16S rRNA Gene-based Amplicon Sequencing Research Articles

The effect of urbanization on planktonic and biofilm bacterial communities in different water bodies of the Danube River in Hungary

Freshwaters play an essential role in providing ecosystem services worldwide, however, the water quality of different water bodies is strongly influenced by human activities such as urbanization, industry and agriculture. In this study, water and biofilm samples were collected from the main channel of the Danube River upstream and downstream of a metropolitan, from a regulated side arm within an urbanized area, and from two differently separated oxbow lakes located in nature conservation areas. The taxonomic diversity of bacterial communities was revealed by 16S rRNA gene-based amplicon sequencing using Illumina MiSeq platform. The results showed that all samples were dominated by phyla Pseudomonadota, Actinobacteriota and Bacteroidota. The bacterial community structures, however, clearly differentiated according to planktonic and epilithic or epiphytic habitats, as well as by riverine body types (main channel, side arm, oxbow lakes). The taxonomic diversity of biofilm communities was higher than that of planktonic ones in all studied habitats. Human impacts were mainly reflected in the slowly changing biofilm composition compared to the planktonic ones. Genera with pollution tolerance and/or degradation potential, such as Acinetobacter, Pseudomonas and Shewanella were mainly detected in biofilm communities of the highly urbanized section of the river side arm.

Changes in the taxonomic composition of soil bacterial communities under different inter-row tillage managements in a sloping vineyard of the Balaton Uplands (Hungary).

The common grape (Vitis vinifera L.) has been cultivated for thousands of years. Nowadays, it is cultivated using a variety of tillage practices that affect the structure of the soil microbial communities and thus the health of the vine. The aim of this study was to explore and compare the effects of tillage (shallow tillage with bare soil) and no-tillage (perennial grass cover) practices on soil physical and chemical properties and soil bacterial community diversities in a small catchment. Soil samples were taken in July and October 2020 at different slope positions of two vineyards exposed to erosion. The two sampling sites were separated by the agricultural inter-row management type: tilled and no-tilled slopes. The taxonomic diversity of bacterial communities was determined using 16S rRNA gene-based amplicon sequencing method on Illumina MiSeq platform. Based on the examined soil properties, the sampling areas were separated from each other according to the positions of the upper and lower slopes and the sampling times. Both the tilled and no-tilled soil samples were dominated by sequences assigned to phyla Pseudomonadota, Acidobacteriota, Bacteroidota, Verrucomicrobiota, Actinobacteriota, and Gemmatimonadota. The results showed that tillage had no significant effect compared to the no-tilled samples in the studied area. Water runoff and seasonally changed soil physical and chemical properties affected mainly the bacterial community structures.

Soil bacterial communities affected by land-use types in a small catchment area of the Balaton Uplands (Hungary).

Changes resulting from different tillage practices can affect the structure of microbial communities, thereby altering soil ecosystems and their functioning. The aim of this study was to explore and compare the physical, chemical properties and bacterial community composition of soils from different land use types (forest, grassland, vineyard, and arable field) in a small catchment. 16S rRNA gene-based amplicon sequencing was used to reveal the taxonomic diversity of summer and autumn soil samples taken from two different slope positions. The greater the anthropogenic impact was on the type of land use, the greater the change was in soil physical and chemical parameters. All sample types were dominated by the phyla Pseudomonadota, Acidobacteriota, Actinobacteriota, Bacteroidota and Verrucomicrobiota. Differences in the relative abundance of various bacterial taxa reflected the different land use types, the seasonality, and the topography. These diversity changes were consistent with the differences in soil properties.

Soil bacteriome diversity and composition of rooftop and surface gardens in urban and peri-urban areas of Bangladesh.

