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In glacier-fed streams, the Windows of Opportunity (WOs) are periods of mild environmental conditions supporting the seasonal development of benthic microorganisms. WOs have been defined based on changes in biofilm biomass, but the responses of microbial diversity to WOs in Alpine streams have been overlooked. A two year (2017-2018) metabarcoding of epilithic and epipsammic biofilm prokaryotes was conducted in Alpine streams fed by glaciers (kryal), rock glaciers (rock glacial), or groundwater/precipitation (krenal) in two catchments of the Central-Eastern European Alps (Italy), aiming at testing the hypothesis that: 1) environmental WOs enhance not only the biomass but also the α-diversity of the prokaryotic biofilm in all stream types, 2) diversity and phenology of prokaryotic biofilm are mainly influenced by the physical habitat in glacial streams, and by water chemistry in the other two stream types. The study confirmed kryal and krenal streams as endmembers of epilithic and sediment prokaryotic α- and β-diversity, with rock glacial streams sharing a large proportion of taxa with the two other stream types. Alpha-diversity appeared to respond to ecological WOs, but, contrary to expectations, seasonality was less pronounced in the turbid kryal than in the clear streams. This was attributed to the small size of the glaciers feeding the studied kryal streams, whose discharge dynamics were those typical of the late phase of deglaciation. Prokaryotic α-diversity of non-glacial streams tended to be higher in early summer than in early autumn. Our findings, while confirming that high altitude streams are heavily threatened by climate change, underscore the still neglected role of rock glacier runoffs as climate refugia for the most stenothermic benthic aquatic microorganism. This advocates the need to define and test strategies for protecting these ecosystems for preserving, restoring, and connecting cold Alpine aquatic biodiversity in the context of the progressing global warming., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Untargeted genetic approaches can be used to explore the high metabolic versatility of cyanobacteria. In this context, a comprehensive metagenomic shotgun analysis was performed on a population of Dolichospermum lemmermannii collected during a surface bloom in Lake Garda in the summer of 2020. Using a phylogenomic approach, the almost complete metagenome-assembled genome obtained from the analysis allowed to clarify the taxonomic position of the species within the genus Dolichospermum and contributed to frame the taxonomy of this genus within the ADA group (Anabaena/Dolichospermum/Aphanizomenon). In addition to common functional traits represented in the central metabolism of photosynthetic cyanobacteria, the genome annotation uncovered some distinctive and adaptive traits that helped define the factors that promote and maintain bloom-forming heterocytous nitrogen-fixing Nostocales in oligotrophic lakes. In addition, genetic clusters were identified that potentially encode several secondary metabolites that were previously unknown in the populations evolving in the southern Alpine Lake district. These included geosmin, anabaenopetins, and other bioactive compounds. The results expanded the knowledge of the distinctive competitive traits that drive algal blooms and provided guidance for more targeted analyses of cyanobacterial metabolites with implications for human health and water resource use., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

Climate change and anthropogenic disturbances are known to influence soil biodiversity. The objectives of this study were to compare the community composition, species coexistence patterns, and ecological assembly processes of soil microbial communities in a paired setting featuring a natural and an anthropogenic ecosystem facing each other at identical climatic, pedological, and vegetational conditions. A transect gradient from forest to seashore allowed for sampling across different habitats within both sites. The field survey was carried out at two adjacent strips of land within the Po River delta lagoon system (Veneto, Italy) one of which is protected within a natural preserve and the other has been converted for decades into a tourist resort. The anthropogenic pressure interestingly led to an increase in the α-diversity of soil microbes but was accompanied by a reduction in β-diversity. The community assembly mechanisms of microbial communities differentiate in natural and anthropic ecosystems: for bacteria, in natural ecosystems deterministic variables and homogeneous selection play a main role (51.92%), while stochastic dispersal limitation (52.15%) is critical in anthropized ecosystems; for fungi, stochastic dispersal limitation increases from 38.1% to 66.09% passing from natural to anthropized ecosystems. We are on calcareous sandy soils and in more natural ecosystems a variation of topsoil pH favors the deterministic selection of bacterial communities, while a divergence of K availability favors stochastic selection. In more anthropized ecosystems, the deterministic variable selection is influenced by the values of SOC. Microbial networks in the natural system exhibited higher numbers of nodes and network edges, as well as higher averages of path length, weighted degree, clustering coefficient, and density than its equivalent sites in the more anthropically impacted environment. The latter on the other hand presented a stronger modularity. Although the influence of stochastic processes increases in anthropized habitats, niche-based selection also proves to impose constraints on communities. Overall, the functionality of the relationships between groups of microorganisms co-existing in communities appeared more relevant to the concept of functional biodiversity in comparison to the plain number of their different taxa. Fewer but functionally more organized lineages displayed traits underscoring a better use of the resources than higher absolute numbers of taxa when those are not equally interconnected in their habitat exploitation. However, considering that network complexity can have important implications for microbial stability and ecosystem multifunctionality, the extinction of complex ecological interactions in anthropogenic habitats may impair important ecosystem services that soils provide us., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier GmbH.. All rights reserved.)

We report the draft genome sequence of strain B0820 of the cyanobacterium Tychonema bourrellyi isolated from the epilimnion of Lake Garda and assembled from a metagenome of a non-axenic culture. The strain analyzed was shown to produce anatoxin-a, a potent neurotoxin that can cause fatal intoxication in exposed organisms., Competing Interests: The authors declare no conflict of interest.

Oral microbiota-host interactions are gaining recognition as potential factors contributing to interindividual variations in taste perception. However, whether such possible links imply specific bacterial co-occurrence networks remains unknown. To address this issue, we used 16 s rRNA gene sequencing to profile the salivary microbiota of 100 healthy individuals (52 % women; 18-30 y/o), who provided hedonic and psychophysical responses to 5 liquid and 5 solid commercially-available foods, each chosen to elicit a target sensation (sweet, sour, bitter, salty, pungent). The same cohort also completed several psychometric measures and a 4-day food diary. Unsupervised data-driven clustering of genus-level Aitchison distances supported the existence of two salivary microbial profiles (CL-1, CL-2). While CL-1 (n = 57; 49.1 % women) exhibited higher α-diversity metrics and was enriched in microbial genera assigned to the class Clostridia (e.g., Lachnospiraceae_[G-3]), CL-2 (n = 43; 55.8 % women) harbored greater amounts of taxa with potential cariogenic effects (e.g., genus Lactobacillus) and significantly lower abundances of inferred MetaCyc pathways related to the metabolic fate of acetate. Intriguingly, CL-2 showed enhanced responsiveness to warning oral sensations (bitter, sour, astringent) and a higher propensity to crave sweet foods or engage in prosocial behaviors. Further, the same cluster reported habitually consuming more simple carbohydrates and fewer beneficial nutrients (vegetable proteins, monounsaturated fatty acids). In summary, while the mediating role of participants' baseline diet on findings can not be definitively excluded, this work provides evidence suggesting that microbe-microbe and microbe-taste interactions may exert an influence on dietary habits and motivates further research to uncover a potential "core" taste-related salivary microbiota., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Microbial communities in agricultural soils are fundamental for plant growth and in vineyard ecosystems contribute to defining regional wine quality. Managing soil microbes towards beneficial outcomes requires knowledge of how community assembly processes vary across taxonomic groups, spatial scales, and through time. However, our understanding of microbial assembly remains limited. To quantify the contributions of stochastic and deterministic processes to bacterial and fungal assembly across spatial scales and through time, we used 16 s rRNA gene and ITS sequencing in the soil of an emblematic wine-growing region of Italy.Combining null- and neutral-modelling, we found that assembly processes were consistent through time, but bacteria and fungi were governed by different processes. At the within-vineyard scale, deterministic selection and homogenising dispersal dominated bacterial assembly, while neither selection nor dispersal had clear influence over fungal assembly. At the among-vineyard scale, the influence of dispersal limitation increased for both taxonomic groups, but its contribution was much larger for fungal communities. These null-model-based inferences were supported by neutral modelling, which estimated a dispersal rate almost two orders-of-magnitude lower for fungi than bacteria.This indicates that while stochastic processes are important for fungal assembly, bacteria were more influenced by deterministic selection imposed by the biotic and/or abiotic environment. Managing microbes in vineyard soils could thus benefit from strategies that account for dispersal limitation of fungi and the importance of environmental conditions for bacteria. Our results are consistent with theoretical expectations whereby larger individual size and smaller populations can lead to higher levels of stochasticity., (© 2022. The Author(s).)

