Summary
The study of complex microbial communities poses unique conceptual and analytical challenges, with microbial species potentially numbering in the thousands. With transient or allochthonous ...microorganisms often adding to this complexity, a ‘core’ microbiota approach, focusing only on the stable and permanent members of the community, is becoming increasingly popular. Given the various ways of defining a core microbiota, it is prudent to examine whether the definition of the core impacts upon the results obtained. Here we used complex marine sponge microbiotas and undertook a systematic evaluation of the degree to which different factors used to define the core influenced the conclusions. Significant differences in alpha‐ and beta‐diversity were detected using some but not all core definitions. However, findings related to host specificity and environmental quality were largely insensitive to major changes in the core microbiota definition. Furthermore, none of the applied definitions altered our perception of the ecological networks summarising interactions among bacteria within the sponges. These results suggest that, while care should still be taken in interpretation, the core microbiota approach is surprisingly robust, at least for comparing microbiotas of closely related samples.
Sponges (phylum Porifera) are early-diverging metazoa renowned for establishing complex microbial symbioses. Here we present a global Porifera microbiome survey, set out to establish the ecological ...and evolutionary drivers of these host-microbe interactions. We show that sponges are a reservoir of exceptional microbial diversity and major contributors to the total microbial diversity of the world's oceans. Little commonality in species composition or structure is evident across the phylum, although symbiont communities are characterized by specialists and generalists rather than opportunists. Core sponge microbiomes are stable and characterized by generalist symbionts exhibiting amensal and/or commensal interactions. Symbionts that are phylogenetically unique to sponges do not disproportionally contribute to the core microbiome, and host phylogeny impacts complexity rather than composition of the symbiont community. Our findings support a model of independent assembly and evolution in symbiont communities across the entire host phylum, with convergent forces resulting in analogous community organization and interactions.
Summary
Marine sponge reefs usually comprise a complex array of taxonomically different sponge species, many of these hosting highly diverse microbial communities. The number of microbial species ...known to occupy a given sponge ranges from tens to thousands, bringing numerous challenges to their analysis. One way to deal with such complexity is to use a core microbiota approach, in which only prevalent and abundant microbes are considered. Here we aimed to test the strength and sensitivity of the core microbiota approach by applying different core definitions to 20 host sponge species. Application of increasingly stringent relative abundance and/or percentage occurrence thresholds to qualify as part of the core microbiota decreased the number of ‘core’ OTUs and phyla and, consequently, changed both alpha‐ and beta‐diversity patterns. Moreover, microbial co‐occurrence patterns explored using correlation networks were also affected by the core microbiota definition. The application of stricter thresholds resulted in smaller and less compartmentalized networks, with different keystone species. These results highlight that the application of different core definitions to phylogenetically disparate host species can result in the drawing of markedly different conclusions. Consequently, we recommend to assess the effects of different core community definitions on the specific system of study before considering its application.
Summary
Many marine sponges contain dense and diverse communities of associated microorganisms. Members of the ‘sponge‐associated unclassified lineage’ (SAUL) are frequently recorded from sponges, ...yet little is known about these bacteria. Here we investigated the distribution and phylogenetic status of SAUL. A meta‐analysis of the available literature revealed the widespread distribution of this clade and its association with taxonomically varied sponge hosts. Phylogenetic analyses, conducted using both 16S rRNA gene‐based phylogeny and concatenated marker protein sequences, revealed that SAUL is a sister clade of the candidate phylum ‘Latescibacteria’. Furthermore, we conducted a comprehensive analysis of two draft genomes assembled from sponge metagenomes, revealing novel insights into the physiology of this symbiont. Metabolic reconstruction suggested that SAUL members are aerobic bacteria with facultative anaerobic metabolism, with the capacity to degrade multiple sponge‐ and algae‐derived carbohydrates. We described for the first time in a sponge symbiont the putative genomic capacity to transport phosphate into the cell and to produce and store polyphosphate granules, presumably constituting a phosphate reservoir for the sponge host in deprivation periods. Our findings suggest that the lifestyle of SAUL is symbiotic with the host sponge, and identify symbiont factors which may facilitate the establishment and maintenance of this relationship.