Soil microbiome science, rapidly evolving, predominantly focuses on field crop soils. However, understanding garden soil microbiomes is essential for enhancing food production sustainability in garden environments. This study aimed to unveil the bacteriome diversity and composition in rooftop garden soils (RGS) and surface garden soils (SGS) across urban (Dhaka North and Dhaka South City Corporations) and peri-urban (Gazipur City Corporation) areas of Dhaka Division, Bangladesh. We analyzed 11 samples, including six RGS and five SGS samples from 11 individual gardens using 16S rRNA (V3-V4 region) gene-based amplicon sequencing. A total of 977 operational taxonomic units (OTUs), including 270 and 707 in RGS and SGS samples, respectively, were identified. The observed OTUs were represented by 21 phyla, 45 classes, 84 orders, 173 families, and 293 genera of bacteria. Alpha diversity indices revealed significantly higher bacterial diversity in SGS samples (p = 0.01), while beta diversity analyses indicated distinct bacteriome compositions between RGS and SGS samples (p = 0.028, PERMANOVA). Despite substantial taxonomic variability between sample categories, there was also a considerable presence of shared bacterial taxa. At the phylum level, Bacilliota (61.14%), Pseudomonadota (23.42%), Actinobacteria (6.33%), and Bacteroidota (3.32%) were the predominant bacterial phyla (comprising > 94.0% of the total abundances) in both types of garden soil samples. Of the identified genera, Bacillus (69.73%) and Brevibacillus (18.81%) in RGS and Bacillus (19.22%), Methylophaga (19.21%), Acinetobacter (6.27%), Corynebacterium (5.06%), Burkholderia (4.78%), Paracoccus (3.98%) and Lysobacter (2.07%) in SGS were the major bacterial genera. Importantly, we detected that 52.90% of genera were shared between RGS and SGS soil samples. Our data reveal unique and shared bacteriomes with probiotic potential in soil samples from both rooftop and surface gardens. Further studies should explore the functional roles of shared bacterial taxa in garden soils and how urban environmental factors affect microbiome composition to optimize soil health and sustainable food production.

The microbiota of ensiled forages and of bulk tank milk on dairy cattle farms in northern Sweden

Factors contributing to variations in the quality and microbiota of ensiled forages and in bulk tank microbiota in milk from cows fed different forages were investigated. Nutritional quality, fermentation parameters and hygiene quality of forage samples and corresponding bulk tank milk samples collected in 3 periods from 18 commercial farms located in northern Sweden were compared. Principal coordinates analysis revealed that the microbiota in forage and bulk milk, analyzed using 16S rRNA gene-based amplicon sequencing, were significantly different. The genera Lactobacillus, Weissella, and Leuconostoc dominated in forage samples, whereas Pseudomonas, Staphylococcus, and Streptococcus dominated in bulk milk samples. Forage quality and forage-associated microbiota were affected by ensiling method and by use of silage additive. Forages stored in bunker and tower silos (confounded with use of additive) were associated with higher levels of acetic and lactic acid and Lactobacillus. Forage ensiled as bales (confounded with no use of additive) was associated with higher DM content, water-soluble carbohydrate content, pH, yeast count, and the genera Weissella, Leuconostoc, and Enterococcus. For bulk tank milk samples, milking system was identified as the major factor affecting the microbiota and type of forage preservation had little effect. Analysis of common amplicon sequence variants (ASV) suggested that forage was not the major source of Lactobacillus found in bulk tank milk.

High-throughput 16S rRNA gene-based amplicon sequencing reveals the functional divergence of halophilic bacterial communities in the Suaeda salsa root compartments on the eastern coast of China