Microorganisms are ubiquitous in the environment and provide genetic and physiological functions to multicellular organisms. Knowledge on the associated microbiota is becoming highly relevant to understand the host's ecology and biology. Among invertebrates, many examples of endosymbiosis have been described, such as those in corals, ants, and termites. At present, however, little is known on the presence, diversity, and putative roles of the microbiota associated to brachyuran crabs in relation to their environment. In this work we investigated the associated microbiota of three populations of the terrestrial brachyuran crab Chiromantes haematocheir to find evidence of a conserved organ-specific microbiome unrelated to the population of origin and dissimilar from environmental microbial assemblages. Bacterial 16S rRNA gene and fungal ITS sequences were obtained from selected crab organs and environmental matrices to profile microbial communities. Despite the presence of truly marine larval stages and the absence of a gregarious behaviour, favouring microbiota exchanges, we found common, organ-specific microbiota, associated with the gut and the gills of crabs from the different populations (with more than 15% of the genera detected specifically enriched only in one organ). These findings suggest the presence of possible functional roles of the organ-specific microbiota., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Bacci, Fratini, Meriggi, Cheng, Ng, Pindo, Iannucci, Mengoni, Cavalieri and Cannicci.)

Grapevine embodies a fascinating species as regards phenotypic plasticity and genotype-per-environment interactions. The terroir, namely the set of agri-environmental factors to which a variety is subjected, can influence the phenotype at the physiological, molecular, and biochemical level, representing an important phenomenon connected to the typicality of productions. We investigated the determinants of plasticity by conducting a field-experiment where all terroir variables, except soil, were kept as constant as possible. We isolated the effect of soils collected from different areas, on phenology, physiology, and transcriptional responses of skin and flesh of a red and a white variety of great economic value: Corvina and Glera. Molecular results, together with physio-phenological parameters, suggest a specific effect of soil on grapevine plastic response, highlighting a higher transcriptional plasticity of Glera in respect to Corvina and a marked response of skin compared to flesh. Using a novel statistical approach, we identified clusters of plastic genes subjected to the specific influence of soil. These findings could represent an issue of applicative value, posing the basis for targeted agricultural practices to enhance the desired characteristics for any soil/cultivar combination, to improve vineyards management for a better resource usage and to valorize vineyards uniqueness maximizing the terroir-effect., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nanjing Agricultural University.)

Grapevines worldwide are grafted onto Vitis spp. rootstocks in order to improve their tolerance to biotic and abiotic stresses. Thus, the response of vines to drought is the result of the interaction between the scion variety and the rootstock genotype. In this work, the responses of genotypes to drought were evaluated on 1103P and 101-14MGt plants, own-rooted and grafted with Cabernet Sauvignon, in three different water deficit conditions (80, 50, and 20% soil water content, SWC). Gas exchange parameters, stem water potential, root and leaf ABA content, and root and leaf transcriptomic response were investigated. Under well-watered conditions, gas exchange and stem water potential were mainly affected by the grafting condition, whereas under sever water deficit they were affected by the rootstock genotype. Under severe stress conditions (20% SWC), 1103P showed an "avoidance" behavior. It reduced stomatal conductance, inhibited photosynthesis, increased ABA content in the roots, and closed the stomata. The 101-14MGt maintained a high photosynthetic rate, limiting the reduction of soil water potential. This behavior results in a "tolerance" strategy. An analysis of the transcriptome showed that most of the differentially expressed genes were detected at 20% SWC, and more significantly in roots than in leaves. A core set of genes has been highlighted on the roots as being related to the root response to drought that are not affected by genotype nor grafting. Genes specifically regulated by grafting and genes specifically regulated by genotype under drought conditions have been identified as well. The 1103P, more than the 101-14MGt, regulated a high number of genes in both own-rooted and grafted conditions. This different regulation revealed that 1103P rootstock readily perceived the water scarcity and rapidly faced the stress, in agreement with its avoidance strategy.

The aim of this research has been to assess the effect of the dietary protein level on piglet growth and post-weaning diarrhea (PWD) incidence. Piglet fecal microbiota and feces composition were also assessed. The experiment was carried out on 144 weaned piglets (Duroc × Large White; 72 piglets per treatment) and lasted from weaning (at 25 days of age) until the end of the post-weaning phase (at 95 days). Two dietary protein levels were compared: high (HP; 17.5% crude protein on average, during the experiment) and low (LP; 15.5% on average). Lower ( p < 0.01) average daily gain and feed conversion ratio were observed in LP piglets in the first growth phase. However, at the end of the post-weaning period, the growth parameters were not significantly different in the two diets. Diarrhea scores were lower in piglets fed LP diets than in piglets fed HP diets (28.6% of the total vs. 71.4% in the HP piglets). Fibrobacteres , Proteobacteria, and Spirochaetes were more abundant in the feces of the piglets fed LP diets. Feces nitrogen content was lower in piglets fed LP diets. In conclusion, low protein levels in the diet can reduce the incidence of PWD while only marginally affecting growth parameters.

CRISPR/Cas9 genome editing is a modern biotechnological approach used to improve plant varieties, modifying only one or a few traits of a specific variety. However, this technology cannot be easily used to improve fruit quality traits in citrus, due to the lack of knowledge of key genes, long juvenile stage, and the difficulty regenerating whole plants of specific varieties. Here, we introduce a genome editing approach with the aim of producing citrus plantlets whose fruits contain both lycopene and anthocyanins. Our method employs a dual single guide RNA (sgRNA)-directed genome editing approach to knockout the fruit-specific β-cyclase 2 gene, responsible for the conversion of lycopene to beta-carotene. The gene is targeted by two sgRNAs simultaneously to create a large deletion, as well as to induce point mutations in both sgRNA targets. The EHA105 strain of Agrobacterium tumefaciens was used to transform five different anthocyanin-pigmented sweet oranges, belonging to the Tarocco and Sanguigno varietal groups, and 'Carrizo' citrange, a citrus rootstock as a model for citrus transformation. Among 58 plantlets sequenced in the target region, 86% of them were successfully edited. The most frequent mutations were deletions (from -1 to -74 nucleotides) and insertions (+1 nucleotide). Moreover, a novel event was identified in six plantlets, consisting of the inversion of the region between the two sgRNAs. For 20 plantlets in which a single mutation occurred, we excluded chimeric events. Plantlets did not show an altered phenotype in vegetative tissues. To the best of our knowledge, this work represents the first example of the use of a genome editing approach to potentially improve qualitative traits of citrus fruit., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Salonia, Ciacciulli, Pappalardo, Poles, Pindo, Larger, Caruso, Caruso and Licciardello.)

Key Message: We present a high-density integrated map for grapevine, allowing refinement and improved understanding of the grapevine genome, while demonstrating the applicability of the Vitis18K SNP chip for linkage mapping. The improvement of grapevine through biotechnology requires identification of the molecular bases of target traits by studying marker-trait associations. The Vitis18K SNP chip provides a useful genotyping tool for genome-wide marker analysis. Most linkage maps are based on single mapping populations, but an integrated map can increase marker density and show order conservation. Here we present an integrated map based on three mapping populations. The parents consist of the well-known wine cultivars 'Cabernet Sauvignon', 'Corvina' and 'Rhine Riesling', the lesser-known wine variety 'Deckrot', and a table grape selection, G1-7720. Three high-density population maps with an average inter-locus gap ranging from 0.74 to 0.99 cM were developed. These maps show high correlations (0.9965-0.9971) with the reference assembly, containing only 93 markers with large order discrepancies compared to expected physical positions, of which a third is consistent across multiple populations. Moreover, the genetic data aid the further refinement of the grapevine genome assembly, by anchoring 104 yet unanchored scaffolds. From these population maps, an integrated map was constructed which includes 6697 molecular markers and reduces the inter-locus gap distance to 0.60 cM, resulting in the densest integrated map for grapevine thus far. A small number of discrepancies, mainly of short distance, involve 88 markers that remain conflictual across maps. The integrated map shows similar collinearity to the reference assembly (0.9974) as the single maps. This high-density map increases our understanding of the grapevine genome and provides a useful tool for its further characterization and the dissection of complex traits., (© 2022. The Author(s).)

