Analysing Microbial Community Composition through Amplicon Sequencing: From Sampling to Hypothesis Testing

Microbial ecology as a scientific field is fundamentally driven by technological advance. The past decade's revolution in DNA sequencing cost and throughput has made it possible for most research groups to map microbial community composition in environments of interest. However, the computational and statistical methodology required to analyse this kind of data is often not part of the biologist training. In this review, we give a historical perspective on the use of sequencing data in microbial ecology and restate the current need for this method; but also highlight the major caveats with standard practices for handling these data, from sample collection and library preparation to statistical analysis. Further, we outline the main new analytical tools that have been developed in the past few years to bypass these caveats, as well as highlight the major requirements of common statistical practices and the extent to which they are applicable to microbial data. Besides delving into the meaning of select alpha- and beta-diversity measures, we give special consideration to techniques for finding the main drivers of community dissimilarity and for interaction network construction. While every project design has specific needs, this review should serve as a starting point for considering what options are available

Elevation and plant species identity jointly shape a diverse arbuscular mycorrhizal fungal community in the High Arctic

Knowledge about the distribution and local diversity patterns of arbuscular mycorrhizal (AM) fungi are limited for extreme environments such as the Arctic, where most studies have focused on spore morphology or root colonization. We here studied the joint effects of plant species identity and elevation on AM fungal distribution and diversity. We sampled roots of 19 plant species in 18 locations in Northeast Greenland, using next generation sequencing to identify AM fungi. We studied the joint effect of plant species, elevation and selected abiotic conditions on AM fungal presence, richness and composition. We identified 29 AM fungal virtual taxa (VT), of which six represent putatively new VT. Arbuscular mycorrhizal fungal presence increased with elevation, and as vegetation cover and the active soil layer decreased. Arbuscular mycorrhizal fungal composition was shaped jointly by elevation and plant species identity. We demonstrate that the Arctic harbours a relatively species-rich and nonrandomly distributed diversity of AM fungi. Given the high diversity and general lack of knowledge exposed herein, we encourage further research into the diversity, drivers and functional role of AM fungi in the Arctic. Such insight is urgently needed for an area with some of the globally highest rates of climate change.Peer reviewe

Observações sobre a mudança tecnológica no setor de máquinas ferramentas do Brasil

Incluye BibliografíaResume la evolucion historica del sector de maquinas herramientas del Brasil en base a cuatro estudios de casos de firmas nacionales. Analiza algunos aspectos del problema tecnologico del sector y presenta cada estudio de caso separadamente

Elevation and plant species identity jointly shape a diverse arbuscular mycorrhizal fungal community in the High Arctic

Knowledge about the distribution and local diversity patterns of arbuscular mycorrhizal (AM) fungi are limited for extreme environments such as the Arctic, where most studies have focused on spore morphology or root colonization. We here studied the joint effects of plant species identity and elevation on AM fungal distribution and diversity. We sampled roots of 19 plant species in 18 locations in Northeast Greenland, using next generation sequencing to identify AM fungi. We studied the joint effect of plant species, elevation and selected abiotic conditions on AM fungal presence, richness and composition. We identified 29 AM fungal virtual taxa (VT), of which six represent putatively new VT. Arbuscular mycorrhizal fungal presence increased with elevation, and as vegetation cover and the active soil layer decreased. Arbuscular mycorrhizal fungal composition was shaped jointly by elevation and plant species identity. We demonstrate that the Arctic harbours a relatively species-rich and nonrandomly distributed diversity of AM fungi. Given the high diversity and general lack of knowledge exposed herein, we encourage further research into the diversity, drivers and functional role of AM fungi in the Arctic. Such insight is urgently needed for an area with some of the globally highest rates of climate change

BARM and BalticMicrobeDB, a reference metagenome and interface to meta-omic data for the Baltic Sea

The Baltic Sea is one of the world’s largest brackish water bodies and is characterised by pronounced physicochemical gradients where microbes are the main biogeochemical catalysts. Meta-omic methods provide rich information on the composition of, and activities within, microbial ecosystems, but are computationally heavy to perform. We here present the Baltic Sea Reference Metagenome (BARM), complete with annotated genes to facilitate further studies with much less computational effort. The assembly is constructed using 2.6 billion metagenomic reads from 81 water samples, spanning both spatial and temporal dimensions, and contains 6.8 million genes that have been annotated for function and taxonomy. The assembly is useful as a reference, facilitating taxonomic and functional annotation of additional samples by simply mapping their reads against the assembly. This capability is demonstrated by the successful mapping and annotation of 24 external samples. In addition, we present a public web interface, BalticMicrobeDB, for interactive exploratory analysis of the dataset

Metapopulation theory identifies biogeographical patterns among core and satellite marine bacteria scaling from tens to thousands of kilometers:Applied metapopulation theory for marine microbes