The rhizosphere environment of plants, which harbors halophilic bacterial communities, faces significant challenges in coping with environmental stressors, particularly saline soil properties. This study utilizes a high-throughput 16S rRNA gene-based amplicon sequencing to investigate the variations in bacterial community dynamics in rhizosphere soil (RH), root surface soil (RS), root endophytic bacteria (PE) compartments of Suaeda salsa roots, and adjoining soils (CK) across six locations along the eastern coast of China: Nantong (NT), Yancheng (YC), Dalian (DL), Tianjin (TJ), Dongying (DY), and Qingdao (QD), all characterized by chloride-type saline soil. Variations in the physicochemical properties of the RH compartment were also evaluated. The results revealed significant changes in pH, electrical conductivity, total salt content, and ion concentrations in RH samples from different locations. Notably, the NT location exhibited the highest alkalinity and nitrogen availability. The pH variations were linked to HCO3− accumulation in S. salsa roots, while salinity stress influenced soil pH through H+ discharge. Despite salinity stress, enzymatic activities such as catalase and urease were higher in soils from various locations. The diversity and richness of bacterial communities were higher in specific locations, with Proteobacteria dominating PE samples from the DL location. Additionally, Vibrio and Marinobacter were prevalent in RH samples. Significant correlations were found between soil pH, salinity, nutrient content, and the abundance and diversity of bacterial taxa in RH samples. Bioinformatics analysis revealed the prevalence of halophilic bacteria, such as Bacillus, Halomonas, and Streptomyces, with diverse metabolic functions, including amino acid and carbohydrate metabolisms. Essential genes, such as auxin response factor (ARF) and GTPase-encoding genes, were abundant in RH samples, suggesting adaptive strategies for harsh environments. Likewise, proline/betaine transport protein genes were enriched, indicating potential bioremediation mechanisms against high salt stress. These findings provide insight into the metabolic adaptations facilitating resilience in saline ecosystems and contribute to understanding the complex interplay between soil conditions, bacterial communities, and plant adaptation.

Gestational diabetes-related gut microbiome dysbiosis is not influenced by different Asian ethnicities and dietary interventions: a pilot study.

Gut microbiome dysbiosis contributes to the pathophysiology of both gestational diabetes mellitus (GDM) and its associated adverse outcomes in the woman and offspring. Even though GDM prevalence, complications, and outcomes vary among different ethnic groups, limited information is available about the influence of ethnicity on gut microbiome dysbiosis in pregnancies complicated by GDM. This pilot prospective cohort study examined the impact of ethnicity on gut dysbiosis in GDM among three Asian ethnic groups (Chinese, Malay, Indian) living in Singapore, and investigated the potential modulatory roles of diet and lifestyle modifications on gut microbiome post-GDM diagnosis. Women with GDM (n = 53) and without GDM (n = 16) were recruited. Fecal samples were collected at 24-28- and 36-40-weeks' gestation and analyzed by targeted 16S rRNA gene-based amplicon sequencing. Permutational multivariate analysis of variance (PERMANOVA) analysis was performed to evaluate differences between groups. Differentially abundant taxa were identified by DeSeq2 based analysis. Functional prediction was performed using thephylogeneticinvestigation of communities by reconstruction of unobserved states (PICRUSt2). Among women with GDM, gut microbiome from different ethnicities harbored common microbial features. However, among those without GDM, there was contrasting microbiome composition between ethnic groups. Microbial members such as Collinsella, Blautia, Ruminococcus, Ruminococcus gnavus, Ruminococcus torques, and Eubacterium hallii groups were differentially enriched (p < 0.05) in women with GDM compared to those without. Among women with GDM, no differences in alpha- and beta- diversity were observed when comparing 24-28weeks' samples with 36-40weeks' samples, a period covering intense dietary and lifestyle modification, suggesting an inability to modulate gut microbiota through classic GDM management. Women with GDM have a distinct gut microbiome profile which harbours common features across different Asian ethnic groups, consistent with the notion that specific microbes are involved in the pathogenesis of insulin resistance, pro-inflammatory conditions, and other metabolic dysregulation known to be present in GDM.

The Vaginal Microbiome Changes During Various Fertility Treatments.

