Background. India is home to 25% of all tuberculosis cases and the second highest number of multidrug resistant cases worldwide. However, little is known about the genetic diversity and resistance determinants of Indian Mycobacterium tuberculosis, particularly for the primary lineages found in India, lineages 1 and 3. Methods. We whole genome sequenced 223 randomly selected M. tuberculosis strains from 196 patients within the Tiruvallur and Madurai districts of Tamil Nadu in Southern India. Using comparative genomics, we examined genetic diversity, transmission patterns, and evolution of resistance. Results. Genomic analyses revealed (1) prevalence of strains from lineages 1 and 3, (2) recent transmission of strains among patients from the same treatment centers, (3) emergence of drug resistance within patients over time, (4) resistance gained in an order typical of strains from different lineages and geographies, (5) underperformance of known resistance-conferring mutations to explain phenotypic resistance in Indian strains relative to studies focused on other geographies, and (6) the possibility that resistance arose through mutations not previously implicated in resistance, or through infections with multiple strains that confound genotype-based prediction of resistance. Conclusions. In addition to substantially expanding the genomic perspectives of lineages 1 and 3, sequencing and analysis of M. tuberculosis whole genomes from Southern India highlight challenges of infection control and rapid diagnosis of resistant tuberculosis using current technologies. Further studies are needed to fully explore the complement of diversity and resistance determinants within endemic M. tuberculosis populations.@en

The haploid Saccharomyces cerevisiae strain CEN.PK113–7D is a popular model system for metabolic engineering and systems biology research. Current genome assemblies are based on short-read sequencing data scaffolded based on homology to strain S288C. However, these assemblies contain large sequence gaps, particularly in subtelomeric regions, and the assumption of perfect homology to S288C for scaffolding introduces bias. In this study, we obtained a near-complete genome assembly of CEN.PK113–7D using only Oxford Nanopore Technology's MinION sequencing platform. Fifteen of the 16 chromosomes, the mitochondrial genome and the 2-μm plasmid are assembled in single contigs and all but one chromosome starts or ends in a telomere repeat. This improved genome assembly contains 770 Kbp of added sequence containing 248 gene annotations in comparison to the previous assembly of CEN.PK113–7D. Many of these genes encode functions determining fitness in specific growth conditions and are therefore highly relevant for various industrial applications. Furthermore, we discovered a translocation between chromosomes III and VIII that caused misidentification of a MAL locus in the previous CEN.PK113–7D assembly. This study demonstrates the power of long-read sequencing by providing a high-quality reference assembly and annotation of CEN.PK113–7D and places a caveat on assumed genome stability of microorganisms.@en

Multidrug-resistant tuberculosis (MDR-TB), caused by drug-resistant strains of Mycobacterium tuberculosis, is an increasingly serious problem worldwide. Here we examined a data set of whole-genome sequences from 5,310 M. tuberculosis isolates from five continents. Despite the great diversity of these isolates with respect to geographical point of isolation, genetic background and drug resistance, the patterns for the emergence of drug resistance were conserved globally. We have identified harbinger mutations that often precede multidrug resistance. In particular, the katG mutation encoding p.Ser315Thr, which confers resistance to isoniazid, overwhelmingly arose before mutations that conferred rifampicin resistance across all of the lineages, geographical regions and time periods. Therefore, molecular diagnostics that include markers for rifampicin resistance alone will be insufficient to identify pre-MDR strains. Incorporating knowledge of polymorphisms that occur before the emergence of multidrug resistance, particularly katG p.Ser315Thr, into molecular diagnostics should enable targeted treatment of patients with pre-MDR-TB to prevent further development of MDR-TB.

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Many disciplines, from human genetics and oncology to plant breeding, microbiology and virology, commonly face the challenge of analyzing rapidly increasing numbers of genomes. In case of Homo sapiens, the number of sequenced genomes will in the next few years. Simply scaling up established bioinformatics pipelines will not be sufficient for leveraging the full potential of such rich genomic data sets. Instead, novel, qualitatively different computational
methods and paradigms are needed. We will witness the rapid extension of computational pan-genomics, a new sub-area of research in computational biology. In this article, we generalize existing definitions and understand a pangenome as any collection of genomic sequences to be analyzed jointly or to be used as a reference. We examine already available approaches to construct and use pan-genomes, discuss the potential benefits of future technologies and methodologies and review open challenges from the vantage point of the above-mentioned biological disciplines. As a prominent example for a computational paradigm shift, we particularly highlight the transition from the representation of reference genomes as strings to representations as graphs. We outline how this and other challenges from different application domains translate into common computational problems, point out relevant bioinformatics techniques and identify open problems in computer science. With this review, we aim to increase awareness that a joint approach to computational pangenomics can help address many of the problems currently faced in various domains.
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We report the whole-genome sequencing of two extensively drug-resistant tuberculosis strains belonging to the Euro-American S lineage. The RSA 114 strain showed single-nucleotide polymorphisms predicted to have drug efflux activity.@en

A more complete understanding of the genetic basis of drug resistance in Mycobacterium tuberculosis is critical for prompt diagnosis and optimal treatment, particularly for toxic second-line drugs such as D-cycloserine. Here we used the whole-genome sequences from 498 strains of M. tuberculosis to identify new resistance-conferring genotypes. By combining association and correlated evolution tests with strategies for amplifying signal from rare variants, we found that loss-of-function mutations in ald (Rv2780), encoding L-alanine dehydrogenase, were associated with unexplained drug resistance. Convergent evolution of this loss of function was observed exclusively among multidrug-resistant strains. Drug susceptibility testing established that ald loss of function conferred resistance to D-cycloserine, and susceptibility to the drug was partially restored by complementation of ald. Clinical strains with mutations in ald and alr exhibited increased resistance to D-cycloserine when cultured in vitro. Incorporation of D-cycloserine resistance in novel molecular diagnostics could allow for targeted use of this toxic drug among patients with susceptible infections.@en

Background
Mycobacterium africanum, made up of lineages 5 and 6 within the Mycobacterium tuberculosis complex (MTC), causes up to half of all tuberculosis cases in West Africa, but is rarely found outside of this region. The reasons for this geographical restriction remain unknown. Possible reasons include a geographically restricted animal reservoir, a unique preference for hosts of West African ethnicity, and an inability to compete with other lineages outside of West Africa. These latter two hypotheses could be caused by loss of fitness or altered interactions with the host immune system.
Methodology/Principal Findings
We sequenced 92 MTC clinical isolates from Mali, including two lineage 5 and 24 lineage 6 strains. Our genome sequencing assembly, alignment, phylogeny and average nucleotide identity analyses enabled us to identify features that typify lineages 5 and 6 and made clear that these lineages do not constitute a distinct species within the MTC. We found that in Mali, lineage 6 and lineage 4 strains have similar levels of diversity and evolve drug resistance through similar mechanisms. In the process, we identified a putative novel streptomycin resistance mutation. In addition, we found evidence of person-to-person transmission of lineage 6 isolates and showed that lineage 6 is not enriched for mutations in virulence-associated genes.
Conclusions
This is the largest collection of lineage 5 and 6 whole genome sequences to date, and our assembly and alignment data provide valuable insights into what distinguishes these lineages from other MTC lineages. Lineages 5 and 6 do not appear to be geographically restricted due to an inability to transmit between West African hosts or to an elevated number of mutations in virulence-associated genes. However, lineage-specific mutations, such as mutations in cell wall structure, secretion systems and cofactor biosynthesis, provide alternative mechanisms that may lead to host specificity.@en