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Abstract Insect gut microbiomes play a crucial role in the insect development and are shaped, among other factors, by the specialized insect diet habits as well as the morphological structure of the gut. Rose chafers (Pachnoda spp.; Coleoptera: Scarabaeidae) have a highly differentiated gut characterized by a pronounced hindgut dilation which resembles a miniaturized rumen. Specifically, the species Pachnoda marginata has not been previously studied in detail in terms of microbial ecology. Here, we show a fine scale study of the highly compartmentalized gut of P. marginata by using amplicon and metagenomic sequencing to shed light on the bacterial, archaeal and fungal communities thriving in each section of the gut. We found a microbial gradient along the gut from aerobic (foregut) to strictly anaerobic communities (hindgut). In addition, we have characterized interesting biological activities and metabolic pathways of gut microbial communities related to cellulose degradation, methane production and sulfate reduction. Taken together, our results reveal the highly diverse microbial community and the potential of P. marginata gut as a source of industrially relevant microbial diversity.

Abstract Background The holistic characterization of different microbiomes in anaerobic digestion (AD) systems can contribute to a better understanding of these systems and provide starting points for bioengineering. The present study investigates the microbiome of 80 European full-scale AD systems. Operational, chemical and taxonomic data were thoroughly collected, analysed and correlated to identify the main drivers of AD processes. Results The present study describes chemical and operational parameters for a broad spectrum of different AD systems. With this data, Spearman correlation and differential abundance analyses were applied to narrow down the role of the individual microorganisms detected. The authors succeeded in further limiting the number of microorganisms in the core microbiome for a broad range of AD systems. Based on 16S rRNA gene amplicon sequencing, MBA03, Proteiniphilum, a member of the family Dethiobacteraceae, the genus Caldicoprobacter and the methanogen Methanosarcina were the most prevalent and abundant organisms identified in all digesters analysed. High ratios for Methanoculleus are often described for agricultural co-digesters. Therefore, it is remarkable that Methanosarcina was surprisingly high in several digesters reaching ratios up to 47.2%. The various statistical analyses revealed that the microorganisms grouped according to different patterns. A purely taxonomic correlation enabled a distinction between an acetoclastic cluster and a hydrogenotrophic one. However, in the multivariate analysis with chemical parameters, the main clusters corresponded to hydrolytic and acidogenic microorganisms, with SAOB bacteria being particularly important in the second group. Including operational parameters resulted in digester-type specific grouping of microbes. Those with separate acidification stood out among the many reactor types due to their unexpected behaviour. Despite maximizing the organic loading rate in the hydrolytic pretreatments, these stages turned into extremely robust methane production units. Conclusions From 80 different AD systems, one of the most holistic data sets is provided. A very distinct formation of microbial clusters was discovered, depending on whether taxonomic, chemical or operational parameters were combined. The microorganisms in the individual clusters were strongly dependent on the respective reference parameters. Graphical Abstract

Abstract Microorganisms are ubiquitously distributed in nature and usually appear as biofilms attached to a variety of surfaces. Here, we report the development of a thick biofilm in the drain pipe of several standard laboratory ice machines, and we describe and characterise, through culture-dependent and -independent techniques, the composition of this oligotrophic microbial community. By using culturomics, 25 different microbial strains were isolated and taxonomically identified. The 16S rRNA high-throughput sequencing analysis revealed that Bacteroidota and Proteobacteria were the most abundant bacterial phyla in the sample, followed by Acidobacteriota and Planctomycetota, while ITS high-throughput sequencing uncovered the fungal community was clearly dominated by the presence of a yet-unidentified genus from the Didymellaceae family. Alpha and beta diversity comparisons of the ice machine microbial community against that of other similar cold oligotrophic and/or artificial environments revealed a low similarity between samples, highlighting the ice machine could be considered a cold and oligotrophic niche with a unique selective pressure for colonisation of particular microorganisms. The recovery and analysis of high-quality metagenome-assembled genomes (MAGs) yielded a strikingly high rate of new species. The functional profiling of the metagenome sequences uncovered the presence of proteins involved in extracellular polymeric substance (EPS) and fimbriae biosynthesis and also allowed us to detect the key proteins involved in the cold adaptation mechanisms and oligotrophic metabolic pathways. The metabolic functions in the recovered MAGs confirmed that all MAGs have the genes involved in psychrophilic protein biosynthesis. In addition, the highest number of genes for EPS biosynthesis was presented in MAGs associated with the genus Sphingomonas, which was also recovered by culture-based method. Further, the MAGs with the highest potential gene number for oligotrophic protein production were closely affiliated with the genera Chryseoglobus and Mycobacterium. Our results reveal the surprising potential of a cold oligotrophic microecosystem within a machine as a source of new microbial taxa and provide the scientific community with clues about which microorganisms are able to colonise this ecological niche and what physiological mechanisms they develop. These results pave the way to understand how and why certain microorganisms can colonise similar anthropogenic environments.

