Curr. Issues Mol. Biol., Volume 24, Issue 1 (October 2017) – 7 articles

This review covers some recent advances in application of omics technologies to studying methylotrophs, with special reference to their activities in natural environments. Some of the developments highlighted in this review are the new outlook at the role of the XoxF-type, lanthanide-dependent methanol dehydrogenase in natural habitats, new mechanistic details of methane oxidation through the reverse methanogenesis pathway, propensity of 'aerobic' methanotrophs to thrive in hypoxic environments and potential connection of this process to denitrification, and a novel outlook at methane oxidation as a community function. Full article

Microbes constitute about a third of the Earth's biomass and are composed by an enormous genetic diversity. In a majority of environments the microbial communities play crucial roles for the ecosystem functioning, where a drastic biodiversity alteration or loss could lead to negative effects on the environment and sustainability. A central goal in microbiome studies is to elucidate the relation between microbial diversity to functions. A better understanding of the relation diversity-function would increase the ability to manipulate that diversity to improve plant and animal health and also setting conservation priorities. The recent advances in genomic methodologies in microbial ecology have provided means to assess highly complex communities in detail, making possible the link between diversity and the functions performed by the microbes. In this work we first explore some advances in bioinformatics tools to connect the microbial community biodiversity to their potential metabolism and after present some examples of how this information can be useful for a better understanding of the microbial role in the environment. Full article

Spatial and temporal variations related to hydric seasonality in abundance and diversity of denitrifier communities were examined in sediments taken from two sites differing in nitrate concentration along a stream in the Doñana National Park during a 3-year study. We found a positive relationship between the relative abundance of denitrifiers, determined as narG, napA, nirK, nirS and nosZ denitrification genes, and sediment nitrate content, with similar spatial and seasonal variations. However, we did not find association between denitrification activity and the community structure of denitrifiers. Because nosZ showed the strongest correlation with the content of nitrate in sediments, we used this gene as a molecular marker to construct eight genomic libraries. Analysis of these genomic libraries revealed that diversity of the nosZ-bearing communities was higher in the site with higher nitrate content. Regardless of nitrate concentration in the sediments, the Bradyrhizobiaceae and Rhodocyclaceae were the most abundant families. On the contrary, Rhizobiaceae was exclusively present in sediments with higher nitrate content. Results showed that differences in sediment nitrate concentration affect the composition and diversity of nosZ-bearing communities. Full article

A crucial step in the molecular detection of viruses in clinical specimens is the efficient extraction of viral nucleic acids. The total yield of viral nucleic acid from a clinical specimen is dependent on the specimen's volume, the initial virus concentration and the effectiveness provided by the extraction method. Recent next generation sequencing (NGS)-based diagnostic approaches (i.e. metagenomics) provide a molecular 'open view' into the sample, as they theoretically generate sequence reads of any nucleic acid present in a specimen in a statistically representative manner. However, since a higher virus-related read output promises better sensitivity in the subsequent bioinformatic analysis, the extraction method selected determines the reliability of diagnostic NGS. In this study nine commercially available kits for nucleic acid extraction were compared regarding the simultaneous isolation of DNA and RNA by real-time PCR, four of which were selected for subsequent comparison by NGS (QIAamp Viral RNA Mini Kit, QIAamp DNA Blood Mini Kit, QIAamp cador Pathogen Mini Kit and QIAamp MinElute Virus Spin Kit). The nucleic acid yields and the sequence read output were compared for four different model viruses–reovirus, orthomyxovirus, orthopoxvirus and paramyxovirus–each at defined but varying concentrations in the same sample. The total amount of nucleic acid was processed to sequence the RNA (as cDNA) and the DNA with quantification by Qubit and virus-specific quantitative real-time PCRs. NGS libraries were prepared for sequencing on the Illumina HiSeq 1500 system. Finally, the percentage of reads assignable to each virus was determined via mapping. Evaluation of different commercial nucleic acid extraction kits with four different viruses indicates little variation in the read numbers obtained for transcribed RNA or DNA by NGS. Since NGS is increasingly being used as a tool in diagnostics of infectious diseases, the individual steps of the complete process have to be validated carefully. Here we could show that for virus identification in liquid clinical specimens, any nucleic acid extraction kit that is performing well for PCR diagnostics can be used for NGS diagnostics as well and that the selection of the kit has only a minor impact on the yield of viral reads. Full article

Surveys of environmental microbial communities using a metagenomic approach produce vast volumes of multidimensional data regarding the phylogenetic and functional composition of the microbiota. Faced with such complex data, a metagenomic researcher needs to select the means for data analysis properly. Data visualization became an indispensable part of the exploratory data analysis and serves a key to the discoveries. While the molecular-genetic analysis of even a single bacterium presents multiple layers of data to be properly displayed and perceived, the studies of microbiota are significantly more challenging. Here we present a review of the state-of-the-art methods for the visualization of metagenomic data in a multilevel manner: from the methods applicable to an in-depth analysis of a single metagenome to the techniques appropriate for large-scale studies containing hundreds of environmental samples. Full article

Metagenomics, the application of high-throughput DNA sequencing for surveys of environmental samples, has revolutionized our view on the taxonomic and genetic composition of complex microbial communities. An enormous richness of microbiota keeps unfolding in the context of various fields ranging from biomedicine and food industry to geology. Primary analysis of metagenomic reads allows to infer semi-quantitative data describing the community structure. However, such compositional data possess statistical specific properties that are important to consider during preprocessing, hypothesis testing and interpreting the results of statistical tests. Failure to account for these specifics may lead to essentially wrong conclusions as a result of the survey. Here we present a researcher introduction to the field of metagenomics with the basic properties of microbial compositional data including statistical power and proposed distribution models, perform a review of the publicly available software tools developed specifically for such data and outline the recommendations for the application of the methods. Full article

Although from its origin metagenomics was concerned with composition of communities of microbial OTUs (Operational Taxonomic Units) living in a given habitat and their diversity and functional heterogeneity (concepts already well rooted in ecology), the new field was more 'environmentally' than 'ecologically' oriented. Probably by circumstantial reasons, metagenomics and ecology followed rather independent trajectories and conceptual and methodological gaps appeared. Recently, calls for the need of integrating the theoretical basis and methodologies coming from metagenomics (and other meta-omics) and ecology have been made. Here I will address some of the principles and methods of field ecology that, although useful in the context of environmental metagenomic studies, have been rather disregarded. In particular, I will emphasize the contribution of some well established concepts and methods of field ecology to a an appropriate field sampling and experimental design of environmental metagenomic studies. Full article