The taxonomic identification of organisms based on the amplification of specific genetic markers (metabarcoding) implicitly requires adequate discriminatory information and taxonomic coverage of environmental DNA sequences in taxonomic databases. These requirements were quantitatively examined by comparing the determination of cyanobacteria and microalgae obtained by metabarcoding and light microscopy. We used planktic and biofilm samples collected in 37 lakes and 22 rivers across the Alpine region. We focused on two of the most used and best represented genetic markers in the reference databases, namely the 16S rRNA and 18S rRNA genes. A sequence gap analysis using blastn showed that, in the identity range of 99-100%, approximately 30% (plankton) and 60% (biofilm) of the sequences did not find any close counterpart in the reference databases (NCBI GenBank). Similarly, a taxonomic gap analysis showed that approximately 50% of the cyanobacterial and eukaryotic microalgal species identified by light microscopy were not represented in the reference databases. In both cases, the magnitude of the gaps differed between the major taxonomic groups. Even considering the species determined under the microscope and represented in the reference databases, 22% and 26% were still not included in the results obtained by the blastn at percentage levels of identity ≥95% and ≥97%, respectively. The main causes were the absence of matching sequences due to amplification and/or sequencing failure and potential misidentification in the microscopy step. Our results quantitatively demonstrated that in metabarcoding the main obstacles in the classification of 16S rRNA and 18S rRNA sequences and interpretation of high-throughput sequencing biomonitoring data were due to the existence of important gaps in the taxonomic completeness of the reference databases and the short length of reads. The study focused on the Alpine region, but the extent of the gaps could be much greater in other less investigated geographic areas., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Erwinia amylovora is a Gram-negative bacterium that colonizes a wide variety of plant species causing recurrent local outbreaks of fire blight in crops of the Rosaceae family. Recent genomic surveys have documented the limited genomic diversity of this species, possibly related to a recent evolutionary bottleneck and a strong correlation between geography and phylogenetic structure of the species. Despite its economic importance, little is known about the genetic variability of co-circulating strains during local outbreaks. Here, we report the genome sequences of 82 isolates of E. amylovora, collected from different host plants in a period of 16 years in Trentino, a small region in the Northeastern Italian Alps that has been characterized by recurrent outbreaks of fire blight in apple orchards. While the genome isolated before 2018 are closely related to other strains already present in Europe, we found a novel subclade composed only by isolates that were sampled starting from 2018 and demonstrate that the endemic population of this pathogen can be composed by mixture of strains., (© 2022 The Authors. Environmental Microbiology Reports published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

Despite the increasing interest in using microbial-based technologies to support human space exploration, many unknowns remain not only on bioprocesses but also on microbial survivability and genetic stability under non-Earth conditions. Here the desert cyanobacterium Chroococcidiopsis sp. CCMEE 029 was investigated for robustness of the repair capability of DNA lesions accumulated under Mars-like conditions (UV radiation and atmosphere) simulated in low Earth orbit using the EXPOSE-R2 facility installed outside the International Space Station. Genomic alterations were determined in a space-derivate of Chroococcidiopsis sp. CCMEE 029 obtained upon reactivation on Earth of the space-exposed cells. Comparative analysis of whole-genome sequences showed no increased variant numbers in the space-derivate compared to triplicates of the reference strain maintained on the ground. This result advanced cyanobacteria-based technologies to support human space exploration., (© 2022. The Author(s).)

Resident microbial communities that can support various host functions play a key role in their development and health. In fishes, microbial symbionts are vertically transferred from the parents to their progeny. Such transfer of microbes in mouthbrooder fish species has not been reported yet. Here, we employed Nile tilapia ( Oreochromis niloticus ) to investigate the vertical transmission of microbes across generations using a 16S rRNA amplicon sequencing approach, based on the presence of bacteria in different generations. Our analysis revealed that the core microbiome in the buccal cavity and posterior intestine of parents shapes the gut microbiome of the progeny across generations. We speculate that the route of this transmission is via the buccal cavity. The identified core microbiome bacteria, namely Nocardioides, Propionibacterium , and Sphingomonas have been reported to play an essential role in the health and development of offspring. These core microbiome members could have specific functions in fish, similar to mammals., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Abdelhafiz, Fernandes, Donati, Pindo and Kiron.)

Background: Trentino is an Italian province with a tourism-based economy, bordering the regions of Lombardy and Veneto, where the two earliest and largest outbreaks of COVID-19 occurred in Italy. The earliest cases in Trentino were reported in the first week of March 2020, with most of the cases occurring in the winter sport areas in the Dolomites mountain range. The number of reported cases decreased over the summer months and was followed by a second wave in the autumn and winter of 2020., Methods: we performed high-coverage Oxford Nanopore sequencing of 253 positive SARS-CoV-2 swabs collected in Trentino between March and December 2020., Results: in this work, we analyzed genome sequences to trace the routes through which the virus entered the area, and assessed whether the autumnal resurgence could be attributed to lineages persisting undetected during summer, or as a consequence of new introductions., Conclusions: Comparing the draft genomes analyzed with a large selection of European sequences retrieved from GISAID we found that multiple introductions of the virus occurred at the early stage of the epidemics; the two epidemic waves were unrelated; the second wave was due to reintroductions of the virus in summer when traveling restrictions were uplifted.

Background: The microbiome is emerging as a crucial player of the immune checkpoint in cancer. Melanoma is a highly immunogenic tumour, and the composition of the gut microbiome has been correlated to prognosis and evolution of advanced melanoma and proposed as a biomarker for immune checkpoint therapy., Objectives: We investigated the gut fungal and bacterial compositions in early-stage melanoma and correlated microbial profiles with histopathological features., Methods: Sequencing of bacterial 16S rRNA and the fungal internal transcribed spacer region was performed on faecal samples of patients with stage I and II melanoma, and healthy controls. A meta-analysis with gut microbiota data from patients with metastatic melanoma was also carried out., Results: We found a combination of gut fungal and bacterial profiles significantly discriminating patients with melanoma from controls. In patients with melanoma, we observed an abundance of Prevotella copri and yeasts belonging to the order Saccharomycetales. We found that the bacterial and fungal community correlated to melanoma invasiveness, whereas the specific fungal profile correlated to melanoma regression. Bacteroides was identified as general marker of immunogenicity, being shared by regressive and invasive melanoma. In addition, the bacterial communities in patients with stage I and II melanoma were different in structure and richer than those from patients with metastatic melanoma., Conclusions: The composition of the gut microbiota in early-stage melanoma changes along the gradient from in situ to invasive (and metastatic) melanoma. Changes in the microbiota and mycobiota are correlated to the histological features of early-stage melanoma, and to the clinical course and response to immune therapies of advanced-stage melanoma, through direct or indirect immunomodulation., (© 2021 The Authors. British Journal of Dermatology published by John Wiley & Sons Ltd on behalf of British Association of Dermatologists.)

The gut microbiota may modulate the disposition and toxicity of environmental contaminants within a host but, conversely, contaminants may also impact gut bacteria. Such contaminant-gut microbial connections, which could lead to alteration of host health, remain poorly known and are rarely studied in free-ranging wildlife. The polar bear (Ursus maritimus) is a long-lived, wide-ranging apex predator that feeds on a variety of high trophic position seal and cetacean species and, as such, is exposed to among the highest levels of biomagnifying contaminants of all Arctic species. Here, we investigate associations between mercury (THg; a key Arctic contaminant), diet, and the diversity and composition of the gut microbiota of polar bears inhabiting the southern Beaufort Sea, while accounting for host sex, age class and body condition. Bacterial diversity was negatively associated with seal consumption and mercury, a pattern seen for both Shannon and Inverse Simpson alpha diversity indices (adjusted R 2  = 0.35, F 1,18  = 8.00, P = 0.013 and adjusted R 2  = 0.26, F 1,18  = 6.04, P = 0.027, respectively). No association was found with sex, age class or body condition of polar bears. Bacteria known to either be involved in THg methylation or considered to be highly contaminant resistant, including Lactobacillales, Bacillales and Aeromonadales, were significantly more abundant in individuals that had higher THg concentrations. Conversely, individuals with higher THg concentrations showed a significantly lower abundance of Bacteroidales, a bacterial order that typically plays an important role in supporting host immune function by stimulating intraepithelial lymphocytes within the epithelial barrier. These associations between diet-acquired mercury and microbiota illustrate a potentially overlooked outcome of mercury accumulation in polar bears., (© 2021. The Author(s).)

