Human genomic data have different statistical properties than the data of randomised controlled trials.

Madole & Harden argue that the Mendelian reshuffling of genes and genomes is analogous to randomised controlled trials. We are not convinced by their arguments. First, their recipe for meeting the demands on randomised experiments is inherently inconsistent. Second, disequilibrium across chromosomes conflicts with their assumption of statistical independence. Third, the genome-wide association study (GWAS) method has many pitfalls, including low repeatability.

BioMiCo: a supervised Bayesian model for inference of microbial community structure.

BACKGROUND: Microbiome samples often represent mixtures of communities, where each community is composed of overlapping assemblages of species. Such mixtures are complex, the number of species is huge and abundance information for many species is often sparse. Classical methods have a limited value for identifying complex features within such data. RESULTS: Here, we describe a novel hierarchical model for Bayesian inference of microbial communities (BioMiCo). The model takes abundance data derived from environmental DNA, and models the composition of each sample by a two-level hierarchy of mixture distributions constrained by Dirichlet priors. BioMiCo is supervised, using known features for samples and appropriate prior constraints to overcome the challenges posed by many variables, sparse data, and large numbers of rare species. The model is trained on a portion of the data, where it learns how assemblages of species are mixed to form communities and how assemblages are related to the known features of each sample. Training yields a model that can predict the features of new samples. We used BioMiCo to build models for three serially sampled datasets and tested their predictive accuracy across different time points. The first model was trained to predict both body site (hand, mouth, and gut) and individual human host. It was able to reliably distinguish these features across different time points. The second was trained on vaginal microbiomes to predict both the Nugent score and individual human host. We found that women having normal and elevated Nugent scores had distinct microbiome structures that persisted over time, with additional structure within women having elevated scores. The third was trained for the purpose of assessing seasonal transitions in a coastal bacterial community. Application of this model to a high-resolution time series permitted us to track the rate and time of community succession and accurately predict known ecosystem-level events. CONCLUSION: BioMiCo provides a framework for learning the structure of microbial communities and for making predictions based on microbial assemblages. By training on carefully chosen features (abiotic or biotic), BioMiCo can be used to understand and predict transitions between complex communities composed of hundreds of microbial species.

Studying genome heterogeneity within the arbuscular mycorrhizal fungal cytoplasm.

Although heterokaryons have been reported in nature, multicellular organisms are generally assumed genetically homogeneous. Here, we investigate the case of arbuscular mycorrhizal fungi (AMF) that form symbiosis with plant roots. The growth advantages they confer to their hosts are of great potential benefit to sustainable agricultural practices. However, measuring genetic diversity for these coenocytes is a major challenge: Within the same cytoplasm, AMF contain thousands of nuclei and show extremely high levels of genetic variation for some loci. The extent and physical location of polymorphism within and between AMF genomes is unclear. We used two complementary strategies to estimate genetic diversity in AMF, investigating polymorphism both on a genome scale and in putative single copy loci. First, we used data from whole-genome pyrosequencing of four AMF isolates to describe genetic diversity, based on a conservative network-based clustering approach. AMF isolates showed marked differences in genome-wide diversity patterns in comparison to a panel of control fungal genomes. This clustering approach further allowed us to provide conservative estimates of Rhizophagus spp. genomes sizes. Second, we designed new putative single copy genomic markers, which we investigated by massive parallel amplicon sequencing for two Rhizophagus irregularis and one Rhizophagus sp. isolates. Most loci showed high polymorphism, with up to 103 alleles per marker. This polymorphism could be distributed within or between nuclei. However, we argue that the Rhizophagus isolates under study might be heterokaryotic, at least for the putative single copy markers we studied. Considering that genetic information is the main resource for identification of AMF, we suggest that special attention is warranted for the study of these ecologically important organisms.

Interactions in the microbiome: communities of organisms and communities of genes.

A central challenge in microbial community ecology is the delineation of appropriate units of biodiversity, which can be taxonomic, phylogenetic, or functional in nature. The term 'community' is applied ambiguously; in some cases, the term refers simply to a set of observed entities, while in other cases, it requires that these entities interact with one another. Microorganisms can rapidly gain and lose genes, potentially decoupling community roles from taxonomic and phylogenetic groupings. Trait-based approaches offer a useful alternative, but many traits can be defined based on gene functions, metabolic modules, and genomic properties, and the optimal set of traits to choose is often not obvious. An analysis that considers taxon assignment and traits in concert may be ideal, with the strengths of each approach offsetting the weaknesses of the other. Individual genes also merit consideration as entities in an ecological analysis, with characteristics such as diversity, turnover, and interactions modeled using genes rather than organisms as entities. We identify some promising avenues of research that are likely to yield a deeper understanding of microbial communities that shift from observation-based questions of 'Who is there?' and 'What are they doing?' to the mechanistically driven question of 'How will they respond?'

Allelic differences within and among sister spores of the arbuscular mycorrhizal fungus Glomus etunicatum suggest segregation at sporulation.

