Whole-Genome and Chromosome Evolution Associated with Host Adaptation and Speciation of the Wheat Pathogen Mycosphaerella graminicola

The fungus Mycosphaerella graminicola has been a pathogen of wheat since host domestication 10,000–12,000 years ago in the Fertile Crescent. The wheat-infecting lineage emerged from closely related Mycosphaerella pathogens infecting wild grasses. We use a comparative genomics approach to assess how the process of host specialization affected the genome structure of M. graminicola since divergence from the closest known progenitor species named M. graminicola S1. The genome of S1 was obtained by Illumina sequencing resulting in a 35 Mb draft genome sequence of 32X. Assembled contigs were aligned to the previously sequenced M. graminicola genome. The alignment covered >90% of the non-repetitive portion of the M. graminicola genome with an average divergence of 7%. The sequenced M. graminicola strain is known to harbor thirteen essential chromosomes plus eight dispensable chromosomes. We found evidence that structural rearrangements significantly affected the dispensable chromosomes while the essential chromosomes were syntenic. At the nucleotide level, the essential and dispensable chromosomes have evolved differently. The average synonymous substitution rate in dispensable chromosomes is considerably lower than in essential chromosomes, whereas the average non-synonymous substitution rate is three times higher. Differences in molecular evolution can be related to different transmission and recombination patterns, as well as to differences in effective population sizes of essential and dispensable chromosomes. In order to identify genes potentially involved in host specialization or speciation, we calculated ratios of synonymous and non-synonymous substitution rates in the >9,500 aligned protein coding genes. The genes are generally under strong purifying selection. We identified 43 candidate genes showing evidence of positive selection, one encoding a potential pathogen effector protein. We conclude that divergence of these pathogens was accompanied by structural rearrangements in the small dispensable chromosomes, while footprints of positive selection were present in only a small number of protein coding genes

Capturing the cloud of diversity reveals complexity and heterogeneity of MRSA carriage, infection and transmission.

Genome sequencing is revolutionizing clinical microbiology and our understanding of infectious diseases. Previous studies have largely relied on the sequencing of a single isolate from each individual. However, it is not clear what degree of bacterial diversity exists within, and is transmitted between individuals. Understanding this 'cloud of diversity' is key to accurate identification of transmission pathways. Here, we report the deep sequencing of methicillin-resistant Staphylococcus aureus among staff and animal patients involved in a transmission network at a veterinary hospital. We demonstrate considerable within-host diversity and that within-host diversity may rise and fall over time. Isolates from invasive disease contained multiple mutations in the same genes, including inactivation of a global regulator of virulence and changes in phage copy number. This study highlights the need for sequencing of multiple isolates from individuals to gain an accurate picture of transmission networks and to further understand the basis of pathogenesis.Thanks to Dr Alex O’Neill, University of Leeds and Dr Matthew Ellington, Public Health England for provision of RN4220 and RN4200mutS. We thank the core sequencing and informatics team at the Wellcome Trust Sanger Institute for sequencing of the isolates described in this study. This work was supported by a Medical Research Council Partnership grant (G1001787/1) held between the Department of Veterinary Medicine, University of Cambridge (M.A.H.), the School of Clinical Medicine, University of Cambridge (S.J.P.), the Moredun Research Institute, and the Wellcome Trust Sanger Institute (J.P. and S.J.P). S.J.P. receives support from the NIHR Cambridge Biomedical Research Centre. M.T.G.H., S.R.H. and J.P. were funded by Wellcome Trust grant no. 098051. G.G.R.M. was funded by an MRC studentship.This is the final version of the article. It first appeared from Nature Publishing Group via http://dx.doi.org/10.1038/ncomms756

GWAS for quantitative resistance phenotypes in Mycobacterium tuberculosis reveals resistance genes and regulatory regions