Wood pellets can sustain the growth of Trichoderma spp. in soil; however, little is known about their side effects on the microbiota. The aims of this study were to evaluate the effect of wood pellets on the growth of Trichoderma spp. in bulk soil and on the soil microbial population's composition and diversity. Trichoderma atroviride SC1 coated wood pellets and non-coated pellets were applied at the level of 10 g∙kg -1 of soil and at the final concentration of 5 × 10 3 conidia∙g -1 of soil and compared to a conidial suspension applied at the same concentration without the wood carrier. Untreated bulk soil served as a control. The non-coated wood pellets increased the total Trichoderma spp. population throughout the experiment (estimated as colony-forming unit g -1 of soil), while wood pellets coated with T. &nbsp; atroviride SC1 did not. The wood carrier increased the richness, and temporarily decreased the diversity, of the bacterial population, with Massilia being the most abundant bacterial genus, while it decreased both the richness and diversity of the fungal community. Wood pellets selectively increased fungal species having biocontrol potential, such as Mortierella , Cladorrhinum, and Stachybotrys , which confirms the suitability of such carriers of Trichoderma spp. for soil application.

The microbial communities associated to the rhizosphere (the rhizomicrobiome) have a substantial impact on plant growth and yield. Understanding the effects of agricultural management on the rhizomicrobiome is very important for selecting efficient practices. By sequencing the V4 region of 16S rRNA for bacteria and the ITS1 regions and fungi, we investigated the influences of agronomic practices, including cucumber grafting on cucurbit hybrid (Cucurbita moschata × C. maxima), cucumber-garlic intercropping, and treatment with fungicide iprodione-carbendazim on cucumber rhizosphere microbial communities during plant growth. Soil dehydrogenase activity (DHA) and plant vegetative parameters were assessed as an indicator of overall soil microbial activity. We found that both treatments and growth stage induced significant shifts in microbial community structure. Grafting had the highest number of differentially abundant OTUs compared to control samples, followed by intercropping and fungicide treatment, while plant development stage affected both alpha and beta diversities indices and composition of the rhizomicrobiome. DHA was more dependent on plant growth stages than on treatments. Among the assessed factors, grafting and plant developmental stage resulted in the greatest changes in the microbial community composition. Grafting also increased the plant growth parameters, suggesting that this method should be further investigated in vegetable production systems., (© The Author(s) 2021. Published by Oxford University Press on behalf of FEMS. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Current biodiversity loss is a major concern and thus biodiversity assessment of modern ecosystems is compelling and needs to be contextualized on a longer timescale. High Throughput Sequencing (HTS) is progressively becoming a major source of data on biodiversity time series. In this multi proxy study, we tested, for the first time, the potential of HTS to estimate plant biodiversity archived in the surface layers of a temperate alpine glacier, amplifying the trnL barcode for vascular plants from eDNA of firn samples. A 573 cm long core was drilled by the Adamello glacier and cut into sections; produced samples were analyzed for physical properties, stable isotope ratio, and plant biodiversity by eDNA metabarcoding and conventional light microscopy analysis. Results highlighted the presence of pollen and plant remains within the distinct layers of snow, firn and ice. While stable isotope ratio showed a scarcely informative pattern, DNA metabarcoding described distinct plant species composition among the different samples, with a broad taxonomic representation of the biodiversity of the catchment area and a high-ranking resolution. New knowledge on climate and plant biodiversity changes of large catchment areas can be obtained by this novel approach, relevant for future estimates of climate change effects.

Diet selection by grazing livestock may affect animal performance as well as the biodiversity of grazed areas. Recent DNA barcoding techniques allow to assess dietary plant composition in faecal samples, which may be additionally integrated by the description of gut microbiota. In this high throughput metabarcoding study, we investigated the diversity of plant, fungal and bacterial taxa in faecal samples of lactating cows of two breeds grazing an Alpine semi-natural grassland during summer. The estimated plant composition of the diet comprised 67 genera and 39 species, which varied remarkably during summer, suggesting a decline of the diet forage value with the advancing of the vegetative season. The fungal community included Neocallimastigomycota gut symbionts, but also Ascomycota and Basidiomycota plant parasite and coprophilous taxa, likely ingested during grazing. The proportion of ingested fungi was remarkably higher than in other studies, and varied during summer, although less than that observed for plants. Some variation related to breed was also detected. The gut bacterial taxa remained stable through the summer but displayed a breed-specific composition. The study provided insights in the reciprocal organisms' interactions affecting, and being affected by, the foraging behaviour: plants showed a high temporal variation, fungi a smaller one, while bacteria had practically none; conversely, the same kingdoms showed the opposite gradient of variation as respect to the animal host breed, as bacteria revealed to be the group mostly characterized by host-specificity.

The factors that influence the diversity and composition of raw milk and fecal microbiota in healthy commercial dairy herds are not fully understood, partially because the majority of metataxonomic studies involve experimental farms and/or single factors. We analyzed the raw milk and fecal microbiota of 100 healthy cows from 10 commercial alpine farms from the Province of Trento, Italy, using metataxonomics and applied statistical modelling to investigate which extrinsic and intrinsic parameters (e.g. herd, diet and milk characteristics) correlated with microbiota richness and composition in these relatively small traditional farms. We confirmed that Firmicutes, Ruminococcaceae and Lachnospiraceae families dominated the fecal and milk samples of these dairy cows, but in addition, we found an association between the number of observed OTUs and Shannon entropy on each farm that indicates higher microbiota richness is associated with increased microbiota stability. Modelling showed that herd was the most significant factor affecting the variation in both milk and fecal microbiota composition. Furthermore, the most important predictors explaining the variation of microbiota richness were milk characteristics (i.e. percentage fat) and diet for milk and fecal samples, respectively. We discuss how high intra-herd variation could affect the development of treatments based on microbiota manipulation., Competing Interests: The authors have declared that no competing interests exist.

Plasmopara viticola is the causal agent of grapevine downy mildew (DM). DM resistant varieties deploy effector-triggered immunity (ETI) to inhibit pathogen growth, which is activated by major resistance loci, the most common of which are Rpv3 and Rpv12. We previously showed that a quick metabolome response lies behind the ETI conferred by Rpv3 TIR-NB-LRR genes. Here we used a grape variety operating Rpv12-mediated ETI, which is conferred by an independent locus containing CC-NB-LRR genes, to investigate the defence response using GC/MS, UPLC, UHPLC and RNA-Seq analyses. Eighty-eight metabolites showed significantly different concentration and 432 genes showed differential expression between inoculated resistant leaves and controls. Most metabolite changes in sugars, fatty acids and phenols were similar in timing and direction to those observed in Rpv3-mediated ETI but some of them were stronger or more persistent. Activators, elicitors and signal transducers for the formation of reactive oxygen species were early observed in samples undergoing Rpv12-mediated ETI and were paralleled and followed by the upregulation of genes belonging to ontology categories associated with salicylic acid signalling, signal transduction, WRKY transcription factors and synthesis of PR-1, PR-2, PR-5 pathogenesis-related proteins.