Marine coastal contamination caused by human activity is a major issue worldwide. The implementation of effective pollution monitoring programs, especially in coastal areas, is important and urgent. ...The use of biological, physiological, or biochemical measurements to monitor the impacts of pollution has garnered increasing interest, particularly for the development of new non-invasive tools to assess water pollution. Fish skin mucus is in direct contact with the marine environment, making it a favourable microenvironment for the formation of biofilm bacterial communities. In this study, we developed a non-invasive technique, sampling fish skin mucus to determine and analyse bacterial community composition using next-generation sequencing. We hypothesised that bacterial communities associated with the skin mucus of a common harbour benthic blennioid triplefin fish, Forsterygion capito, would reflect conditions of different marine environments. We detected clear differences in bacterial community alpha-diversity between contaminated and reference sites. Beta-diversity analysis also revealed differences in the bacterial community structure of the skin mucus of fish inhabiting different geographical areas. The relative abundance of different bacterial orders varied among sites, as determined by linear discriminant analysis (LDA) and effect size (LEfSe) analyses. The observed variation in bacterial community compositions correlated more strongly with variation in hydrocarbons than to various metal concentrations. Using advanced DNA sequencing technologies, we have developed a novel non-invasive, low-cost and effective tool to monitor the impacts of pollution through analysis of the bacterial communities associated with fish skin mucus.
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•α-diversity in contaminated sites were richer and more diverse.•β-diversity revealed microbial differences in different geographical areas.•Bacterial community compositions in contaminated sites were more strongly associated with hydrocarbons.
A non-invasive, novel method using bacterial DNA sequencing of fish skin mucus was successfully developed and applied on blennioid fish.
Microorganisms play fundamental roles in the diversity and functional stability of environments, including nutrient and energy cycling. However, microbial biodiversity loss and change because of ...global climate and land use change remain poorly understood. Many microbial taxa exhibit fast growth rates and are highly sensitive to environmental change. This suggests they have potential to be efficient biological indicators to assess and monitor the state of the habitats within which they occur. Here, we describe and illustrate a range of univariate and multivariate statistical approaches that can be used to identify effective microbial indicators of environmental perturbations and quantify changes in microbial communities. We show that the integration of multiple approaches, such as linear discriminant analysis effect size and indicator value analysis, is optimal for the quantification of the effects of perturbation on microbial communities. We demonstrate the most prevalent techniques using microbial community data derived from soils under different land uses. We discuss the limitations to the development and use of microbial bioindicators and identify future research directions, such as the creation of reliable, standardised reference databases to provide baseline metrics that are indicative of healthy microbial communities. If reliable and globally-relevant microbial indicators of environmental health can be developed, there is enormous potential for their use, both as a standalone monitoring tool and via their integration with existing physical, chemical and biological measures of environmental health.
Adenotonsillar and middle ear diseases result in some of the most frequently performed operations in the pediatric population worldwide. The pathogen reservoir hypothesis (PRH) suggests that the ...adenoids act as a reservoir of bacteria which play a potential pathogenic role in otitis media. Evidence supporting this hypothesis is limited. This study sought to comprehensively determine and compare associations between the adenotonsillar and middle ear bacterial microbiota within individual patients via next-generation sequencing and microbial network analyses.
Bacterial 16S rRNA gene-targeted amplicon sequencing was used to determine the bacterial composition of ten pediatric patients undergoing adenotonsillectomy and ventilation tube insertion for otitis media with effusion. At the time of surgery, swabs were taken from the adenoid surface, tonsil crypts and middle ear clefts (through the myringotomy incision).
The most abundant sequences within the bacterial community at genus level across all anatomical sites were Fusobacterium, Haemophilus, Neisseria, and Porphyromonas. There was an observable difference in the relative abundance of bacterial communities, with a higher proportion of Haemophilus and Moraxella in the adenoid when compared with the middle ear. Furthermore, only one module (consisting of 4 bacterial OTUs) from one patient was identified through microbial network analyses to be significantly associated between middle ear and adenoid. In addition, microbial network analysis revealed that the adenoid and tonsil microbiota share greater similarity than do the adenoid and middle ear.
The results of this study suggest that the adenoid microenvironment does not correlate to the middle ear microenvironment. A future study at the species level, and over time, is required to further investigate whether the differing relationship between the microbiota of the adenoid and middle ear rejects the pathogen reservoir hypothesis.