Abstract Bioprospecting of microorganisms suitable for bioremediation of fuel or oil spills is often carried out in contaminated environments such as gas stations or polluted coastal areas. Using next-generation sequencing (NGS) we analyzed the microbiota thriving below the lids of the fuel deposits of diesel and gasoline cars. The microbiome colonizing the tank lids differed from the diversity found in other hydrocarbon-polluted environments, with Proteobacteria being the dominant phylum and without clear differences between gasoline or diesel-fueled vehicles. We observed differential growth when samples were inoculated in cultures with gasoline or diesel as the main carbon source, as well as an increase in the relative abundance of the genus Pseudomonas in diesel. A collection of culturable strains was established, mostly Pseudomonas, Stenotrophomonas, Staphylococcus, and Bacillus genera. Strains belonging to Bacillus, Pseudomonas, Achromobacter, and Isoptericola genera showed a clear diesel degradation pattern when analyzed by GC-MS, suggesting their potential use for bioremediation and a possible new species of Isoptericola was further characterized as hydrocarbon degrader.

Abstract Despite the increasing evidence of links between human gut and health, the number of gut microbiomes that have been studied to date at a country level are surprisingly low. Mediterranean countries, including some of the most long-lived and healthy countries in the world, have not been considered so far in those studies at a large scale. The main objective of this work is to characterize the gut microbiome of a healthy adult population of a Mediterranean, paradigmatically healthy country: Spain. Stool samples from 530 healthy volunteers were collected, total metagenomic DNA extracted, and the microbial profiles determined through 16S rRNA metataxonomic sequencing. Our results confirm the associations between several microbial markers and different variables, including sex, age, BMI and diet choices, and bring new insights into the relationship between microbiome and diet in the Spanish population. Remarkably, some of the associations found, such as the decrease of Faecalibacterium with age or the link of Flavonifractor with less healthy dietary habits, have been barely noticed in other large-scale cohorts. On the other hand, a range of links between microorganisms, diet, and lifestyle coincide with those reported in other populations, thus increasing the robustness of such associations and confirming the importance of these microbial markers across different countries. Overall, this study describes the Spanish “normal” microbiome, providing a solid baseline for future studies investigating the effects of gut microbiome composition and deviations in the adherence to the Mediterranean diet.

Abstract Background Recent research articles indicate that direct interspecies electron transfer (DIET) is an alternative metabolic route for methanogenic archaea that improves microbial methane productivity. It has been shown that multiple conductive materials such as biochar can be supplemented to anaerobic digesters to increase the rate of DIET. However, the industrial applicability, as well as the impact of such supplements on taxonomic profiles, has not been sufficiently assessed to date. Results Seven industrial biogas plants were upgraded with a shock charge of 1.8 kg biochar per ton of reactor content and then 1.8 kg per ton were added to the substrate for one year. A joint analysis for all seven systems showed a decreasing trend for the concentration of acetic acid (p

A demonstrator plant of a recently patented process for improved sludge degradation has been implemented on a municipal scale. In a 1500 m3 sewage sludge digester, an intermediary stage with aerobic sewage sludge reactivation was implemented. This oxic activation increased the biogas yield by up to 55% with a 25% reduction of the remaining fermentation residue volume. Furthermore, this process allowed an NH4-N removal of over 90%. Additionally, 16S rRNA gene amplicon high-throughput sequencing of the reactivated digestate showed a reduced number of methane-forming archaea compared to the main digester. Multiple ammonium-oxidizing bacteria were detected. This includes multiple genera belonging to the family Chitinophagaceae (the highest values reached 18.8% of the DNA sequences) as well as a small amount of the genus Candidatus nitrosoglobus (

