Publications by authors named "Sean Sykes"

21 Publications

  • Page 1 of 1

Tracing Genetic Exchange and Biogeography of var. at the Global Population Level.

Genetics 2017 09 5;207(1):327-346. Epub 2017 Jul 5.

Infectious Disease and Microbiome Program, Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, Massachusetts 02142

var. is the causative agent of cryptococcal meningitis, a significant source of mortality in immunocompromised individuals, typically human immunodeficiency virus/AIDS patients from developing countries. Despite the worldwide emergence of this ubiquitous infection, little is known about the global molecular epidemiology of this fungal pathogen. Here we sequence the genomes of 188 diverse isolates and characterize the major subdivisions, their relative diversity, and the level of genetic exchange between them. While most isolates of var. belong to one of three major lineages (VNI, VNII, and VNB), some haploid isolates show hybrid ancestry including some that appear to have recently interbred, based on the detection of large blocks of each ancestry across each chromosome. Many isolates display evidence of aneuploidy, which was detected for all chromosomes. In diploid isolates of var. serotype AA) and of hybrids with var. (serotype AD) such aneuploidies have resulted in loss of heterozygosity, where a chromosomal region is represented by the genotype of only one parental isolate. Phylogenetic and population genomic analyses of isolates from Brazil reveal that the previously "African" VNB lineage occurs naturally in the South American environment. This suggests migration of the VNB lineage between Africa and South America prior to its diversification, supported by finding ancestral recombination events between isolates from different lineages and regions. The results provide evidence of substantial population structure, with all lineages showing multi-continental distributions; demonstrating the highly dispersive nature of this pathogen.
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http://dx.doi.org/10.1534/genetics.117.203836DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5586382PMC
September 2017

Population genomics and the evolution of virulence in the fungal pathogen .

Genome Res 2017 07;27(7):1207-1219

Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA.

is an opportunistic fungal pathogen that causes approximately 625,000 deaths per year from nervous system infections. Here, we leveraged a unique, genetically diverse population of from sub-Saharan Africa, commonly isolated from mopane trees, to determine how selective pressures in the environment coincidentally adapted for human virulence. Genome sequencing and phylogenetic analysis of 387 isolates, representing the global VNI and African VNB lineages, highlighted a deep, nonrecombining split in VNB (herein, VNBI and VNBII). VNBII was enriched for clinical samples relative to VNBI, while phenotypic profiling of 183 isolates demonstrated that VNBI isolates were significantly more resistant to oxidative stress and more heavily melanized than VNBII isolates. Lack of melanization in both lineages was associated with loss-of-function mutations in the transcription factor. A genome-wide association study across all VNB isolates revealed sequence differences between clinical and environmental isolates in virulence factors and stress response genes. Inositol transporters and catabolism genes, which process sugars present in plants and the human nervous system, were identified as targets of selection in all three lineages. Further phylogenetic and population genomic analyses revealed extensive loss of genetic diversity in VNBI, suggestive of a history of population bottlenecks, along with unique evolutionary trajectories for mating type loci. These data highlight the complex evolutionary interplay between adaptation to natural environments and opportunistic infections, and that selection on specific pathways may predispose isolates to human virulence.
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http://dx.doi.org/10.1101/gr.218727.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5495072PMC
July 2017

Genome Diversity, Recombination, and Virulence across the Major Lineages of .

mSphere 2016 Sep-Oct;1(5). Epub 2016 Sep 28.

Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

The genus includes two species of thermally dimorphic fungi that cause paracoccidioidomycosis, a neglected health-threatening human systemic mycosis endemic to Latin America. To examine the genome evolution and the diversity of spp., we conducted whole-genome sequencing of 31 isolates representing the phylogenetic, geographic, and ecological breadth of the genus. These samples included clinical, environmental and laboratory reference strains of the S1, PS2, PS3, and PS4 lineages of and also isolates of species We completed the first annotated genome assemblies for the PS3 and PS4 lineages and found that gene order was highly conserved across the major lineages, with only a few chromosomal rearrangements Comparing whole-genome assemblies of the major lineages with single-nucleotide polymorphisms (SNPs) predicted from the remaining 26 isolates, we identified a deep split of the S1 lineage into two clades we named S1a and S1b. We found evidence for greater genetic exchange between the S1b lineage and all other lineages; this may reflect the broad geographic range of S1b, which is often sympatric with the remaining, largely geographically isolated lineages. In addition, we found evidence of positive selection for the and antigen genes and genes coding for other secreted proteins and proteases and lineage-specific loss-of-function mutations in cell wall and protease genes; these together may contribute to virulence and host immune response variation among natural isolates of spp. These insights into the recent evolutionary events highlight important differences between the lineages that could impact the distribution, pathogenicity, and ecology of . Characterization of genetic differences between lineages of the dimorphic human-pathogenic fungus can identify changes linked to important phenotypes and guide the development of new diagnostics and treatments. In this article, we compared genomes of 31 diverse isolates representing the major lineages of spp. and completed the first annotated genome sequences for the PS3 and PS4 lineages. We analyzed the population structure and characterized the genetic diversity among the lineages of , including a deep split of S1 into two lineages (S1a and S1b), and differentiated S1b, associated with most clinical cases, as the more highly recombining and diverse lineage. In addition, we found patterns of positive selection in surface proteins and secreted enzymes among the lineages, suggesting diversifying mechanisms of pathogenicity and adaptation across this species complex. These genetic differences suggest associations with the geographic range, pathogenicity, and ecological niches of lineages.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5040785PMC
http://dx.doi.org/10.1128/mSphere.00213-16DOI Listing
September 2016