Metapopulation theory developed in terrestrial ecology provides applicable frameworks for interpreting the role of local and regional processes in shaping species distribution patterns. Yet, empirical testing of metapopulation models on microbial communities is essentially lacking. We determined regional bacterioplankton dynamics from monthly transect sampling in the Baltic Sea Proper using 16S rRNA gene sequencing. A strong positive trend was found between local relative abundance and occupancy of populations. Notably, the occupancy-frequency distributions were significantly bimodal with a satellite mode of rare endemic populations and a core mode of abundant cosmopolitan populations (e.g. Synechococcus, SAR11 and SAR86 clade members). Temporal changes in population distributions supported several theoretical frameworks. Still, bimodality was found among bacterioplankton communities across the entire Baltic Sea, and was also frequent in globally distributed datasets. Datasets spanning waters with widely different physicochemical characteristics or environmental gradients typically lacked significant bimodal patterns. When such datasets were divided into subsets with coherent environmental conditions, bimodal patterns emerged, highlighting the importance of positive feedbacks between local abundance and occupancy within specific biomes. Thus, metapopulation theory applied to microbial biogeography can provide novel insights into the mechanisms governing shifts in biodiversity resulting from natural or anthropogenically induced changes in the environment

Multiple identification of most important waterborne protozoa in surface water used for irrigation purposes by 18S rRNA amplicon-based metagenomics

[EN] Understanding waterborne protozoan parasites (WPPs) diversity has important implications in public health. In this study, we evaluated a NGS-based method as a detection approach to identify simultaneously most important WPPs using 18S rRNA high-throughput sequencing. A set of primers to target the V4 18S rRNA region of WPPs such as Cryptosporidium spp., Giardia sp., Blastocystis sp., Entamoeba spp, Toxoplasma sp. and free-living amoebae (FLA) was designed. In order to optimize PCR conditions before sequencing, both a mock community with a defined composition of representative WPPs and a real water sample inoculated with specific WPPs DNA were prepared. Using the method proposed in this study, we have detected the presence of Giardia intestinalis, Acanthamoeba castellanii, Toxoplasma gondii, Entamoeba histolytica and Blastocystis sp. at species level in real irrigation water samples. Our results showed that untreated surface irrigation water in open fields can provide an important source of WPPs. Therefore, the methodology proposed in this study can establish a basis for an accurate and effective diagnostic of WPPs to provide a better understanding of the risk associated to irrigation water.This work was supported through the project funded by the Spanish Ministry of Economy and Competitiveness (MINECO) in the frame of the collaborative international consortium JPIW2013-095-C03-02 of the Water Challenges for a Changing World Joint Programming Initiative (Water JPI) Pilot Call. R. Perez acknowledges support from MINECO program "Promocion de Empleo Joven e Implantacion de la Garantia Juvenil en I + D + i".Moreno Trigos, MY.; Moreno-Mesonero, L.; Amoros, I.; Pérez-Santonja, R.; Morillo, J.; Alonso Molina, JL. (2018). Multiple identification of most important waterborne protozoa in surface water used for irrigation purposes by 18S rRNA amplicon-based metagenomics. International Journal of Hygiene and Environmental Health. 221(1):102-111. https://doi.org/10.1016/j.ijheh.2017.10.008S102111221

The nasal and gut microbiome in Parkinson's disease and idiopathic rapid eye movement sleep behavior disorder.

BACKGROUND: Increasing evidence connects the gut microbiota and the onset and/or phenotype of Parkinson's disease (PD). Differences in the abundances of specific bacterial taxa have been reported in PD patients. It is, however, unknown whether these differences can be observed in individuals at high risk, for example, with idiopathic rapid eye movement sleep behavior disorder, a prodromal condition of alpha-synuclein aggregation disorders including PD. OBJECTIVES: To compare microbiota in carefully preserved nasal wash and stool samples of subjects with idiopathic rapid eye movement sleep behavior disorder, manifest PD, and healthy individuals. METHODS: Microbiota of flash-frozen stool and nasal wash samples from 76 PD patients, 21 idiopathic rapid eye movement sleep behavior disorder patients, and 78 healthy controls were assessed by 16S and 18S ribosomal RNA amplicon sequencing. Seventy variables, related to demographics, clinical parameters including nonmotor symptoms, and sample processing, were analyzed in relation to microbiome variability and controlled differential analyses were performed. RESULTS: Differentially abundant gut microbes, such as Akkermansia, were observed in PD, but no strong differences in nasal microbiota. Eighty percent of the differential gut microbes in PD versus healthy controls showed similar trends in idiopathic rapid eye movement sleep behavior disorder, for example, Anaerotruncus and several Bacteroides spp., and correlated with nonmotor symptoms. Metagenomic sequencing of select samples enabled the reconstruction of genomes of so far uncharacterized differentially abundant organisms. CONCLUSION: Our study reveals differential abundances of gut microbial taxa in PD and its prodrome idiopathic rapid eye movement sleep behavior disorder in comparison to the healthy controls, and highlights the potential of metagenomics to identify and characterize microbial taxa, which are enriched or depleted in PD and/or idiopathic rapid eye movement sleep behavior disorder. (c) 2017 International Parkinson and Movement Disorder Society