Arbuscular mycorrhizal fungi (AMF) are root-inhabiting fungi that form mutualistic symbioses with their host plants. AMF are made up of coenocytic networks of hyphae through which nuclei and organelles can freely migrate. In this study, we investigated the possibility of a genetic bottleneck and segregation of allelic variation at sporulation for a low-copy Polymerase1-like gene, PLS. Specifically, our objectives were (1) to estimate what allelic diversity is passed on to a single spore (2) to determine whether this diversity is less than the total amount of variation found in all spores (3) to investigate whether there is any differential segregation of allelic variation. We inoculated three tomato plants with a single spore of Glomus etunicatum each and after six months sampled between two and three daughter spores per tomato plant. Pyrosequencing PLS amplicons in eight spores revealed high levels of allelic diversity; between 43 and 152 alleles per spore. We corroborated the spore pyrosequencing results with Sanger- and pyrosequenced allele distributions from the original parent isolate. Both sequencing methods retrieved the most abundant alleles from the offspring spore allele distributions. Our results indicate that individual spores contain only a subset of the total allelic variation from the pooled spores and parent isolate. Patterns of allele diversity between spores suggest the possibility for segregation of PLS alleles among spores. We conclude that a genetic bottleneck could potentially occur during sporulation in AMF, with resulting differences in genetic variation among sister spores. We suggest that the effects of this bottleneck may be countered by anastomosis (hyphal fusion) between related hyphae.

Molecular biodiversity of arbuscular mycorrhizal fungi in trace metal-polluted soils.

We assessed the indigenous arbuscular mycorrhizal fungi (AMF) community structure from the roots and associated soil of Plantago major (plantain) plants growing on sites polluted with trace metals (TM) and on unpolluted sites. Uncontaminated and TM-contaminated sites containing As, Cd, Cu, Pb, Sn and Zn were selected based on a survey of the TM concentration in soils of community gardens in the City of Montréal. Total genomic DNA was extracted directly from these samples. PCR followed by denaturing gradient gel electrophoresis (PCR-DGGE), augmented by cloning and sequencing, as well as direct sequencing techniques, was all used to investigate AMF community structure. We found a decreased diversity of native AMF (assessed by the number of AMF ribotypes) in soils and plant roots harvested from TM-polluted soils compared with unpolluted soils. We also found that community structure was modified by TM contamination. Various species of Glomus, Scutellospora aurigloba and S. calospora were the most abundant ribotypes detected in unpolluted soil; ribotypes of G. etunicatum, G. irregulare/G. intraradices and G. viscosum were found in both polluted and unpolluted soils, while ribotypes of G. mosseae and Glomus spp. (B9 and B13) were dominant in TM-polluted soils. The predominance of G. mosseae in metal-polluted sites suggests the tolerance of this species to TM stress, as well as its potential use for phytoremediation. These data are relevant for our understanding of how AMF microbial communities respond to natural environments that contain a broad variety of toxic inorganic compounds and will substantially expand our knowledge of AMF ecology and biodiversity.

The flow of antimicrobial peptide genes through a genetic barrier between Mytilus edulis and M. galloprovincialis.

We studied the population genetics of two antimicrobial peptide (AMP) loci, called Mytilin B and Mytilus galloprovincialis defensin 2 (MGD2), in the secondary contact mosaic hybrid zone between Mytilus edulis and M. galloprovincialis. The isolation period between the two species was estimated to be approximately 1 million years (range, 0.5 million to 2 million years) long. During this period, coevolution between microbes and the immune system has likely occurred. The secondary contact, which would date back to approximately 25,000 (0-200,000) years, recently allowed these coadaptations to be rearranged through hybridization. Distinctive polymorphisms were uncovered in coding sequences of the two AMP loci such as insertion/deletion of codons or bisubstituted codons. Very low levels of differentiation were observed between populations of the two species at both loci, while other nuclear loci often showed marked structure among the same samples. The absence of population differentiation proved to be the consequence of secondary introgression of highly divergent alleles. While only a few recombinants were observed at the Mytilin B locus, the MGD2 locus showed a high intragenic recombination rate, which increased in the exon coding for the mature peptide. In addition, standard neutrality tests revealed significant deviations from the mutation-drift equilibrium at both loci. These results suggest that either balancing or directional selection is likely to play a role in the evolution of the two AMPs and introgression would be adaptive. However, evidence accumulated at the Mytilin B locus allows neither for identification of the direction of selection nor for any conclusions on whether selection acted directly on the antimicrobial peptide itself. At the MGD2 locus, a spatial variation of polymorphism patterns along the sequence suggests that selection was direct, although the precise nature of the selection (directional vs. balancing) remains unclear. This study concurs with previous reports of an effect of slight selection on AMP genes evolution in other invertebrates, although selection does not necessarily act on the mature peptides.

Low effective population size and evidence for inbreeding in an overexploited flatfish, plaice (Pleuronectes platessa L.).

Overexploitation and subsequent collapses of major worldwide fisheries has made it clear that marine stocks are no inexhaustible. Unfortunately, the perception remains that marine fished are resilient to large population reductions, as even a commercially 'collapsed' stock will still consist of millions of individuals. Coupled with this notion is the idea that fisheries can, therefore, have little effect on the genetic diversity of stocks. We used DNA from archived otoliths collected between 1924 and 1972 together with 2002 juvenile;s tissue to estimate effective population size (Ne) in plaice (Pleuronrctes platessa). Ne was estimated at 20,000 in the North Sea and 2000 in Iceland. These values are five orders of magnitude smaller than the estimated census size foe the two locations. Populations examined between 1924 and 1960 were in Hardy-Weinberg equilibrium, whereas populations examined after 1970 were not. Extensive testing was performed to rule out genotyping artefacts and Wahlund effects. The significant heterozygote deficiencies found from 1970 onward were attributed to inbreeding. The emergence of inbreeding between 1905 and 19070 coincides with the increase in fishing mortality after World War II. Although the biological mechanisms remain speculative, our demonstration of inbreeding signals the need for understanding the social and mating behaviour in commercially important fishes.