Mountain glacier shrinkage represents a major effect of the current global warming and 80-100% of the Alpine glaciers are predicted to vanish within the next few decades. As the thawing rate of mountain permafrost ice is much lower than for glacier ice, a shift from glacial to periglacial dynamics is predicted for Alpine landscapes during the 21st century. Despite the growing literature on the impacts of deglaciation on Alpine hydrology and ecosystems, chemical and biological features of waters emerging from Alpine rock glaciers (i.e. permafrost landforms composed by a mixture of ice and debris) have been poorly investigated so far, and knowledge on microbial biodiversity of headwaters is still sparse. A set of glacier-, rock glacier- and groundwater/precipitation-fed streams was investigated in the Italian Central Alps in late summer 2016, aiming at exploring bacterial community composition and diversity in epilithic and surface sediment biofilm and at verifying the hypothesis that rock glacier-fed headwaters represent peculiar ecosystems from both a chemical and biological point of view. Rock glacier-fed waters showed high values of electrical conductivity and trace elements related to their bedrock lithology, and their highly diverse bacterial assemblages significantly differed from those detected in glacier-fed streams. Bacterial taxonomic composition appeared to be mainly related to water and substrate type, as well as to water chemistry, the latter including concentrations of nutrients and trace metals. The results of this study confirm the chemical and biological peculiarity of rock glacier-fed waters compared to glacial waters, and suggest a potential driving role of thawing permafrost in modulating future ecological traits of Alpine headwaters within the context of progressing deglaciation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The structure of microbial communities, microalgae, heterotrophic protozoa and fungi contributes to characterize food webs and productivity and, from an anthropogenic point of view, the qualitative characteristics of water bodies. Traditionally, in freshwater environments many investigations have been directed to the study of pelagic microalgae ("phytoplankton") and periphyton (i.e., photosynthetic and mixotrophic protists) through the use of light microscopy (LM). While the number of studies on bacterioplankton communities have shown a substantial increase after the advent of high-throughput sequencing (HTS) approaches, the study of the composition, structure, and spatio-temporal patterns of microbial eukaryotes in freshwater environments was much less widespread. Moreover, the understanding of the correspondence between the relative phytoplankton abundances estimated by HTS and LM is still incomplete. Taking into account these limitations, this study examined the biodiversity and seasonality of the community of eukaryotic microplankton in the epilimnetic layer of a large and deep perialpine lake (Lake Garda) using HTS. The analyses were carried out at monthly frequency during 2014 and 2015. The results highlighted the existence of a rich and well diversified community and the presence of numerous phytoplankton taxa that were never identified by LM in previous investigations. Furthermore, the relative abundances of phytoplankton estimated by HTS and LM showed a significant relationship at different taxonomic ranks. In the 2 years of investigation, the temporal development of the whole micro-eukaryotic community showed a clear non-random and comparable distribution pattern, with the main taxonomic groups coherently distributed in the individual seasons. In perspective, the results obtained in this study highlight the importance of HTS approaches in assessing biodiversity and the relative importance of the main protist groups along environmental gradients, including those caused by anthropogenic impacts (e.g., eutrophication and climate change)., (Copyright © 2020 Salmaso, Boscaini and Pindo.)

'Glera' and 'Ribolla Gialla' are the most economically relevant local grapevine cultivars of Friuli Venezia Giulia region (north-eastern Italy). 'Glera' is used to produce the world-renowned Prosecco wine. 'Ribolla Gialla' cultivation is constantly increasing due to the strong demand for sparkling wine and is the most important variety in Brda (Slovenia). Knowledge of local varieties history in terms of migration and pedigree relationships has scientific and marketing appeal. Following prospections, genotyping and ampelographic characterization of minor germplasm in Friuli Venezia Giulia, a further research was developed to understand the parentage relationships among the grapevine varieties grown in this region. An integrated strategy was followed combining the analysis of nuclear and chloroplast microsatellites with the Vitis 18k SNP chip. Two main recurrent parents were found, which can be regarded as "founders": 'Vulpea', an Austrian variety parent-offspring related with at least ten Friuli Venezia Giulia cultivars, among them 'Glera', and 'Refosco Nostrano', first degree related with other six Friuli Venezia Giulia varieties. 'Ribolla Gialla' was shown to be another member of the impressively long list of offspring derived from the prolific 'Heunisch Weiss'. Combining molecular markers and historical references was a high-performance strategy for retracing and adjusting the history of cultivars.

The bacterium Erwinia amylovora, the causal agent of fire blight disease in apple, triggers its infection through the DspA/E effector which interacts with the apple susceptibility protein MdDIPM4. In this work, MdDIPM4 knockout has been produced in two Malus × domestica susceptible cultivars using the CRISPR/Cas9 system delivered via Agrobacterium tumefaciens. Fifty-seven transgenic lines were screened to identify CRISPR/Cas9-induced mutations. An editing efficiency of 75% was obtained. Seven edited lines with a loss-of-function mutation were inoculated with the pathogen. Highly significant reduction in susceptibility was observed compared to control plants. Sequencing of five potential off-target sites revealed no mutation event. Moreover, our construct contained a heat-shock inducible FLP/FRT recombination system designed specifically to remove the T-DNA harbouring the expression cassettes for CRISPR/Cas9, the marker gene and the FLP itself. Six plant lines with reduced susceptibility to the pathogen were heat-treated and screened by real-time PCR to quantify the exogenous DNA elimination. The T-DNA removal was further validated by sequencing in one plant line. To our knowledge, this work demonstrates for the first time the development and application of a CRISPR/Cas9-FLP/FRT gene editing system for the production of edited apple plants carrying a minimal trace of exogenous DNA., (© 2019 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Mgaloblishvili, a Vitis vinifera cultivar, exhibits unique resistance traits against Plasmopara viticola , the downy mildew agent. This offers the unique opportunity of exploring the molecular responses in compatible and incompatible plant-pathogen interaction. In this study, whole transcriptomes of Mgaloblishvili, Pinot noir (a V. vinifera susceptible cultivar), and Bianca (a resistant hybrid) leaves, inoculated and non-inoculated with the pathogen , were used to identify P. viticola effector-encoding genes and plant susceptibility/resistance genes. Multiple effector-encoding genes were identified in P. viticola transcriptome, with remarkable expression differences in relation to the inoculated grapevine cultivar. Intriguingly, five apoplastic effectors specifically associated with resistance in V. vinifera . Gene coexpression network analysis identified specific modules and metabolic changes occurring during infection in the three grapevine cultivars. Analysis of these data allowed, for the first time, the detection in V. vinifera of a putative P. viticola susceptibility gene, encoding a LOB domain-containing protein. Finally, the de novo assembly of Mgaloblishvili, Pinot noir, and Bianca transcriptomes and their comparison highlighted novel candidate genes that might be at the basis of the resistant phenotype. These results open the way to functional analysis studies and to new perspectives in molecular breeding of grapevine for resistance to P. viticola ., Competing Interests: The authors declare no conflict of interest.

Since it has been understood that gut microbiota of vector mosquitoes can influence their vector competence, efforts have been undertaken to develop new control strategies based on host microbiota manipulation, and aimed at suppressing the vector population or replacing it with a less competent one. For the proper design of such control strategies it is necessary to know the microbiota composition of the target vector species, how it is acquired, and how it changes throughout the host's life cycle. In this study, 16S rRNA amplicon high-throughput sequencing was used to characterize the changes in microbiota from the aquatic environment (larval breeding sites) to the different developmental stages of field-collected Aedes koreicus in Italy, an emerging invasive mosquito species in Europe and a potential vector of several pathogens. The bacterial communities of the aquatic breeding sites, larvae, pupae and adults showed distinctive structures to one another. Indeed, 84% of community members were unique to a given sample type. Nevertheless, almost 40% of the sequences generated were assigned to bacteria detected in all sample types, suggesting the importance of bacteria transstadially transmitted from water to the adult stage in constituting mosquito microbiota. Among these, genus C39 largely constituted water microbiota, family Burkholderiaceae was the most abundant in larvae and pupae, and genus Asaia dominated adult communities. In addition, Asaia constituted a core microbiota across all sample types. Our results suggest that the microbiota of Ae. koreicus mosquitoes is composed by a community which derives from the aquatic bacteria of the larval breeding sites, is then filtered by the larval gut, where only certain members are able to persist, rearranged by metamorphosis and finally modified by the change in diet at the adult stage. Understanding how the microbiota of Ae. koreicus changes through the mosquito life cycle represents a first step in selecting bacterial candidates for use in microbiota-based intervention measures for this species. The properties which Asaia exhibits in this species, such as dominance, high prevalence and transstadial transmission, prevent the use of Wolbachia but make Asaia an ideal candidate for paratransgenesis., (Copyright © 2019 Alfano, Tagliapietra, Rosso, Manica, Arnoldi, Pindo and Rizzoli.)

