The distribution of fitness effects among synonymous mutations in a gene under selection, bioRxiv, 2019-02-19

AbstractThe fitness effects of synonymous mutations, nucleotide changes that do not alter the encoded amino acid, have often been assumed to be neutral, but a growing body of evidence suggests otherwise. We used site-directed mutagenesis coupled with direct measures of competitive fitness to estimate the distribution of fitness effects among synonymous mutations for a gene under selection. Synonymous mutations had highly variable fitness effects, both deleterious and beneficial, resembling those of nonsynonymous mutations in the same gene. This variation in fitness was underlain by changes in transcription linked to the creation of internal promoter sites. A positive correlation between fitness and the presence of synonymous substitutions across a phylogeny of related Pseudomonads suggests these mutations may be common in nature. Taken together, our results provide the most compelling evidence to date that synonymous mutations with non-neutral fitness effects may in fact be commonplace.

biorxiv evolutionary-biology 0-100-users 2019

Evolutionary dynamics of phage resistance in bacterial biofilms, bioRxiv, 2019-02-17

Interactions among bacteria and their viral predators, the bacteriophages, are likely among the most common ecological phenomena on Earth. The constant threat of phage infection to bacterial hosts, and the imperative of achieving infection on the part of phages, drives an evolutionary contest in which phage-resistant bacteria emerge, often followed by phages with new routes of infection. This process has received abundant theoretical and experimental attention for decades and forms an important basis for molecular genetics and theoretical ecology and evolution. However, at present, we know very little about the nature of phage-bacteria interaction -- and the evolution of phage resistance -- inside the surface-bound communities that microbes usually occupy in natural environments. These communities, termed biofilms, are encased in a matrix of secreted polymers produced by their microbial residents. Biofilms are spatially constrained such that interactions become limited to neighbors or near-neighbors; diffusion of solutes and particulates is reduced; and there is pronounced heterogeneity in nutrient access and therefore physiological state. These factors can dramatically impact the way phage infections proceed even in simple, single-strain biofilms, but we still know little of their effect on phage resistance evolutionary dynamics. Here we explore this problem using a computational simulation framework customized for implementing phage infection inside multi-strain biofilms. Our simulations predict that it is far easier for phage-susceptible and phage- resistant bacteria to coexist inside biofilms relative to planktonic culture, where phages and hosts are well-mixed. We characterize the negative frequency dependent selection that underlies this coexistence, and we then test and confirm this prediction using an experimental model of biofilm growth measured with confocal microscopy at single-cell and single-phage resolution.

biorxiv microbiology 0-100-users 2019

Large, three-generation CEPH families reveal post-zygotic mosaicism and variability in germline mutation accumulation, bioRxiv, 2019-02-17

AbstractThe number of de novo mutations (DNMs) found in an offspring’s genome increases with both paternal and maternal age. But does the rate of mutation accumulation in human gametes differ across families? Using sequencing data from 33 large, three-generation CEPH families, we observed significant variability in parental age effects on DNM counts across families, with estimates ranging from 0.19 to 3.24 DNMs per year. Additionally, we found that approximately 3% of DNMs originated following primordial germ cell specification (PGCS) in a parent, and differed from non-mosaic germline DNMs in their mutational spectra. We also discovered that nearly 10% of candidate DNMs in the second generation were post-zygotic, and present in both somatic and germ cells; these gonosomal mutations occurred at equivalent frequencies on both parental haplotypes. Our results demonstrate that the rate of germline mutation accumulation varies among families with similar ancestry, and confirm that post-zygotic mosaicism is a substantial source of de novo mutations in humans.Data and code availability. Code used for statistical analysis and figure generation has been deposited on GitHub as a collection of annotated Jupyter Notebooks <jatsext-link xmlnsxlink=httpwww.w3.org1999xlink ext-link-type=uri xlinkhref=httpsgithub.comquinlan-labceph-dnm-manuscript>httpsgithub.comquinlan-labceph-dnm-manuscript<jatsext-link>. Data files containing high-confidence de novo mutations, as well as the gonosomal and post-primordial germ cell specification (PGCS) mosaic mutations, are included with these Notebooks. To mitigate compatibility issues, we have also made all notebooks available in a Binder environment, accessible at the above GitHub repository.

biorxiv genetics 0-100-users 2019

Enterococcus faecium genome dynamics during long-term asymptomatic patient gut colonization, bioRxiv, 2019-02-16