Genome analysis of three Pneumocystis species reveals adaptation mechanisms to life exclusively in mammalian hosts.

Nat Commun 2016 Feb 22;7:10740. Epub 2016 Feb 22.

Critical Care Medicine Department, NIH Clinical Center, National Institutes of Health, Building 10, Room 2C145, 10 Center Drive, Bethesda, Maryland 20892, USA.

Pneumocystis jirovecii is a major cause of life-threatening pneumonia in immunosuppressed patients including transplant recipients and those with HIV/AIDS, yet surprisingly little is known about the biology of this fungal pathogen. Here we report near complete genome assemblies for three Pneumocystis species that infect humans, rats and mice. Pneumocystis genomes are highly compact relative to other fungi, with substantial reductions of ribosomal RNA genes, transporters, transcription factors and many metabolic pathways, but contain expansions of surface proteins, especially a unique and complex surface glycoprotein superfamily, as well as proteases and RNA processing proteins. Unexpectedly, the key fungal cell wall components chitin and outer chain N-mannans are absent, based on genome content and experimental validation. Our findings suggest that Pneumocystis has developed unique mechanisms of adaptation to life exclusively in mammalian hosts, including dependence on the lungs for gas and nutrients and highly efficient strategies to escape both host innate and acquired immune defenses.
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http://dx.doi.org/10.1038/ncomms10740DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4764891PMC
February 2016

Analysis of a food-borne fungal pathogen outbreak: virulence and genome of a Mucor circinelloides isolate from yogurt.

mBio 2014 Jul 8;5(4):e01390-14. Epub 2014 Jul 8.

Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, USA

Food-borne pathogens are ongoing problems, and new pathogens are emerging. The impact of fungi, however, is largely underestimated. Recently, commercial yogurts contaminated with Mucor circinelloides were sold, and >200 consumers became ill with nausea, vomiting, and diarrhea. Mucoralean fungi cause the fatal fungal infection mucormycosis, whose incidence has been continuously increasing. In this study, we isolated an M. circinelloides strain from a yogurt container, and multilocus sequence typing identified the strain as Mucor circinelloides f. circinelloides. M. circinelloides f. circinelloides is the most virulent M. circinelloides subspecies and is commonly associated with human infections, whereas M. circinelloides f. lusitanicus and M. circinelloides f. griseocyanus are less common causes of infection. Whole-genome analysis of the yogurt isolate confirmed it as being close to the M. circinelloides f. circinelloides subgroup, with a higher percentage of divergence with the M. circinelloides f. lusitanicus subgroup. In mating assays, the yogurt isolate formed sexual zygospores with the (-) M. circinelloides f. circinelloides tester strain, which is congruent with its sex locus encoding SexP, the (+) mating type sex determinant. The yogurt isolate was virulent in murine and wax moth larva host systems. In a murine gastromucormycosis model, Mucor was recovered from fecal samples of infected mice for up to 10 days, indicating that Mucor can survive transit through the GI tract. In interactions with human immune cells, M. circinelloides f. lusitanicus induced proinflammatory cytokines but M. circinelloides f. circinelloides did not, which may explain the different levels of virulence in mammalian hosts. This study demonstrates that M. circinelloides can spoil food products and cause gastrointestinal illness in consumers and may pose a particular risk to immunocompromised patients. Importance: The U.S. FDA reported that yogurt products were contaminated with M. circinelloides, a mucoralean fungal pathogen, and >200 consumers complained of symptoms, including vomiting, nausea, and diarrhea. The manufacturer voluntarily withdrew the affected yogurt products from the market. Compared to other food-borne pathogens, including bacteria, viruses, and parasites, less focus has been placed on the risk of fungal pathogens. This study evaluates the potential risk from the food-borne fungal pathogen M. circinelloides that was isolated from the contaminated commercial yogurt. We successfully cultured an M. circinelloides isolate and found that the isolate belongs to the species M. circinelloides f. circinelloides, which is often associated with human infections. In murine and insect host models, the isolate was virulent. While information disseminated in the popular press would suggest this fungal contaminant poses little or no risk to consumers, our results show instead that it is capable of causing significant infections in animals.
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http://dx.doi.org/10.1128/mBio.01390-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4161253PMC
July 2014