YesDrug resistance diagnostics that rely on the detection of resistance-related mutations could expedite patient care and TB eradication. We perform minimum inhibitory concentration testing for 12 anti-TB drugs together with Illumina whole-genome sequencing on 1452 clinical Mycobacterium tuberculosis (MTB) isolates. We evaluate genome-wide associations between mutations in MTB genes or non-coding regions and resistance, followed by validation in an independent data set of 792 patient isolates. We confirm associations at 13 non-canonical loci, with two involving non-coding regions. Promoter mutations are measured to have smaller average effects on resistance than gene body mutations. We estimate the heritability of the resistance phenotype to 11 anti-TB drugs and identify a lower than expected contribution from known resistance genes. This study highlights the complexity of the genomic mechanisms associated with the MTB resistance phenotype, including the relatively large number of potentially causal loci, and emphasizes the contribution of the non-coding portion of the genome.Biomedical research grant from the American Lung Association (PI MF, RG-270912-N), a K01 award from the BD2K initiative (PI MF, ES026835), and an NIAID U19 CETR grant (P.I. M.M., AI109755), the Belgian Science Policy (Belspo) (L.R., C.J.M.)

Molecular adaptation during a rapid adaptive radiation.

"Explosive" adaptive radiations on islands remain one of the most puzzling evolutionary phenomena and the evolutionary genetic processes behind such radiations remain unclear. Rapid morphological and ecological evolution during island radiations suggests that many genes may be under fairly strong selection, although this remains untested. Here, we report that during a rapid recent diversification in the Hawaiian endemic plant genus Schiedea (Caryophyllaceae), 5 in 36 studied genes evolved under positive selection. Positively selected genes are involved in defence mechanisms, photosynthesis, and reproduction. Comparison with eight mainland plant groups demonstrates both the relaxation of purifying selection and more widespread positive selection in Hawaiian Schiedea. This provides compelling evidence that adaptive evolution of protein-coding genes may play a significant role during island adaptive radiations

Prevalence of Staphylococcus aureus protein A (spa) mutants in the community and hospitals in Oxfordshire.

BACKGROUND: Staphylococcal protein A (spa) is an important virulence factor which enables Staphylococcus aureus to evade host immune responses. Genotypes known as "spa-types", based on highly variable Xr region sequences of the spa-gene, are frequently used to classify strains. A weakness of current spa-typing primers is that rearrangements in the IgG-binding region of the gene cause 1-2% of strains to be designated as "non-typeable". RESULTS: We developed an improved primer which enabled sequencing of all strains, containing any type of genetic rearrangement, in a large study among community carriers and hospital inpatients in Oxfordshire, UK (6110 isolates). We identified eight novel spa-gene variants, plus one previously described. Three of these rearrangements would be designated "non-typeable" using current spa-typing methods; they occurred in 1.8% (72/3905) asymptomatically carried and 0.6% (14/2205) inpatient S. aureus strains. Some individuals were simultaneously colonized by both formerly non-typeable and typeable strains; previously such patients would have been identified as carrying only currently typeable strains, underestimating mixed carriage prevalence and diversity. Formerly non-typeable strains were found in more spa-types associated with multilocus sequence type ST398 (35%), common among livestock, compared to other groups with any non-typeable strains (1-4%), suggesting particular spa-types may have been under-represented in previous human studies. CONCLUSIONS: This improved method allows us to spa-type previously non-typeable strains with rearrangements in the spa-gene and to resolve cases of mixed colonization with deletions in one or more strains, thus accounting for hidden diversity of S. aureus in both community and hospital environments

Línea proletaria : plataforma de expresión de Núcleos Obreros Comunistas

VILADOT-CEDO

Mycobacterial DNA extraction for whole-genome sequencing from early positive liquid (MGIT) cultures

We developed a low-cost and reliable method of DNA extraction from as little as 1 ml of early positive mycobacterial growth indicator tube (MGIT) cultures that is suitable for whole-genome sequencing to identify mycobacterial species and predict antibiotic resistance in clinical samples. The DNA extraction method is based on ethanol precipitation supplemented by pretreatment steps with a MolYsis kit or saline wash for the removal of human DNA and a final DNA cleanup step with solid-phase reversible immobilization beads. The protocol yielded ≥0.2 ng/μl of DNA for 90% (MolYsis kit) and 83% (saline wash) of positive MGIT cultures. A total of 144 (94%) of the 154 samples sequenced on the MiSeq platform (Illumina) achieved the target of 1 million reads, with 90% coverage achieved. The DNA extraction protocol, therefore, will facilitate fast and accurate identification of mycobacterial species and resistance using a range of bioinformatics tools