Background E. faecium is a gut commensal of humans and animals. In addition, it has recently emerged as an important nosocomial pathogen through the acquisition of genetic elements that confer resistance to antibiotics and virulence. We performed a whole-genome sequencing based study on 96 multidrug-resistant E. faecium strains that asymptomatically colonized five patients with the aim to describe the genome dynamics of this species. Results The patients were hospitalized on multiple occasions and isolates were collected over periods ranging from 15 months to 6.5 years. Ninety-five of the sequenced isolates belonged to E. faecium clade A1, which was previously determined to be responsible for the vast majority of clinical infections. The clade A1 strains clustered into six clonal groups of highly similar isolates, three of which entirely consisted of isolates from a single patient. We also found evidence of concurrent colonization of patients by multiple distinct lineages and transfer of strains between patients during hospitalisation. We estimated the evolutionary rate of two clonal groups that colonized a single patient at 12.6 and 25.2 single nucleotide polymorphisms (SNPs)genomeyear. A detailed analysis of the accessory genome of one of the clonal groups revealed considerable variation due to gene gain and loss events, including the chromosomal acquisition of a 37 kbp prophage and the loss of an element containing carbohydrate metabolism-related genes. We determined the presence and location of twelve different Insertion Sequence (IS) elements, with ISEfa5 showing a unique pattern of location in 24 of the 25 isolates, suggesting widespread ISEfa5 excision and insertion into the genome during gut colonization. Conclusions Our findings show that the E. faecium genome is highly dynamic during asymptomatic colonization of the patient gut. We observe considerable genomic flexibility due to frequent horizontal gene transfer and recombination, which can contribute to the generation of genetic diversity within the species and, ultimately, can contribute to its success as a nosocomial pathogen.

biorxiv microbiology 0-100-users 2019

Semi-quantitative characterisation of mixed pollen samples using MinION sequencing and Reverse Metagenomics (RevMet), bioRxiv, 2019-02-16

The ability to identify and quantify the constituent plant species that make up a mixed-species sample of pollen has important applications in ecology, conservation, and agriculture. Recently, metabarcoding protocols have been developed for pollen that can identify constituent plant species, but there are strong reasons to doubt that metabarcoding can accurately quantify their relative abundances. A PCR-free, shotgun metagenomics approach has greater potential for accurately quantifying species relative abundances, but applying metagenomics to eukaryotes is challenging due to low numbers of reference genomes. We have developed a pipeline, RevMet (Reverse Metagenomics), that allows reliable and semi-quantitative characterization of the species composition of mixed-species eukaryote samples, such as bee-collected pollen, without requiring reference genomes. Instead, reference species are represented only by 'genome skims' low-cost, low-coverage, shortread sequence datasets. The skims are mapped to individual long reads sequenced from mixed-species samples using the MinION, a portable nanopore sequencing device, and each long read is uniquely assigned to a plant species. We genome-skimmed 49 wild UK plant species, validated our pipeline with mock DNA mixtures of known composition, and then applied RevMet to pollen loads collected from wild bees. We demonstrate that RevMet can identify plant species present in mixed-species samples at proportions of DNA &gt;1%, with few false positives and false negatives, and reliably differentiate species represented by high versus low amounts of DNA in a sample. The RevMet pipeline could readily be adapted to generate semi-quantitative datasets for a wide range of mixed eukaryote samples, which could include characterising diets, quantifying allergenic pollen from air samples, quantifying soil fauna, and identifying the compositions of algal and diatom communities. Our per-sample costs were GBP 90 per genome skim and GBP 60 per pollen sample, and new versions of sequencers available now will further reduce these costs.

biorxiv ecology 0-100-users 2019

Cannabis use, depression and self-harm phenotypic and genetic relationships, bioRxiv, 2019-02-15

Background and Aims The use of cannabis has previously been linked to both depression and self-harm, however the role of genetics in this relationship are unclear. We aimed to examine the phenotypic and genetic relationships between these traits.Design Genetic and cross-sectional phenotypic data collected through UK Biobank, together with consortia genome-wide association study summary statistics. These data were used to assess the phenotypic and genetic relationship between cannabis use, depression and self harm.Setting UK, with additional international consortia dataParticipants N=126,291 British adults aged between 40 and 70 years, recruited into UK Biobank.Measurements Genome-wide genetic data, phenotypic data on lifetime history of cannabis use, depression and self-harm.Findings In UK Biobank, cannabis use is associated with increased likelihood of depression (OR=1.64, 95% CI=1.59-1.70, p=1.19x10-213) and self-harm (OR=2.85, 95% CI=2.69-3.01, p=3.46x10-304). The strength of this phenotypic association is stronger when more severe trait definitions of cannabis use and depression are considered. Additionally, significant genetic correlations are seen between cannabis use and depression using consortia summary statistics (rg=0.289, SE=0.036, p=1.45x10-15). Polygenic risk scores for cannabis use and depression both explain a small but significant proportion of variance in cannabis use, depression and self harm within a UK Biobank target sample. However, two-sample Mendelian randomisation analyses were not significant.Conclusions Cannabis use is both phenotypically and genetically associated with depression and self harm. Future work dissecting the causal mechanism linking these traits may have implications for cannabis users.

biorxiv genetics 0-100-users 2019