The gut microbiota plays a critical role in host health, yet remains poorly studied in wild species. Polar bears (Ursus maritimus), key indicators of Arctic ecosystem health and environmental change, are currently affected by rapid shifts in habitat that may alter gut homeostasis. Declining sea ice has led to a divide in the southern Beaufort Sea polar bear subpopulation such that an increasing proportion of individuals now inhabit onshore coastal regions during the open-water period ('onshore bears') while others continue to exhibit their typical behaviour of remaining on the ice ('offshore bears'). We propose that bears that have altered their habitat selection in response to climate change will exhibit a distinct gut microbiota diversity and composition, which may ultimately have important consequences for their health. Here, we perform the first assessment of abundance and diversity in the faecal microbiota of wild polar bears using 16S rRNA Illumina technology. We find that bacterial diversity is significantly higher in onshore bears compared to offshore bears. The most enriched OTU abundance in onshore bears belonged to the phylum Proteobacteria, while the most depleted OTU abundance within onshore bears was seen in the phylum Firmicutes. We conclude that climate-driven changes in polar bear land use are associated with distinct microbial communities. In doing so, we present the first case of global change mediated alterations in the gut microbiota of a free-roaming wild animal.

Background: Despite their importance as a reservoir of biodiversity, the factors shaping soil microbial communities and the extent by which these are impacted by cultivation are still poorly understood. Using 16S rRNA gene and ITS sequencing, we characterized the soil microbiota of vineyards and of neighboring permanent grassland soils in the Italian province of Trentino, and correlated their structure and composition to location, chemical properties of the soil, and land management., Results: Bacterial communities had a core of conserved taxa accounting for more than 60% of the reads of each sample, that was influenced both by geography and cultivation. The core fungal microbiota was much smaller and dominated by geography alone. Cultivation altered the structure and composition of the soil microbiota both for bacteria and fungi, with site-specific effects on their diversity. The diversity of bacterial and fungal communities was generally inversely correlated across locations. We identified several taxa that were impacted by the chemical properties and texture of the soil., Conclusions: Our results highlight the different responses of bacterial and fungal communities to environmental factors and highlight the need to characterize both components of the soil microbiota to fully understand the factors that drive their variability.

Community composition of freshwater prokaryotes and protists varies through time. Few studies contemporarily investigate temporal variation of these freshwater communities for more than 1 year. We compared the temporal patterns of prokaryotes and protists in three distinct habitats for 4 years (2014-2017) in Lake Tovel, a cold-water lake. This lake showed a marked temperature increase in 2017 linked to altered precipitation patterns. We investigated whether microbial communities reflected this change across habitats and whether changes occurred at the same time and to the same extent. Furthermore, we tested the concept of hydrological year emphasizing the ecological effect of water renewal on communities for its explanatory power of community changes. Microbe diversity was assessed by Illumina sequencing of the V3-V4 hypervariable region of the 16S rRNA gene and 18S rRNA gene, and we applied co-inertia analysis and asymmetric eigenvector maps modelling to infer synchrony and temporal patterns of prokaryotes and protists. When considering community composition, microbes were invariable in synchrony across habitats and indicated a temporal gradient linked to decreasing precipitation; however, when looking at temporal patterns, the extent of synchrony was reduced. Small-scale patterns were similar across habitats and microbes and linked to seasonally varying environmental variables, while large-scale patterns were different and partially linked to an ecosystem change as indicated by increasing water transparency and temperature and decreasing dissolved oxygen. Our advanced statistical approach outlined the multifaceted aspect of synchrony when linked to community composition and temporal patterns., (© 2019 John Wiley & Sons Ltd.)

Purpose: Olive pomace is a major waste product of olive oil production but remains rich in polyphenols and fibres. We measured the potential of an olive pomace-enriched biscuit formulation delivering 17.1 ± 4.01 mg/100 g of hydroxytyrosol and its derivatives, to modulate the composition and metabolic activity of the human gut microbiota., Methods: In a double-blind, controlled parallel dietary intervention 62 otherwise healthy hypercholesterolemic (total plasma cholesterol 180-240 mg/dl) subjects were randomly assigned to eat 90 g of olive pomace-enriched biscuit (olive-enriched product, OEP) or an isoenergetic control (CTRL) for 8 weeks. Fasted blood samples, 24-h urine and faecal samples were collected before and after dietary intervention for measurement of microbiota, metabolites and clinical parameters., Results: Consumption of OEP biscuits did not impact on the diversity of the faecal microbiota and there was no statistically significant effect on CVD markers. A trend towards reduced oxidized LDL cholesterol following OEP ingestion was observed. At the genus level lactobacilli and Ruminococcus were reduced in OEP compared to CTRL biscuits. A trend towards increased bifidobacteria abundance was observed after OEP ingestion in 16S rRNA profiles, by fluorescent in situ hybridization and by qPCR. Targeted LC-MS revealed significant increases phenolic acid concentrations in 24-h urine following OEP ingestion and 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid, derivatives of hydroxytyrosol, were elevated in blood. A sex effect was apparent in urine small phenolic acid concentrations, and this sex effect was mirrored by statistically significant differences in relative abundances of faecal bacteria between men and women., Conclusion: Ingestion of OEP biscuits led to a significant increase in the metabolic output of the gut microbiota with an apparent sex effect possibly linked to differences in microbiota makeup. Increased levels of homovanillic acid and DOPAC, thought to be involved in reducing oxidative LDL cholesterol, were observed upon OEP ingestion. However, OEP did not induce statistically significant changes in either ox-LDL or urinary isoprostane in this study.

Temporal dynamics of bacterioplankton are rarely investigated for multiple habitats and years within individual lakes, limiting our understanding of the variability of bacterioplankton community (BC) composition with respect to environmental factors. We assessed the BC composition of a littoral and two pelagic habitats (euphotic zone and hypolimnion) of Lake Tovel monthly from April 2014 to May 2017 by high-throughput sequencing of the V3-V4 hypervariable region of the 16S rRNA gene. The three habitats differed in temperature, light, oxygen and hydrology. In particular, the littoral was the most hydrologically unstable because it receives most of the lake inflow, the hypolimnion was the most stable because of its hydrologically sheltered position, and the pelagic euphotic habitat was intermediate. Consequently, we hypothesized different temporal patterns of BC composition for all three habitats according to their environmental differences. We applied PERMANOVA, nonmetric multidimensional scaling and source-sink analysis to characterize BC composition. Overall, BCs were different among habitats with the littoral showing the highest variability and the hypolimnion the highest stability. The BC of rainy 2014 was distinct from the BCs of other years irrespective of the habitats considered. Seasonal differences in BCs were limited to spring, probably linked to meltwater inflow and mixing. Thus, temporal effects related to year and season were linked to the hydrological gradient of habitats. We suggest that despite potential within-lake dispersal of bacterioplankton by water flow and mixing, local environmental conditions played a major role in Lake Tovel, fostering distinct BCs in the three habitats., (© 2018 John Wiley & Sons Ltd.)

Aedes mosquitoes are considered highly successful global invasive species and vectors of several pathogens of relevance for public health. Their midgut's microbiota can play an important role in affecting not only their vectorial competence but also their fitness, physiology, food digestion, metabolism, immunity and adaptation to new environmental conditions. Using high-throughput sequencing we compared the microbiota of Aedes albopictus collected in Italy with those reported in populations from France and Vietnam. We also analysed Aedes koreicus gut microbiota for the first time. We found remarkable individual difference along with common bacterial taxa in both species. Ae. albopictus collected in Italy had a lower richness and a different composition of microbiota in respect to specimens collected in France and Vietnam. It also showed a core microbiota formed mainly of bacteria of the genus Pseudomonas. Overall, the two Aedes species (Ae. albopictus and Ae. koreicus) collected in Italy, showed a large core microbiota with 75.98% of the identified Operational Taxonomic Units. Furthermore, Ae. albopictus had 2.5% prevalence of Wolbachia and 0.07% of Asaia spp, while Ae. koreicus had 14.42% of Asaia spp. and no Wolbachia. This study provides new informations on the spatial variation of the midgut bacterial communities in mosquitoes of medical relevance within areas of recent invasion and provide the basis for further studies aimed at assessing the effects of such variation on vectorial capacity for a range of pathogens.

