut microbiota and ARGs in the endangered migratory M. squamatus. This species could serve as a reservoir for pathogenic bacteria and ARGs. Therefore, monitoring the diversity of gut microbiota and ARGs in M. squamatus across different habitats is essential for a comprehensive assessment of its health status and the ecological risk of ARG transmission in the future.

Effect of long-term ammonia deficiency on nitrification performance and microbial dynamics in downflow sponge biofilm reactor

The downflow sponge biofilm (DSB) reactor has a promising nitrification capacity for ammonium (NH4+-N)-polluted raw water treatment. However, the highly fluctuating NH4+-N levels in raw water may pose a challenge to a stable nitrification process. It is unclear how the DSB reactor contends with NH4+-N deficiency. This study evaluated the robustness of a DSB reactor under long-term NH4+-N abundant and NH4+-N deficient conditions. Four NH4+-N deficient periods were investigated, comprising two 14-day partial NH4+-N deficient phases, one 56-day phase involving both partial and complete NH4+-N deficient, and one 90-day complete NH4+-N deficient phase. Repeated 14-day partial NH4+-N deficient phases had no impact on nitrification performance. However, shifting from partial to complete NH4+-N deficient condition resulted in a delay in NOx−-N production when NH4+-N was replenished. Following an additional 90 days of NH4+-N deficient phase, NH4+-N removal performance recovered faster than nitrite (NO2−-N) removal performance, which took 5 and 29 days, respectively. Furthermore, long-term NH4+-N deficiency resulted in a nominal loss of biomass in the DSB reactor. These findings suggested that the nitrification capacity of the DSB reactor is robust when subjected to long-term NH4+-N deficient conditions. The 16S rRNA amplicon sequencing results revealed that the growth of Nitrosomonas was suppressed, but Candidatus Nitrosotenuis predominated in NH4+-N deficient conditions, while Nitrospira persisted throughout the study. The observed persistence of Nitrospira may be attributed to their diverse metabolic capabilities, which include mixotrophy and complete ammonia oxidation metabolisms. Collectively, the co-existence of Ca. Nitrosotenius, Nitrosomonas, and Nitrospira contributed to stable NH4+-N removal performance in the robust DSB reactor. The DSB reactor shows promise for use in raw water treatment with highly fluctuating NH4+-N availability.

Significant role of symbiotic bacteria in the blood digestion and reproduction of Dermanyssus gallinae mites

Abstract Endosymbiotic bacteria exert profound effects on the fitness of their arthropod hosts. Dermanyssus gallinae, the poultry red mite, is a blood-feeding ectoparasite that feeds solely on the blood of its avian hosts. A relatively comprehensive understanding of its microbial community structures across developmental stages based on 16S rRNA sequencing is available. Yet, their functional integration within the host's physiology remains elusive. This study endeavors to illuminate the role of symbiotic bacteria in the biology of D. gallinae mites. 16S rRNA amplicon sequencing and fluorescence in situ hybridization revealed a prominent midgut-confinement bacterial microbiota with considerable diversity, out of which Kocuria and Bartonella A acted as the predominant bacterial genera inhabiting D. gallinae. The relative abundance of Bartonella A exhibited rapid increments subsequent to blood-sucking, reflecting the bacteria's adaptation to grow in blood or indicating its pivotal role in the mites' post-engorgement life activities. Some of the isolated bacterial strains from D. gallinae display hemolytic activity on blood agar, which could potentially contribute to the blood digestion process in mites. To corroborate this in vivo, antibiotic-mediated clearance was exploited to generate dysbiosed cohorts of D. gallinae mites, lacking some of the key bacterial species. Phenotypic scoring of dysbiosed D. gallinae mites revealed that this cohort resulted in delayed blood digestion and diminished reproductive capacity. Whole genome sequencing revealed that Bartonella A, identified as a new species within the genus Bartonella, displays the characteristics of an obligate symbiont. These findings underscore the significance of microbiota in poultry red mites and suggest microbiota-targeted strategies for controlling mite populations in poultry farms.