This study aimed to investigate the influence of hormonal treatment on the vaginal microbiome during fertility treatments. Bacterial vaginosis (BV) could affect fecundity, particularly in the in vitro fertilization (IVF) population, where negative effects on pregnancy outcomes have been reported. It is hypothesized that the hormone treatment during fertility treatments could influence the abundance of Lactobacilli, with negative effects on the pregnancy results. A total of 53 couples attending a fertility clinic in the Netherlands between July 2019 and August 2022 were included in this prospective cohort study. Vaginal samples were collected at start of treatment, oocyte retrieval or insemination from subjects undergoing intra uterine insemination (IUI) with mild ovarian stimulation, and IVF or intra cytoplasmatic sperm injection (ICSI) with controlled ovarian hyperstimulation. AmpliSens® Florocenosis/Bacterial vaginosis-FRT qPCR and 16S rRNA gene-based amplicon sequencing were performed on all samples. In total, 140 swabs were analyzed, with a median of two swabs per person. 33 (24%) tested qPCR BV positive. Lactobacilli percentage decreased during fertility treatments, leading to changes in the vaginal microbiome. Shannon diversity index was not significantly different. Of the total of 53 persons, nine switched from qPCR BV negative to positive during treatment. The persons switching to qPCR BV positive had already a (not significant) higher Shannon diversity index at start of treatment. If the vaginal microbiome of persons deteriorates during fertility treatments, timing of following treatments, lifestyle modifications, or a freeze all strategy could be of possible benefit.

Process conditions affect microbial diversity and activity in a haloalkaline biodesulfurization system.

Biodesulfurization (BD) systems that treat sour gas employ mixtures of haloalkaliphilic sulfur-oxidizing bacteria to convert sulfide to elemental sulfur. In the past years, these systems have seen major technical innovations that have led to changes in microbial community composition. Different studies have identified and discussed the microbial communities in both traditional and improved systems. However, these studies do not identify metabolically active community members and merely focus on members' presence/absence. Therefore, their results cannot confirm the activity and role of certain bacteria in the BD system. To investigate the active community members, we determined the microbial communities of six different runs of a pilot-scale BD system. 16S rRNA gene-based amplicon sequencing was performed using both DNA and RNA. A comparison of the DNA- and RNA-based sequencing results identified the active microbes in the BD system. Statistical analyses indicated that not all the existing microbes were actively involved in the system and that microbial communities continuously evolved during the operation. At the end of the run, strains affiliated with Alkalilimnicola ehrlichii and Thioalkalivibrio sulfidiphilus were confirmed as the most active key bacteria in the BD system. This study determined that microbial communities were shaped predominantly by the combination of hydraulic retention time (HRT) and sulfide concentration in the anoxic reactor and, to a lesser extent, by other operational parameters.IMPORTANCEHaloalkaliphilic sulfur-oxidizing bacteria are integral to biodesulfurization (BD) systems and are responsible for converting sulfide to sulfur. To understand the cause of conversions occurring in the BD systems, knowing which bacteria are present and active in the systems is essential. So far, only a few studies have investigated the BD system's microbial composition, but none have identified the active microbial community. Here, we reveal the metabolically active community, their succession, and their influence on product formation.

Solitary Bees Host More Bacteria and Fungi on Their Cuticle than Social Bees.

Bees come into contact with bacteria and fungi from flowering plants during their foraging trips. The Western honeybee (Apis mellifera) shows a pronounced hygienic behavior with social interactions, while the solitary red mason bee (Osmia bicornis) lacks a social immune system. Since both visit the same floral resources, it is intriguing to speculate that the body surface of a solitary bee should harbor a more complex microbiome than that of the social honeybee. We compared the cuticular microbiomes of A. mellifera (including three European subspecies) and O. bicornis for the first time by bacterial 16S rRNA and fungal ITS gene-based high-throughput amplicon sequencing. The cuticular microbiome of the solitary O. bicornis was significantly more complex than that of the social A. mellifera. The microbiome composition of A. mellifera subspecies was very similar. However, we counted significantly different numbers of fungi and a higher diversity in the honeybee subspecies adapted to warmer climates. Our results suggest that the cuticular microbiome of bees is strongly affected by visited plants, lifestyle and adaptation to temperature, which have important implications for the maintenance of the health of bees under conditions of global change.

Relationship between microorganisms and volatile components in each fermentation process in the kusaya gravy that plays an important role in the manufacturing of kusaya, a traditional Japanese fermented fish product.