Nucleotide-binding Oligomerization Domain-2 (NOD2) mutations are associated with an increased risk to develop Crohn's Disease. In previous studies, we have shown that Nod2-/- mice manifest increased proportion of Lamina Propria (LP) CD4+ LAP+ Foxp3- regulatory cells, when compared with Nod2+/+ mice, while CD4+ Foxp3 + regulatory cells were not affected. Here, we investigated the Nod2 gut microbiota, by 16S rRNA pyrosequencing, at steady state and after TNBS-colitis induction in mice reared separately or in cohousing, correlating the microbial profiles with LP regulatory T cells proportion and tissue cytokines content. We found that enrichment of Rikenella and Alistipes (Rikenellaceae) in Nod2-/- mice at 8 weeks of age reared separately was associated with increased proportion of CD4+ LAP+ Foxp3- cells and less severe TNBS-colitis. In co-housed mice the acquisition of Rickenellaceae by Nod2+/+ mice was associated with increased CD4+ LAP+ Foxp3- proportion and less severe colitis. Severe colitis was associated with enrichment of gram-negative pathobionts (Escherichia and Enterococcus), while less severe colitis with protective bacteria (Barnesiella, Odoribacter and Clostridium IV). Environmental factors acting on genetic background with different outcomes according to their impact on microbiota, predispose in different ways to inflammation. These results open a new scenario for therapeutic attempt to re-establish eubiosis in Inflammatory Bowel Disease patients with NOD2 polymorphisms.

The Eurasian grapevine (Vitis vinifera), an Old World species now cultivated worldwide for high-quality wine production, is extremely susceptible to the agent of downy mildew, Plasmopara viticola. The cultivation of resistant V. vinifera varieties would be a sustainable way to reduce the damage caused by the pathogen and the impact of disease management, which involves the economic, health and environmental costs of frequent fungicide application. We report the finding of unique downy mildew resistance traits in a winemaking cultivar from the domestication center of V. vinifera, and characterize the expression of a range of genes associated with the resistance mechanism. Based on comparative experimental inoculations, confocal microscopy and transcriptomics analyses, our study shows that V. vinifera cv. Mgaloblishvili, native to Georgia (South Caucasus), exhibits unique resistance traits against P. viticola. Its defense response, leading to a limitation of P. viticola growth and sporulation, is determined by the overexpression of genes related to pathogen recognition, the ethylene signaling pathway, synthesis of antimicrobial compounds and enzymes, and the development of structural barriers. The unique resistant traits found in Mgaloblishvili highlight the presence of a rare defense system in V. vinifera against P. viticola which promises fresh opportunities for grapevine genetic improvement.

High-throughput sequencing (HTS) was used to analyze the seasonal variations in the bacterioplankton community composition (BCC) in the euphotic layer of a large and deep lake south of the Alps (Lake Garda). The BCC was analyzed throughout two annual cycles by monthly samplings using the amplification and sequencing of the V3-V4 hypervariable region of the 16S rRNA gene by the MiSeq Illumina platform. The dominant and most diverse bacterioplankton phyla were among the more frequently reported in freshwater ecosystems, including the Proteobacteria, Cyanobacteria, Bacteroidetes, Verrucomicrobia, Actinobacteria, and Planctomycetes. As a distinctive feature, the development of the BCC showed a cyclical temporal pattern in the two analyzed years and throughout the euphotic layer. The recurring temporal development was controlled by the strong seasonality in water temperature and thermal stratification, and by cyclical temporal changes in nutrients and, possibly, by the remarkable annual cyclical development of cyanobacteria and eukaryotic phytoplankton hosting bacterioplankton that characterizes Lake Garda. Further downstream analyses of operational taxonomic units associated to cyanobacteria allowed confirming the presence of the most abundant taxa previously identified by microscopy and/or phylogenetic analyses, as well as the presence of other small Synechococcales/Chroococcales and rare Nostocales never identified so far in the deep lakes south of the Alps. The implications of the high diversity and strong seasonality are relevant, opening perspectives for the definition of common and discriminating patterns characterizing the temporal and spatial distribution in the BCC, and for the application of the new sequencing technologies in the monitoring of water quality in large and deep lakes.

Plant tissues host complex fungal and bacterial communities, and their composition is determined by host traits such as tissue age, plant genotype and environmental conditions. Despite the importance of bark as a possible reservoir of plant pathogenic microorganisms, little is known about the associated microbial communities. In this work, we evaluated the composition of fungal and bacterial communities in the pear (Abate and Williams cultivars) and apple (Golden Delicious and Gala cultivars) bark of three/four-year-old shoots (old bark) or one-year-old shoots (young bark), using a meta-barcoding approach. The results showed that both fungal and bacterial communities are dominated by genera with ubiquitous attitudes, such as Aureobasidium, Cryptococcus, Deinococcus and Hymenobacter, indicating intense microbial migration to surrounding environments. The shoot age, plant species and plant cultivar influenced the composition of bark fungal and bacterial communities. In particular, bark communities included potential biocontrol agents that could maintain an equilibrium with potential plant pathogens. The abundance of fungal (e.g. Alternaria, Penicillium, Rosellinia, Stemphylium and Taphrina) and bacterial (e.g. Curtobacterium and Pseudomonas) plant pathogens was affected by bark age and host genotype, as well as those of fungal genera (e.g. Arthrinium, Aureobasidium, Rhodotorula, Sporobolomyces) and bacterial genera (e.g. Bacillus, Brevibacillus, Methylobacterium, Sphingomonas and Stenotrophomonas) with possible biocontrol and plant growth promotion properties., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

Developing fish farming to meet the demands of food security and sustainability in the 21st century will require new farming systems and improved feeds. Diet and microbe interactions in the gut is an important variable with the potential to make a significant impact on future fish farming diets and production systems. It was monitored the gut microbiota of farmed rainbow trout using 16S rRNA profiling over 51 weeks during standard rearing conditions and feeding diet with supplementation of an essential oils (MixOil) mixture from plants (at a concentration in diet of 200 mg/kg). Gut microbiota 16S rRNA profiling indicated that the fish gut was dominated by Actinobacteria, Proteobacteria, Bacteroidetes and Firmicutes. Although the dietary supplementation with MixOil had no impact on either the composition or architecture of gut microbiota, significant changes in alpha and beta diversity and relative abundance of groups of gut bacteria were evident during growth stages on test feeds, especially upon prolonged growth on finishing feed. Fish fillet quality to guarantee palatability and safety for human consumption was also evaluated. Significant differences within the gut microbiota of juvenile and adult trout under the same rearing conditions were observed, The addition of essential oil blend affected some physicochemical characteristics of trout fillets, including their resistance to oxidative damage and their weight loss (as liquid loss and water holding capacity) during the first period of storage, that are two important parameters related to product shelf life and susceptibility to spoilage. The results highlighted the need for further studies concern dietary microbiome modulation at different life stages and its influence on animal health, growth performance and final product quality.

Main Conclusion: A coordinated regulation of different branches of the flavonoid pathway was highlighted that may contribute to elucidate the role of this important class of compounds during the early stages of apple fruit development. Apple (Malus × domestica Borkh.) is an economically important fruit appreciated for its organoleptic characteristics and its benefits for human health. The first stages after fruit set represent a very important and still poorly characterized developmental process. To enable the profiling of genes involved in apple early fruit development, we combined the suppression subtractive hybridization (SSH) protocol to next-generation sequencing. We identified and characterized genes induced and repressed during fruit development in the apple cultivar 'Golden Delicious'. Our results showed an opposite regulation of genes coding for enzymes belonging to flavonoid and monolignol pathways, with a strong induction of the former and a simultaneous repression of the latter. Two isoforms of phenylalanine ammonia-lyase and 4-coumarate:CoA ligase, key enzymes located at the branching point between flavonoid and monolignol pathways, showed opposite expression patterns during the period in analysis, suggesting a possible regulation mechanism. A targeted metabolomic analysis supported the SSH results and revealed an accumulation of the monomers catechin and epicatechin as well as several forms of procyanidin oligomers in apple fruitlets starting early after anthesis, together with a decreased production of other classes of flavonoids such as some flavonols and the dihydrochalcone phlorizin. Moreover, gene expression and metabolites accumulation of 'Golden Delicious' were compared to a wild apple genotype of Manchurian crabapple (Malus mandshurica (Maxim.) Kom.). Significant differences in both gene expression and metabolites accumulation were found between the two genotypes.