Fecal Microbiome Composition Correlates with Pathologic Complete Response in Patients with Operable Esophageal Cancer Treated with Combined Chemoradiotherapy and Immunotherapy

Background: Preclinical and clinical data indicate that chemoradiotherapy (CRT) in combination with checkpoint inhibitors may prime an anti-tumor immunological response in esophageal cancer. However, responses to neoadjuvant therapy can vary widely and the key biomarkers to determine response remain poorly understood. The fecal microbiome is a novel and potentially modifiable biomarker of immunotherapy response, and both fecal and tumor microbes have been found to associate with outcomes in esophageal cancer. Methods: Fecal and tumor samples were collected from patients with stage II–III resectable esophageal or gastroesophageal junction carcinoma treated with neoadjuvant immune checkpoint inhibitors (ICIs) plus CRT prior to surgical resection. Microbiome profiles were analyzed by 16S rRNA amplicon sequencing and taxonomic data were integrated with fecal metabolite analysis to assess microbial function. Results: The fecal microbiome of patients with pathological complete response (PCR) grouped in distinct clusters compared to patients with residual viable tumor (RVT) by Bray–Curtis diversity metric. Integrated taxonomic and metabolomic analysis of fecal samples identified a sphingolipid and primary bile acid as enriched in the PCR, the levels of which correlated with several bacterial species: Roseburis inulinivorans, Ruminococcus callidus, and Fusicantenibacter saccharivorans. Analysis of the tumor microbiome profiles identified several bacterial genera previously associated with esophageal tumors, including Streptococcus and Veillonella. Conclusions: These results further characterize the fecal and tumor microbiome of patients with operable esophageal cancer and identify specific microbes and metabolites that may help elucidate how microbes contribute to tumor response with neoadjuvant CRT combined with ICI.

Long-Term Outdoor Cultivation of Nannochloropsis in California, Hawaii, and New Mexico

The project “Optimizing Selection Pressures and Pest Management to Maximize Cultivation Yield” (OSPREY, award #DE-EE08902) was undertaken to enhance the annual productivity, stability, and quality of algal production strains for biofuels and bioproducts. The foundation of this project was the year-round cultivation of a Nannochloropsis strain across three outdoor systems in California, Hawaii, and New Mexico. We aimed to leverage environmental selection pressures to drive strain improvement and use metagenomic techniques to inform pest management tools. The resulting dataset includes environmental and biological parameters from these cultivation campaigns, captured in a single CSV file. This dataset aims to serve a wide range of end users, from biologists to algal farmers, addressing the scarcity of publicly available data on algae cultivation. Further data releases will include 16S rRNA amplicon sequencing and shotgun sequencing datasets.

Effect of the 16S rRNA Gene Hypervariable Region on the Microbiome Taxonomic Profile and Diversity in the Endangered Fish Totoaba macdonaldi

Understanding the intricate dynamics of fish microbiota through 16S rRNA amplicon sequencing is pivotal for ecological insights and effective disease management. However, this approach faces challenges including the co-amplification of host mitochondrial sequences and the variability in bacterial composition influenced by the selected 16S rRNA gene regions. To overcome these limitations, we conducted a comprehensive investigation to identify the most suitable 16S rRNA region for bacterial microbial analysis in endangered fish Totoaba macdonaldi, an endemic species of significant ecological and economic importance in Mexico. Targeting four distinct hypervariable regions (V1–V2, V2–V3, V3–V4, and V5–V7) of the 16S rRNA gene, we determined the microbial composition within the distal intestine. A total of 40 microbiomes were sequenced. Our findings underscore the critical impact of region selection on the accuracy of microbiota analysis. The V3–V4 region detected the highest number of bacterial taxa and exhibited significantly higher alpha diversity indices, demonstrating the highest taxonomic resolution. This study emphasizes the necessity of meticulous 16S rRNA region selection for fish microbiota analysis, particularly in native species of ecological and economic significance such as the endangered T. macdonaldi, where information is limited. Such optimization enhances the reliability and applicability of microbiota studies in fisheries management and conservation efforts.

Dynamics of the gut microbiome in individuals at risk of rheumatoid arthritis: a cross-sectional and longitudinal observational study

ObjectivesThis work aimed to resolve the conflicting reports on Prevotellaceae abundance in the development of rheumatoid arthritis (RA) and to observe structural, functional and temporal changes in the gut microbiome...