The relationship between the microbiota and volatile components of kusaya gravy involved in the manufacturing of kusaya, a traditional Japanese fermented fish product, in the Izu Islands (Niijima and Hachijojima) and the fermentation processes are not clear. In this study, we aimed to investigate the relationship between the microbiota and volatile compounds involved in the manufacturing and management of kusaya gravy. 16S ribosomal RNA (rRNA) gene-based amplicon sequencing revealed that the microbiota in kusaya gravy was significantly different between the two islands, and the microbiota hardly changed during each fermentation process. Gas chromatography-mass spectrometry analysis also revealed that the volatile components were strongly related to the microbiota in kusaya gravy, with Hachijojima samples containing sulfur-containing compounds and Niijima samples containing short-chain fatty acids. Therefore, our findings suggest that kusaya gravy is a characteristic fermented gravy with a stable microbiota, and the fermented pickling gravy is fermented by microorganisms.

Sulfate reduction and homoacetogenesis at various hypersaline conditions: Implications for H2 underground gas storage

While interest in underground gas storage (UGS) of hydrogen (H2) in salt caverns is increasing in a growing H2 economy, knowledge of the microbial communities inhabiting those hypersaline environments is still scarce. High salt concentrations and limited availability of carbon (C) sources in cavern environments reduce microbial growth rates and metabolic activities. Growth conditions potentially change once H2 is stored in salt caverns. H2 is a universal electron donor that can facilitate autotrophic growth and subsequently growth of heterotrophs. In this study, a mixed culture enriched from hypersaline UGS sites was investigated in microcosm experiments with H2 atmosphere, testing the effect of different salt concentrations and C sources on methanogenesis, sulfate reduction and homoacetogenesis. Sulfate, acetate and lactate concentrations were quantified throughout a time span of 125 days of incubation and correlations with the microbial community structure and function were explored through 16S rRNA gene-based amplicon sequencing at the end of the experiment. Many of the amplicon sequence variants (ASVs) were only assigned to family or order level, reflecting that a large number of ASVs belong to previously undescribed taxa. At 4.4 M NaCl, close to cavern brine salinity, members of the Desulfovibrionales were absent when no C source other than CO2 was offered as the sole C source. This is in line with the finding that no sulfate reduction occurred at these conditions. Acetogenic Halanaerobiia dominated in these high salinity levels. Based on metagenome sequencing of four selected samples, we found that acetogenesis at autotrophic conditions relies on the activity of a halophilic homoacetogen, Acetohalobium sp. and that sulfate reduction can most likely be associated with a so far undescribed member of the Desulfonatronovibrionales. We further discuss the discrepancy between sulfate reduction at heterotrophic conditions, while no sulfate reduction was observed under autotrophic conditions although acetate was produced through the homoacetogenic activity of Acetohalobium. For the application of UGS of H2, this means that the presence and activity of the aforementioned microorganisms must be investigated, as they can eventually lead to the formation of acetate and allow sulfate reduction at relevant concentrations.

제주 해안 노출 시기에 따른 해조류 유래 미생물 군집 구조 및 다양성 비교 분석

Seaweeds have become a global industrial source material in food and cosmetic area. In recent reports, seaweed-associated bacteria play a vital role in nutrient shifts with secondary metabolites in direct and/or indirect ways. Here, we investigated the shift of bacterial communities in two different seaweed samples along with incubation and exposure duration to seashore using 16S rRNA gene-based amplicon sequencing. Bacterial communities belonging to the phyla Proteobacteria and Bacteroidetes are the most abundant on seaweeds surface. Bacterial diversity in Ulva pertusa is increased along with distances from seashore while that of Sargassum horneri are similar. Remarkably, Cobetia sp. having alginate-degrading activity represent with high proportion about 80% in. Also, the abundance of Agar-Degrading Bacteria(ADB) are fluctuated with incubation periods in mesocosm experiment, ADB predominantly present after 7 days incubation while ADB are reduced their portion after 18 days. These features would be consequences of seaweed importance to proper periods reached land. To understand the functional roles of associated bacteria in seaweeds, combined further studies should be performed coping with biochemical, physiological and microbiological approach.