Comparative genomic and transcriptomic analysis of wangiella dermatitidis, a major cause of phaeohyphomycosis and a model black yeast human pathogen.

G3 (Bethesda) 2014 Apr 16;4(4):561-78. Epub 2014 Apr 16.

Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, Massachusetts 02142.

Black or dark brown (phaeoid) fungi cause cutaneous, subcutaneous, and systemic infections in humans. Black fungi thrive in stressful conditions such as intense light, high radiation, and very low pH. Wangiella (Exophiala) dermatitidis is arguably the most studied phaeoid fungal pathogen of humans. Here, we report our comparative analysis of the genome of W. dermatitidis and the transcriptional response to low pH stress. This revealed that W. dermatitidis has lost the ability to synthesize alpha-glucan, a cell wall compound many pathogenic fungi use to evade the host immune system. In contrast, W. dermatitidis contains a similar profile of chitin synthase genes as related fungi and strongly induces genes involved in cell wall synthesis in response to pH stress. The large portfolio of transporters may provide W. dermatitidis with an enhanced ability to remove harmful products as well as to survive on diverse nutrient sources. The genome encodes three independent pathways for producing melanin, an ability linked to pathogenesis; these are active during pH stress, potentially to produce a barrier to accumulated oxidative damage that might occur under stress conditions. In addition, a full set of fungal light-sensing genes is present, including as part of a carotenoid biosynthesis gene cluster. Finally, we identify a two-gene cluster involved in nucleotide sugar metabolism conserved with a subset of fungi and characterize a horizontal transfer event of this cluster between fungi and algal viruses. This work reveals how W. dermatitidis has adapted to stress and survives in diverse environments, including during human infections.
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http://dx.doi.org/10.1534/g3.113.009241DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4059230PMC
April 2014

Distinctive expansion of potential virulence genes in the genome of the oomycete fish pathogen Saprolegnia parasitica.

PLoS Genet 2013 Jun 13;9(6):e1003272. Epub 2013 Jun 13.

Broad Institute of MIT and Harvard, Cambridge, Massachusetts, United States of America.

Oomycetes in the class Saprolegniomycetidae of the Eukaryotic kingdom Stramenopila have evolved as severe pathogens of amphibians, crustaceans, fish and insects, resulting in major losses in aquaculture and damage to aquatic ecosystems. We have sequenced the 63 Mb genome of the fresh water fish pathogen, Saprolegnia parasitica. Approximately 1/3 of the assembled genome exhibits loss of heterozygosity, indicating an efficient mechanism for revealing new variation. Comparison of S. parasitica with plant pathogenic oomycetes suggests that during evolution the host cellular environment has driven distinct patterns of gene expansion and loss in the genomes of plant and animal pathogens. S. parasitica possesses one of the largest repertoires of proteases (270) among eukaryotes that are deployed in waves at different points during infection as determined from RNA-Seq data. In contrast, despite being capable of living saprotrophically, parasitism has led to loss of inorganic nitrogen and sulfur assimilation pathways, strikingly similar to losses in obligate plant pathogenic oomycetes and fungi. The large gene families that are hallmarks of plant pathogenic oomycetes such as Phytophthora appear to be lacking in S. parasitica, including those encoding RXLR effectors, Crinkler's, and Necrosis Inducing-Like Proteins (NLP). S. parasitica also has a very large kinome of 543 kinases, 10% of which is induced upon infection. Moreover, S. parasitica encodes several genes typical of animals or animal-pathogens and lacking from other oomycetes, including disintegrins and galactose-binding lectins, whose expression and evolutionary origins implicate horizontal gene transfer in the evolution of animal pathogenesis in S. parasitica.
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http://dx.doi.org/10.1371/journal.pgen.1003272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3681718PMC
June 2013

Sequencing and characterization of the complete mitochondrial genomes of three Pneumocystis species provide new insights into divergence between human and rodent Pneumocystis.