The choice of 16S rRNA gene sequence analysis impacted characterization of highly variable surface microbiota in dairy processing environments.

Accurate knowledge of the microbiota collected from surfaces in food processing environments is important for food quality and safety. This study assessed discrepancies in taxonomic composition and alpha and beta diversity values generated from eight different bioinformatic workflows for the analysis of 16S rRNA gene sequences extracted from the microbiota collected from surfaces in dairy processing environments. We found that the microbiota collected from environmental surfaces varied widely in density (0-9.09 log10 CFU/cm2) and Shannon alpha diversity (0.01-3.40). Consequently, depending on the sequence analysis method used, characterization of low-abundance genera (i.e., below 1% relative abundance) and the number of genera identified (114-173 genera) varied considerably. Some low-abundance genera, including Listeria, varied between the amplicon sequence variant (ASV) and operational taxonomic unit (OTU) methods. Centered log-ratio transformation inflated alpha and beta diversity values compared to rarefaction. Furthermore, the ASV method also inflated alpha and beta diversity values compared to the OTU method (P < 0.05). Therefore, for sparse, uneven, low-density data sets, the OTU method and rarefaction are better for taxonomic and ecological characterization of surface microbiota.IMPORTANCECulture-dependent environmental monitoring programs are used by the food industry to identify foodborne pathogens and spoilage biota on surfaces in food processing environments. The use of culture-independent 16S rRNA amplicon sequencing to characterize this surface microbiota has been proposed as a tool to enhance environmental monitoring. However, there is no consensus on the most suitable bioinformatic analyses to accurately capture the diverse levels and types of bacteria on surfaces in food processing environments. Here, we quantify the impact of different bioinformatic analyses on the results and interpretation of 16S rRNA amplicon sequences collected from three cultured dairy facilities in New York State. This study provides guidance for the selection of appropriate 16S rRNA analysis procedures for studying environmental microbiota in dairy processing environments.

Oral, Vaginal, and Stool Microbial Signatures in Patients With Endometriosis as Potential Diagnostic Non-Invasive Biomarkers: A Prospective Cohort Study.

To identify a microbial signature for endometriosis for use as a diagnostic non-invasive biomarker. Prospective cohort pilot study. Nepean Hospital and UNSW Microbiome Research Centre, Australia. Sixty-four age- and sex-matched subjects (n = 19 healthy control (HC); n = 24 non-endometriosis (N-ENDO) and n = 21 confirmed endometriosis (ENDO)). All study participants, besides healthy controls, underwent laparoscopic surgical assessment for endometriosis, and histology was performed on excised lesions. Oral, stool and, vaginal samples were self-collected at a single time point for healthy controls, and preoperatively for patients undergoing laparoscopy. Samples underwent 16S rRNA amplicon sequencing, followed by bioinformatics analysis. Compositional differences between cohorts as identified by diversity analyses, and differentially abundant microbial taxa, as identified by LEfSE analysis. The composition of the oral (adjusted p = 0.003), and stool (adjusted p = 0.042) microbiota is different between the three cohorts. Differentially abundant taxa are present within each cohort as identified by LEfSE analysis. Particularly, Fusobacterium was enriched in the oral samples of patients with moderate/severe endometriosis. Taxonomic and compositional differences were found between the microbiota in the mouth, gut and, vagina of patients with and without endometriosis and healthy controls. Fusobacterium was enriched in patients with moderate/severe endometriosis. Fusobacterium is noted as a key pathogen in periodontal disease, a common comorbidity in endometriosis. These findings suggest a role for the oral, stool and, vaginal microbiome in endometriosis, and present potential for microbial-based treatments and the design of a diagnostic swab.

Selection and application of methanol-utilizing bacteria from tomato leaves for biocontrol of gray mold.