Background: A critical aspect regarding the global dispersion of pathogenic microorganisms is associated with atmospheric movement of soil particles. Especially, desert dust storms can transport alien microorganisms over continental scales and can deposit them in sensitive sink habitats. In winter 2014, the largest ever recorded Saharan dust event in Italy was efficiently deposited on the Dolomite Alps and was sealed between dust-free snow. This provided us the unique opportunity to overcome difficulties in separating dust associated from "domestic" microbes and thus, to determine with high precision microorganisms transported exclusively by desert dust., Results: Our metagenomic analysis revealed that sandstorms can move not only fractions but rather large parts of entire microbial communities far away from their area of origin and that this microbiota contains several of the most stress-resistant organisms on Earth, including highly destructive fungal and bacterial pathogens. In particular, we provide first evidence that winter-occurring dust depositions can favor a rapid microbial contamination of sensitive sink habitats after snowmelt., Conclusions: Airborne microbial depositions accompanying extreme meteorological events represent a realistic threat for ecosystem and public health. Therefore, monitoring the spread and persistence of storm-travelling alien microbes is a priority while considering future trajectories of climatic anomalies as well as anthropogenically driven changes in land use in the source regions.

Alteration of gut microbiota is involved in several chronic inflammatory and autoimmune diseases, including rheumatoid arthritis, and gut microbial "pro-arthritogenic" profiles have been hypothesized. Intestinal inflammation may be involved in spondyloarthropathies and in a subset of patients affected by Juvenile Idiopathic Arthritis (JIA), the most common chronic rheumatic disease of childhood. We compared the fecal microbiota composition of JIA patients with healthy subjects (HS), evaluating differences in microbial profiles between sub-categories of JIA, such as enthesitis-related arthritis (JIA-ERA), in which inflammation of entheses occurs, and polyarticular JIA, non-enthesitis related arthritis (JIA-nERA). Through taxon-level analysis, we discovered alteration of fecal microbiota components that could be involved in subclinical gut inflammation, and promotion of joint inflammation. We observed abundance in Ruminococcaceae in both JIA categories, reduction in Clostridiaceae and Peptostreptococcaceae in JIA-ERA, and increase in Veillonellaceae in JIA-nERA, respectively, compared with HS. Among the more relevant genera, we found an increase in Clostridium cluster XIVb , involved in colitis and arthritis, in JIA-ERA patients compared with HS, and a trend of decrease in Faecalibacterium , known for anti-inflammatory properties, in JIA-nERA compared with JIA-ERA and HS. Differential abundant taxa identified JIA patients for the HLA-B27 allele, including Bilophila, Clostridium cluster XIVb, Oscillibacter , and Parvimonas . Prediction analysis of metabolic functions showed that JIA-ERA metagenome was differentially enriched in bacterial functions related to cell motility and chemotaxis, suggesting selection of potential virulence traits. We also discovered differential microbial profiles and intra-group variability among active disease and remission, suggesting instability of microbial ecosystem in autoimmune diseases with respect to healthy status. Similarly to other chronic autoimmune and inflammatory diseases, different microbial profiles, as observed among different JIA subgroups compared to HS, and potential functional acquisition related to migration, could promote inflammation and contribute to the disease pathogenesis.

Background: The human gut microbiota directly affects human health, and its alteration can lead to gastrointestinal abnormalities and inflammation. Rett syndrome (RTT), a progressive neurological disorder mainly caused by mutations in MeCP2 gene, is commonly associated with gastrointestinal dysfunctions and constipation, suggesting a link between RTT's gastrointestinal abnormalities and the gut microbiota. The aim of this study was to evaluate the bacterial and fungal gut microbiota in a cohort of RTT subjects integrating clinical, metabolomics and metagenomics data to understand if changes in the gut microbiota of RTT subjects could be associated with gastrointestinal abnormalities and inflammatory status., Results: Our findings revealed the occurrence of an intestinal sub-inflammatory status in RTT subjects as measured by the elevated values of faecal calprotectin and erythrocyte sedimentation rate. We showed that, overall, RTT subjects harbour bacterial and fungal microbiota altered in terms of relative abundances from those of healthy controls, with a reduced microbial richness and dominated by microbial taxa belonging to Bifidobacterium, several Clostridia (among which Anaerostipes, Clostridium XIVa, Clostridium XIVb) as well as Erysipelotrichaceae, Actinomyces, Lactobacillus, Enterococcus, Eggerthella, Escherichia/Shigella and the fungal genus Candida. We further observed that alterations of the gut microbiota do not depend on the constipation status of RTT subjects and that this dysbiotic microbiota produced altered short chain fatty acids profiles., Conclusions: We demonstrated for the first time that RTT is associated with a dysbiosis of both the bacterial and fungal component of the gut microbiota, suggesting that impairments of MeCP2 functioning favour the establishment of a microbial community adapted to the costive gastrointestinal niche of RTT subjects. The altered production of short chain fatty acids associated with this microbiota might reinforce the constipation status of RTT subjects and contribute to RTT gastrointestinal physiopathology.

Protein derivatives and carbohydrates can stimulate plant growth, increase stress tolerance, and activate plant defense mechanisms. However, these molecules can also act as a nutritional substrate for microbial communities living on the plant phyllosphere and possibly affect their biocontrol activity against pathogens. We investigated the mechanisms of action of a protein derivative (nutrient broth, NB) against grapevine downy mildew, specifically focusing on the effects of foliar treatments on plant defense stimulation and on the composition and biocontrol features of the phyllosphere microbial populations. NB reduced downy mildew symptoms and induced the expression of defense-related genes in greenhouse- and in vitro-grown plants, indicating the activation of grapevine resistance mechanisms. Furthermore, NB increased the number of culturable phyllosphere bacteria and altered the composition of bacterial and fungal populations on leaves of greenhouse-grown plants. Although, NB-induced changes on microbial populations were affected by the structure of indigenous communities originally residing on grapevine leaves, degrees of disease reduction and defense gene modulation were consistent among the experiments. Thus, modifications in the structure of phyllosphere populations caused by NB application could partially contribute to downy mildew control by competition for space or other biocontrol strategies. Particularly, changes in the abundance of phyllosphere microorganisms may provide a contribution to resistance induction, partially affecting the hormone-mediated signaling pathways involved. Modifying phyllosphere populations by increasing natural biocontrol agents with the application of selected nutritional factors can open new opportunities in terms of sustainable plant protection strategies.

Cultivated apple (Malus × domestica Borkh.) is one of the most important fruit crops in temperate regions, and has great economic and cultural value. The apple genome is highly heterozygous and has undergone a recent duplication which, combined with a rapid linkage disequilibrium decay, makes it difficult to perform genome-wide association (GWA) studies. Single nucleotide polymorphism arrays offer highly multiplexed assays at a relatively low cost per data point and can be a valid tool for the identification of the markers associated with traits of interest. Here, we describe the development and validation of a 487K SNP Affymetrix Axiom(®) genotyping array for apple and discuss its potential applications. The array has been built from the high-depth resequencing of 63 different cultivars covering most of the genetic diversity in cultivated apple. The SNPs were chosen by applying a focal points approach to enrich genic regions, but also to reach a uniform coverage of non-genic regions. A total of 1324 apple accessions, including the 92 progenies of two mapping populations, have been genotyped with the Axiom(®) Apple480K to assess the effectiveness of the array. A large majority of SNPs (359 994 or 74%) fell in the stringent class of poly high resolution polymorphisms. We also devised a filtering procedure to identify a subset of 275K very robust markers that can be safely used for germplasm surveys in apple. The Axiom(®) Apple480K has now been commercially released both for public and proprietary use and will likely be a reference tool for GWA studies in apple., (© 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.)

The expansion of agriculture is shrinking pristine forest areas worldwide, jeopardizing the persistence of their wild inhabitants. The Udzungwa red colobus monkey (Procolobus gordonorum) is among the most threatened primate species in Africa. Primarily arboreal and highly sensitive to hunting and habitat destruction, they provide a critical model to understanding whether anthropogenic disturbance impacts gut microbiota diversity. We sampled seven social groups inhabiting two forests (disturbed vs. undisturbed) in the Udzungwa Mountains of Tanzania. While Ruminococcaceae and Lachnospiraceae dominated in all individuals, reflecting their role in extracting energy from folivorous diets, analysis of genus composition showed a marked diversification across habitats, with gut microbiota α-diversity significantly higher in the undisturbed forest. Functional analysis suggests that such variation may be associated with food plant diversity in natural versus human-modified habitats, requiring metabolic pathways to digest xenobiotics. Thus, the effects of changes in gut microbiota should not be ignored to conserve endangered populations.