Summary Solar panel surfaces can be colonized by microorganisms adapted to desiccation, temperature fluctuations and solar radiation. Although the taxonomic and functional composition of these communities has been studied, the microbial colonization process remains unclear. In the present work, we have monitored this microbial colonization process during 24 months by performing weekly measurements of the photovoltaic efficiency, carrying out 16S rRNA gene high‐throughput sequencing, and studying the effect of antimicrobial compounds on the composition of the microbial biocenosis. This is the first time a long‐term study of the colonization process of solar panels has been performed, and our results reveal that species richness and biodiversity exhibit seasonal fluctuations and that there is a trend towards an increase or decrease of specialist (solar panel‐adapted) and generalist taxa, respectively. On the former, extremophilic bacterial genera Deinococcus, Hymenobacter and Roseomonas and fungal Neocatenulostroma, Symmetrospora and Sporobolomyces tended to dominate the biocenosis; whereas Lactobacillus sp or Stemphyllium exhibited a decreasing trend. This profile was deeply altered by washing the panels with chemical agents (Virkon), but this did not lead to an increase of the solar panels efficiency. Our results show that solar panels are extreme environments that force the selection of a particular microbial community.

Abstract Metagenomic sequencing has allowed for the recovery of previously unexplored microbial genomes. Whereas short-read sequencing platforms often result in highly fragmented metagenomes, nanopore-based sequencers could lead to more contiguous assemblies due to their potential to generate long reads. Nevertheless, there is a lack of updated and systematic studies evaluating the performance of different assembly tools on nanopore data. In this study, we have benchmarked the ability of different assemblers to reconstruct two different commercially-available mock communities that have been sequenced using Oxford Nanopore Technologies platforms. Among the tested tools, only metaFlye, Raven, and Canu performed well in all the datasets. These tools retrieved highly contiguous genomes (or even complete genomes) directly from the metagenomic data. Despite the intrinsic high error of nanopore sequencing, final assemblies reached high accuracy (~ 99.5 to 99.8% of consensus accuracy). Polishing strategies demonstrated to be necessary for reducing the number of indels, and this had an impact on the prediction of biosynthetic gene clusters. Correction with high quality short reads did not always result in higher quality draft assemblies. Overall, nanopore metagenomic sequencing data-adapted to MinION’s current output-proved sufficient for assembling and characterizing low-complexity microbial communities.

Abstract Ocean pollution is a worldwide environmental challenge that could be partially tackled through microbial applications. To shed light on the diversity and applications of the bacterial communities that inhabit the sediments trapped in artificial containers, we analyzed residues (polyethylene terephthalate [PET] bottles and aluminum cans) collected from the Mediterranean Sea by scanning electron microscopy and next generation sequencing. Moreover, we set a collection of culturable bacteria from the plastisphere that were screened for their ability to use PET as a carbon source. Our results reveal that Proteobacteria are the predominant phylum in all the samples and that Rhodobacteraceae, Woeseia, Actinomarinales, or Vibrio are also abundant in these residues. Moreover, we identified marine isolates with enhanced growth in the presence of PET: Aquimarina intermedia, Citricoccus spp., and Micrococcus spp. Our results suggest that the marine environment is a source of biotechnologically promising bacterial isolates that may use PET or PET additives as carbon sources.

Bioprospecting expeditions are often performed in remote locations, in order to access previously unexplored samples. Nevertheless, the actual potential of those samples is only assessed once scientists are back in the laboratory, where a time-consuming screening must take place. This work evaluates the suitability of using Nanopore sequencing during a journey to the Tabernas Desert (Spain) for forecasting the potential of specific samples in terms of bacterial diversity and prevalence of radiation- and desiccation-resistant taxa, which were the target of the bioprospecting activities. Samples collected during the first day were analyzed through 16S rRNA gene sequencing using a mobile laboratory. Results enabled the identification of locations showing the greatest and the least potential, and a second, informed sampling was performed focusing on those sites. After finishing the expedition, a culture collection of 166 strains belonging to 50 different genera was established. Overall, Nanopore and culturing data correlated well, since samples holding a greater potential at the microbiome level also yielded a more interesting set of microbial isolates, whereas samples showing less biodiversity resulted in a reduced (and redundant) set of culturable bacteria. Thus, we anticipate that portable sequencers hold potential as key, easy-to-use tools for in situ-informed bioprospecting strategies.