Microbiome-scale analysis of aerosol facemask contamination during nebulization therapy in hospital

Microbial contamination of aerosol facemasks could be a source of nosocomial infections during nebulization therapy in hospital, prompting efforts to identify these contaminants. Identification of micro-organisms in medical devices has traditionally relied on culture-dependent methods, which are incapable of detecting the majority of these microbial contaminants. This challenge could be overcome with culture-independent sequencing-based techniques that are suited for the profiling of complex microbiomes. To characterize the microbial contaminants in aerosol facemasks used for nebulization therapy, and identify factors influencing the composition of these microbial contaminants with the acquisition and analysis of comprehensive microbiome-scale profiles using culture-independent high-throughput sequencing. Used aerosol facemasks collected from hospitalized patients were analysed with culture-independent 16S rRNA gene-based amplicon sequencing to acquire microbiome-scale comprehensive profiles of the microbial contaminants. Microbiome-based analysis was performed to identify potential sources of microbial contamination in facemasks. Culture-independent high-throughput sequencing was demonstrated for the capacity to acquire microbiome-scale profiles of microbial contaminants on aerosol facemasks. Microbial source identification enabled by the microbiome-scale profiles linked microbial contamination on aerosol facemasks to the human skin and oral microbiota. Antibiotic treatment with levofloxacin was found to reduce contamination of the facemasks by oral microbiota. Sequencing-based microbiome-scale analysis is capable of providing comprehensive characterization of microbial contamination in aerosol facemasks. Insight gained from microbiome-scale analysis facilitates the development of effective strategies for the prevention and mitigation of the risk of nosocomial infections arising from exposure to microbial contamination of aerosol facemasks, such as targeted elimination of potential sources of contamination.

A unified and simple medium for growing model methanogens.

Apart from their archetypic use in anaerobic digestion (AD) methanogenic archaea are targeted for a wide range of applications. Using different methanogenic archaea for one specific application requires the optimization of culture media to enable the growth of different strains under identical environmental conditions, e.g., in microbial electrochemical technologies (MET) for (bio)electromethanation. Here we present a new culture medium (BFS01) adapted from the DSM-120 medium by omitting resazurin, yeast extract, casitone, and using a low salt concentration, that was optimized for Methanosarcina barkeri, Methanobacterium formicicum, and Methanothrix soehngenii. The aim was to provide a medium for follow-up co-culture studies using specific methanogens and Geobacter spp. dominated biofilm anodes. All three methanogens showed growth and activity in the BFS01 medium. This was demonstrated by estimating the specific growth rates ( ) and doubling times ( ) of each methanogen. The and based on methane accumulation in the headspace showed values consistent with literature values for M. barkeri and M. soehngenii. However, and based on methane accumulation in the headspace differed from literature data for M. formicicum but still allowed sufficient growth. The lowered salt concentration and the omission of chemically complex organic components in the medium may have led to the observed deviation from and for M. formicicum as well as the changed morphology. 16S rRNA gene-based amplicon sequencing and whole genome nanopore sequencing further confirmed purity and species identity.

Engineering of the chloroaniline-catabolic plasmid pDCA-1 and its potential for genetic bioaugmentation