Gray mold, caused by Botrytis cinerea, is a significant threat to tomato production. Traditional chemical control methods have become increasingly ineffective because of the development of resistance. This study aimed to isolate methanol-utilizing bacteria from tomato leaves and evaluate their biocontrol potential against gray mold. To obtain bacterial suspensions, tomato leaf samples were collected and washed. We analyzed the microbial communities of these samples using 16S rRNA amplicon sequencing and identified several methylotrophic strains. Among these, 405 isolated strains were cultivated on a solid low-nutrient inorganic salt medium containing methanol, and 7 strains exhibiting considerable antifungal activity against B. cinerea were identified. Greenhouse tests revealed that two strains-SY163 and SY183-significantly reduced the severity of gray mold on tomato leaves. Disease index scores and the area under the disease progress curve values confirmed the efficacy of these strains as biocontrol agents. Statistical analysis indicated the effectiveness of pre- and co-application of these strains with B. cinerea. Phylogenetic analysis identified Serratia rubidaea as the inhibitory strain. The biocontrol activity is likely mediated through the production of antifungal compounds and suppression of B. cinerea sporulation. This study provides the basis for developing a technology of gray mold suppression by controlling the abundance of S. rubidaea in plant microbial communities.

Asaia spp. accelerate development of the yellow fever mosquito, Aedes aegypti, via interactions with the vertically transmitted larval microbiome.

A wide range of vector control programmes rely on the efficient production and release of male mosquito. Asaia bacteria are described as potential symbionts of several mosquito species but their relationship with Aedes aegypti has never been rigorously tested. Here, we aimed to quantify the benefits of three Asaia species on host development in Ae. aegypti, and the ability of these bacteria to form a stable symbiotic association with growing larvae. In order to disentangle direct and indirect effects of Asaia inoculation on host development, experiments used insects with an intact microbiome and those reared in near-aseptic conditions, while we characterized bacterial communities and Asaia densities with culture dependent and independent methods (16S rRNA amplicon sequencing). Neonate larvae were inoculated with Asaia spp. for 24 h, or left as uninoculated controls, all were reared on sterile food. Aseptic larvae were produced by surface sterilization of eggs. Although all Asaia were transient members of the gut community, two species accelerated larval development relative to controls. The two mutualistic species had lasting impacts on the larval microbiome, largely by altering the relative abundance of dominant bacteria, namely Klebsiella and Pseudomonas. Axenic larvae were dominated by Asaia when inoculated with this species but showed slower development than conventionally reared insects, indicating that Asaia alone could not restore normal development. Our results reveal Asaia as a poor mutualist for Ae. aegypti, but with a species-specific positive effect on improving host performance mediated by interactions with other bacteria.

Fecal microbiome analysis uncovers hidden stress effects of low stocking density on rainbow trout

BackgroundRecirculating aquaculture systems can cause chronic stress in fish when stocking density is too high. However, this study tested whether low stocking density can cause fish stress. Adult rainbow trout, with an average weight of 1.517 kg (± 0.39), were subjected to low (12 kg/m3 ± 0.94) and moderate (43 kg/m3 ± 2.03) stocking densities for 24 days in a recirculating system maintained at 15 °C. At the end of the experiment, fecal microbiome analysis was carried out using a 16S rRNA amplicon sequencing. Additionally, an untargeted plasma metabolomics analysis was conducted.ResultsThe moderate stocking density group harboured greater numbers of commensals, such as C. somerae, R. lituseburensis, and L. plantarum. In contrast, detrimental species such as S. putrifacens and P. putida were abundant in the low-stocking density fish. Functional microbiome profiling revealed vitamin B12 salvage and synthesis in moderate stocking densities, which may support intestinal tight junction function. Additionally, vitamin B1 biosynthesis pathways were more abundant in the moderate stocking density group, which may function towards oxidative energy metabolism and protect against oxidative stress. A complementary plasma metabolomics study, although done at slightly different stocking densities and duration, confirmed the presence of blood metabolic stress markers. Elevated levels of L-lactic acid and L-Norvaline, L-Valine, and L-glutamine, indicate low stocking density fish were under stress. Furthermore, a P4HA2 stress gene biomarker confirmed the occurrence of stress in low-density fish. This study suggests that low stocking density can induce stress in fish. Moreover, moderate stocking density leads to a distinct and beneficial fecal microbiome profile.ConclusionOur study highlights the potential benefits of optimizing the stocking density of fish in recirculating aquaculture systems. This can improve fish health and welfare, promoting a more resilient fecal microbiome.Graphical abstract