Background & Aims Acid sphingomyelinase (ASMase) is activated in non-alcoholic steatohepatitis (NASH). However, the contribution of ASMase to NASH is poorly understood and limited to hepatic ...steatosis and glucose metabolism. Here we examined the role of ASMase in high fat diet (HFD)-induced NASH. Methods Autophagy, endoplasmic reticulum (ER) stress and lysosomal membrane permeabilization (LMP) were determined in ASMase−/− mice fed a HFD. The impact of pharmacological ASMase inhibition on NASH was analyzed in wild type mice fed a HFD. Results ASMase deficiency determined resistance to hepatic steatosis mediated by a HFD or methionine-choline deficient diet. ASMase−/− mice were resistant to HFD-induced hepatic ER stress, but sensitive to tunicamycin-mediated ER stress, indicating selectivity in the resistance of ASMase−/− mice to ER stress and steatosis. Autophagic flux, determined in the presence of rapamycin and/or chloroquine, was lower in primary mouse hepatocytes (PMH) from ASMase−/− mice and accompanied by increased p62 levels, suggesting autophagic impairment. Moreover, autophagy suppression by chloroquine and brefeldin A caused ER stress in PMH from ASMase+/+ mice but not in ASMase−/− mice. ASMase−/− PMH exhibited increased lysosomal cholesterol loading, decreased LMP and apoptosis resistance induced by O-methyl-serine dodecylamide hydrochloride or palmitic acid, effects that were reversed by decreasing cholesterol levels by oxysterol 25-hydroxycholesterol. In vivo pharmacological ASMase inhibition by amitriptyline, a widely used tricyclic antidepressant, protected wild type mice against HFD-induced hepatic steatosis, fibrosis, and liver damage, effects indicative of early-stage NASH. Conclusions These findings underscore a critical role for ASMase in diet-induced NASH and suggest the potential of amitriptyline as a treatment for patients with NASH.
Obstructive sleep apnea (OSA) is now a more common indication for tonsillectomy than recurrent tonsillitis (RT) 1,2. Few studies have addressed possible differences in pathogenesis between these two ...conditions. Children with RT and OSA are often being treated in the community with multiple courses of antibiotics before surgery. Current understanding of the role of bacteria in disorders of the tonsils is mainly based on the culture of tonsil swabs. Swab cultures reflect only a very small fraction of the bacteria present on the mucosal surface and may not represent the bacterial communities within the tonsil crypts 3,4. This study aimed to evaluate the local lymphocyte response and associations with bacterial community composition using molecular techniques of the tonsils removed from children for RT or OSA.
The palatine tonsils were removed by extracapsular dissection from 24 patients with age range one to ten years, 14 of whom had RT and 10 had OSA. The fixed tonsil tissues were evaluated for bacteria by Gram-staining and presence of connective tissue by safranin staining. B lymphocytes and T lymphocytes were also measured immunohistochemically. Finally, previously published bacterial community data for this cohort were reassessed for associations with RT and OSA, and with the observed lymphocyte patterns.
In tonsils from patients with RT, large micro-colonies of bacteria were observed in the tonsil crypts, and a large number of B and T lymphocytes were noted immediately adjacent to the tonsil crypt itself. In marked contrast, the tonsils from patients with OSA had no bacteria identified, and no significant skewing of lymphocytes based on site (such as follicles or crypts). We observed that the majority of lymphocytes surrounding the bacterial micro-colonies were B lymphocytes with a mean ratio of 109:55 (B lymphocytes: T lymphocytes). Bacterial community diversity was not different between the two cohorts; however, there were significant differences in bacterial community composition. Children with RT had a higher relative abundance of members from the genera Parvimonas, Prevotella, and Treponema. While children with OSA had a higher relative abundance of Haemophilus, and Capnocytophaga.
These results demonstrate significant differences in the local lymphocyte response and bacterial community composition in tonsil tissue between RT and OSA patients. It suggests that the response to antibiotics used in the treatment of these two conditions may be different. Furthermore, the presence of lymphocytes in RT within the tonsil crypt outside the tonsil epithelium is a unique observation of the location of these cells.
Abstract
Marine sponges (phylum Porifera) are a diverse, phylogenetically deep-branching clade known for forming intimate partnerships with complex communities of microorganisms. To date, 16S rRNA ...gene sequencing studies have largely utilised different extraction and amplification methodologies to target the microbial communities of a limited number of sponge species, severely limiting comparative analyses of sponge microbial diversity and structure. Here, we provide an extensive and standardised dataset that will facilitate sponge microbiome comparisons across large spatial, temporal, and environmental scales. Samples from marine sponges (n = 3569 specimens), seawater (n = 370), marine sediments (n = 65) and other environments (n = 29) were collected from different locations across the globe. This dataset incorporates at least 268 different sponge species, including several yet unidentified taxa. The V4 region of the 16S rRNA gene was amplified and sequenced from extracted DNA using standardised procedures. Raw sequences (total of 1.1 billion sequences) were processed and clustered with (i) a standard protocol using QIIME closed-reference picking resulting in 39 543 operational taxonomic units (OTU) at 97% sequence identity, (ii) a de novo clustering using Mothur resulting in 518 246 OTUs, and (iii) a new high-resolution Deblur protocol resulting in 83 908 unique bacterial sequences. Abundance tables, representative sequences, taxonomic classifications, and metadata are provided. This dataset represents a comprehensive resource of sponge-associated microbial communities based on 16S rRNA gene sequences that can be used to address overarching hypotheses regarding host-associated prokaryotes, including host specificity, convergent evolution, environmental drivers of microbiome structure, and the sponge-associated rare biosphere.
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