Plasmids disseminate pollutant-catabolic genes through horizontal transfer and contribute to the catabolic potential of the host microbial community. Genetic bioaugmentation of conjugative catabolic plasmids has recently been considered an effective and novel approach for long-term bioremediation of contaminated sites. In this study, we engineered an IncP-1β-2 subgroup plasmid pDCA-1 from Achromobacter sp. ANB-1 for its potential application in genetic bioaugmentation. A dcaA1A2B cluster responsible for the deamination of chloroaniline to chlorocatechol and a 1,2-dioxygenase gene (ccdC) for the ring cleavage of chlorocatechol were found to locate on the broad host range plasmid pDCA-1. Insertion of an amidase gene (phh or tccA2) at the accessory region of plasmid pDCA-1 greatly expanded its catabolic substrate spectrum from chloroaniline (3-chloroaniline and 3,4-dichloroaniline) to herbicides (linuron, propanil, propham, and chlorpropham), bacteriostatic agent (triclocarban), plant regulator (forchlorfenuron), and insecticide (diflubenzuron). Taking advantages of high-throughput cell sorting and 16S rRNA gene-based amplicon sequencing, we depicted the diversity of bacterial recipients for the engineered plasmid pDCA-1-gfp-phh in a soil bacterial consortium, showing an excellent conjugative transfer capacity of the pDCA-1 derivative to various Gram negative and even positive strains in natural environment. All in all, the engineered plasmid pDCA-1 had a great potential in genetic bioaugmentation of the sites contaminated with chloroanilines and their derivatives.

Microbiome Profiles of Nebulizers in Hospital Use.

Background: Nebulizers are used to provide treatment to respiratory patients. Concerns over nosocomial infection risks from contaminated nebulizers raise the critical need to identify all microbial populations in nebulizers used by patients. However, conventional culture-dependent techniques are inadequate with the ability to identify specific microbial populations only. Therefore, the aims of this study were to acquire complete profiles of microbiomes in nebulizers used by in-patients with culture-independent high-throughput sequencing and identify sources of microbial contaminants for the development of effective practices to reduce microbial contamination in nebulizer devices. Methods: This study was conducted at the University of Tennessee Medical Center in Knoxville, TN. Nebulizers were collected between May 2018 and October 2018 from inpatients admitted to the floors for pneumonia or chronic obstructive pulmonary disease exacerbations. Nebulizers were sampled for 16S rRNA gene-based amplicon sequencing to profile nebulizer microbiomes and perform phylogenetic analysis. A Bayesian community-wide culture-independent microbial source tracking technique was used to quantify the contribution of human-associated microbiota as potential sources of nebulizer contamination. Results: Culture-independent sequencing detected diverse microbial populations in nebulizers, represented by 18 abundant genera. Stenotrophomonas was identified as the most abundant genus, accounting for 12.4% of the nebulizer microbiome, followed by Rhizobium, Staphylococcus, Streptococcus, and Ralstonia. Phylogenetic analysis revealed the presence of multiple phylotypes with close relationship to potential pathogens. Contributing up to 15% to nebulizer microbiomes, human-associated microbiota was not identified as the primary sources of nebulizer contamination. Conclusion: Culture-independent sequencing was demonstrated to be capable of acquiring comprehensive profiles of microbiomes in nebulizers used by in-patients. Phylogenetic analysis identified differences in pathogenicity between closely related phylotypes. Microbiome profile-enabled community-wide culture-independent microbial source tracking suggested greater importance of environmental sources than human sources as contributors to nebulizer microbiomes, providing important insight for the development of effective strategies for the monitoring and control of nebulizer devices to mitigate infection risks in the hospital.

Changes of the vaginal microbiota in HPV infection and cervical intraepithelial neoplasia: a cross-sectional analysis

This study aimed to explore the changes of the vaginal microbiota and enzymes in the women with high-risk human papillomavirus (HR-HPV) infection and cervical lesions. A total of 448 participants were carried out HPV genotyping, cytology tests, and microecology tests, and 28 participants were treated as sub-samples, in which vaginal samples were characterized by sequencing the bacterial 16S V4 ribosomal RNA (rRNA) gene region. The study found the prevalence of HR-HPV was higher in patients with BV (P = 0.036). The HR-HPV infection rate was 72.73% in G. vaginalis women, which was significantly higher than that of women with lactobacillus as the dominant microbiota (44.72%) (P = 0.04). The positive rate of sialidase (SNA) was higher in women with HR-HPV infection (P = 0.004) and women diagnosed with cervical intraepithelial neoplasia (CIN) (P = 0.041). In HPV (+) women, the α-diversity was significantly higher than that in HPV (−) women. The 16S rRNA gene-based amplicon sequencing results showed that Lactobacillus was the dominant bacteria in the normal vaginal microbiota. However, the proportion of Gardnerella and Prevotella were markedly increased in HPV (+) patients. Gardnerella and Prevotella are the most high-risk combination for the development of HPV (+) women. The SNA secreted by Gardnerella and Prevotella may play a significant role in HPV infection progress to cervical lesions.