Commercial table salt is a condiment with food preservative properties by decreasing water activity and increasing osmotic pressure. Salt is also a source of halophilic bacteria and archaea. In the present research, the diversity of halotolerant and halophilic microorganisms was studied in six commercial table salts by culture-dependent and culture-independent techniques. Three table salts were obtained from marine origins: Atlantic Ocean, Mediterranean (Ibiza Island), and Odiel marshes (supermarket marine salt). Other salts supplemented with mineral and nutritional ingredients were also used: Himalayan pink, Hawaiian black, and one with dried vegetables known as Viking salt. The results of 16S rRNA gene sequencing reveal that the salts from marine origins display a similar archaeal taxonomy, but with significant variations among genera. Archaeal taxa Halorubrum, Halobacterium, Hallobellus, Natronomonas, Haloplanus, Halonotius, Halomarina, and Haloarcula were prevalent in those three marine salts. Furthermore, the most abundant archaeal genera present in all salts were Natronomonas, Halolamina, Halonotius, Halapricum, Halobacterium, Haloarcula, and uncultured Halobacterales. Sulfitobacter sp. was the most frequent bacteria, represented almost in all salts. Other genera such as Bacillus, Enterococcus, and Flavobacterium were the most frequent taxa in the Viking, Himalayan pink, and black salts, respectively. Interestingly, the genus Salinibacter was detected only in marine-originated salts. A collection of 76 halotolerant and halophilic bacterial and haloarchaeal species was set by culturing on different media with a broad range of salinity and nutrient composition. Comparing the results of 16S rRNA gene metataxonomic and culturomics revealed that culturable bacteria Acinetobacter, Aquibacillus, Bacillus, Brevundimonas, Fictibacillus, Gracilibacillus, Halobacillus, Micrococcus, Oceanobacillus, Salibacterium, Salinibacter, Terribacillus, Thalassobacillus, and also Archaea Haloarcula, Halobacterium, and Halorubrum were identified at least in one sample by both methods. Our results show that salts from marine origins are dominated by Archaea, whereas salts from other sources or salt supplemented with ingredients are dominated by bacteria.

Anaerobic digestion is a technology known for its potential in terms of methane production. During the digestion process, multiple metabolites of high value are synthesized. However, recent works have demonstrated the high robustness and resilience of the involved microbiomes; these attributes make it difficult to manipulate them in such a way that a specific metabolite is predominantly produced. Therefore, an exact understanding of the manipulability of anaerobic microbiomes may open up a treasure box for bio-based industries. In the present work, the effect of nalidixic acid, γ-aminobutyric acid (GABA), and sodium phosphate on the microbiome of digested sewage sludge from a water treatment plant fed with glucose was investigated. Despite of the induced process perturbations, high stability was observed at the phylum level. However, strong variations were observed at the genus level, especially for the genera Trichococcus, Candidatus Caldatribacterium, and Phascolarctobacterium. Ecological interactions were analyzed based on the Lotka–Volterra model for Trichococcus, Rikenellaceae DMER64, Sedimentibacter, Candidatus Cloacimonas, Smithella, Cloacimonadaceae W5 and Longilinea. These genera dynamically shifted among positive, negative or no correlation, depending on the applied stressor, which indicates a surprisingly dynamic behavior. Globally, the presented work suggests a massive resilience and stability of the methanogenic communities coupled with a surprising flexibility of the particular microbial key players involved in the process.

Natural products have traditionally been discovered through the screening of culturable microbial isolates from diverse environments. The sequencing revolution allowed the identification of dozens of biosynthetic gene clusters (BGCs) within single bacterial genomes, either from cultured or uncultured strains. However, we are still far from fully exploiting the microbial reservoir, as most of the species are non-model organisms with complex regulatory systems that can be recalcitrant to engineering approaches. Genomic and metagenomic data produced by laboratories worldwide covering the range of natural and artificial environments on Earth, are an invaluable source of raw information from which natural product biosynthesis can be accessed. In the present work, we describe the environmental distribution and evolution of the abyssomicin BGC through the analysis of publicly available genomic and metagenomic data. Our results demonstrate that the selection of a pathway-specific enzyme to direct genome mining is an excellent strategy; we identified 74 new Diels–Alderase homologs and unveiled a surprising prevalence of the abyssomicin BGC within terrestrial habitats, mainly soil and plant-associated. We also identified five complete and 12 partial new abyssomicin BGCs and 23 new potential abyssomicin BGCs. Our results strongly support the potential of genome and metagenome mining as a key preliminary tool to inform bioprospecting strategies aimed at the identification of new bioactive compounds such as -but not restricted to- abyssomicins.