Impacts of Different Habitats on the Composition of Bacterial Communities at the Discharging Endpoints of a Hypogene Thermal Karst System

The discharging zones of thermal karst caves provide special environmental conditions for bacteria involved in the biogeochemical processes. The taxonomic composition of bacterial communities inhabiting these still active natural habitats is largely unknown. The aim of this study was to reveal and compare the bacterial diversity of upwelling thermal waters, rock surface biofilms, and cave deposits using high-throughput next-generation sequencing. Two thermal karst caves formed in carbonate rocks and located at the discharging endpoints of the Transdanubian Range (Hungary) were studied: a sublacustrine spring cave and a phreatic hypogene cave. The results of 16S rRNA gene-based Illumina MiSeq amplicon sequencing revealed that almost all samples were dominated by Proteobacteria and/or Epsilonbacteraeota. Other abundant phyla showed habitat-dependent and diverse distributions. Sequences of the thermal waters were mainly associated with different chemolithotrophic sulfur-oxidizing bacteria (genera Thiovirga, Thiofaba, and Arcobacter in the sublacustrine thermal spring cave, whereas genera Sulfurovum, Sulfuricurvum, and Sulfurifustis were characteristic of the phreatic hypogene thermal cave) in accordance with the difference in environmental parameters. Sequences belonging to ammonia-oxidizing (unclassified Nitrosococcaceae) and nitrite-oxidizing bacteria (Nitrospira) were characteristic of the hypogene cave. Sequences related to photoautotrophic bacteria from the lake water and chemoheterotrophic bacteria from the sediment were only found in the sublacustrine spring cave samples.

Characterization of External Mucosal Microbiomes of Nile Tilapia and Grey Mullet Co-cultured in Semi-Intensive Pond Systems.

The external mucosal surfaces of the fish harbor complex microbial communities, which may play pivotal roles in the physiological, metabolic, and immunological status of the host. Currently, little is known about the composition and role of these communities, whether they are species and/or tissue specific and whether they reflect their surrounding environment. Co-culture of fish, a common practice in semi-intensive aquaculture, where different fish species cohabit in the same contained environment, is an easily accessible and informative model toward understanding such interactions. This study provides the first in-depth characterization of gill and skin microbiomes in co-cultured Nile tilapia (Oreochromis niloticus) and grey mullet (Mugil capito) in semi-intensive pond systems in Egypt using 16S rRNA gene-based amplicon sequencing. Results showed that the microbiome composition of the external surfaces of both species and pond water was dominated by the following bacterial phyla: Proteobacteria, Fusobacteriota, Firmicutes, Planctomycetota, Verrucomicrobiota, Bacteroidota, and Actinobacteriota. However, water microbial communities had the highest abundance and richness and significantly diverged from the external microbiome of both species; thus, the external autochthonous communities are not a passive reflection of their allochthonous communities. The autochthonous bacterial communities of the skin were distinct from those of the gill in both species, indicating that the external microbiome is likely organ specific. However, gill autochthonous communities were clearly species specific, whereas skin communities showed higher commonalities between both species. Core microbiome analysis identified the presence of shared core taxa between both species and pond water in addition to organ-specific taxa within and between the core community of each species. These core taxa included possibly beneficial genera such as Uncultured Pirellulaceae, Exiguobacterium, and Cetobacterium and opportunistic potential pathogens such as Aeromonas, Plesiomonas, and Vibrio. This study provides the first in-depth mapping of bacterial communities in this semi-intensive system that in turn provides a foundation for further studies toward enhancing the health and welfare of these cultured fish and ensuring sustainability.