FASEB J 2013 May 7;27(5):1962-72. Epub 2013 Feb 7.

Critical Care Medicine Department, NIH Clinical Center, 10 Center Dr., Bethesda, MD 20892, USA.

Pneumocystis jirovecii is an important opportunistic pathogen associated with AIDS and other immunodeficient conditions. Currently, very little is known about its nuclear and mitochondrial genomes. In this study, we sequenced the complete mitochondrial genome (mtDNA) of this organism and its closely related species Pneumocystis carinii and Pneumocystis murina by a combination of sequencing technologies. Our study shows that P. carinii and P. murina mtDNA share a nearly identical number and order of genes in a linear configuration, whereas P. jirovecii has a circular mtDNA containing nearly the same set of genes but in a different order. Detailed studies of the mtDNA terminal structures of P. murina and P. carinii suggest a unique replication mechanism for linear mtDNA. Phylogenetic analysis supports a close association of Pneumocystis species with Taphrina, Saitoella, and Schizosaccharomyces, and divergence within Pneumocystis species, with P. murina and P. carinii being more closely related to each other than either is to P. jirovecii. Comparative analysis of four complete P. jirovecii mtDNA sequences in this study and previously reported mtDNA sequences for diagnosing and genotyping suggests that the current diagnostic and typing methods can be improved using the complete mtDNA data. The availability of the complete P. jirovecii mtDNA also opens the possibility of identifying new therapeutic targets.
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http://dx.doi.org/10.1096/fj.12-224444DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3633818PMC
May 2013

The malaria parasite Plasmodium vivax exhibits greater genetic diversity than Plasmodium falciparum.

Nat Genet 2012 Sep 5;44(9):1046-50. Epub 2012 Aug 5.

Broad Institute, Cambridge, Massachusetts, USA.

We sequenced and annotated the genomes of four P. vivax strains collected from disparate geographic locations, tripling the number of genome sequences available for this understudied parasite and providing the first genome-wide perspective of global variability in this species. We observe approximately twice as much SNP diversity among these isolates as we do among a comparable collection of isolates of P. falciparum, a malaria-causing parasite that results in higher mortality. This indicates a distinct history of global colonization and/or a more stable demographic history for P. vivax relative to P. falciparum, which is thought to have undergone a recent population bottleneck. The SNP diversity, as well as additional microsatellite and gene family variability, suggests a capacity for greater functional variation in the global population of P. vivax. These findings warrant a deeper survey of variation in P. vivax to equip disease interventions targeting the distinctive biology of this neglected but major pathogen.
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http://dx.doi.org/10.1038/ng.2373DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3432710PMC
September 2012

Optimizing read mapping to reference genomes to determine composition and species prevalence in microbial communities.

PLoS One 2012 13;7(6):e36427. Epub 2012 Jun 13.

The Genome Institute, Washington University School of Medicine, St Louis, Missouri, United States of America.

The Human Microbiome Project (HMP) aims to characterize the microbial communities of 18 body sites from healthy individuals. To accomplish this, the HMP generated two types of shotgun data: reference shotgun sequences isolated from different anatomical sites on the human body and shotgun metagenomic sequences from the microbial communities of each site. The alignment strategy for characterizing these metagenomic communities using available reference sequence is important to the success of HMP data analysis. Six next-generation aligners were used to align a community of known composition against a database comprising reference organisms known to be present in that community. All aligners report nearly complete genome coverage (>97%) for strains with over 6X depth of coverage, however they differ in speed, memory requirement and ease of use issues such as database size limitations and supported mapping strategies. The selected aligner was tested across a range of parameters to maximize sensitivity while maintaining a low false positive rate. We found that constraining alignment length had more impact on sensitivity than does constraining similarity in all cases tested. However, when reference species were replaced with phylogenetic neighbors, similarity begins to play a larger role in detection. We also show that choosing the top hit randomly when multiple, equally strong mappings are available increases overall sensitivity at the expense of taxonomic resolution. The results of this study identified a strategy that was used to map over 3 tera-bases of microbial sequence against a database of more than 5,000 reference genomes in just over a month.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0036427PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3374613PMC
December 2012

Genomic epidemiology of the Escherichia coli O104:H4 outbreaks in Europe, 2011.

Proc Natl Acad Sci U S A 2012 Feb 6;109(8):3065-70. Epub 2012 Feb 6.

Department of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA.

The degree to which molecular epidemiology reveals information about the sources and transmission patterns of an outbreak depends on the resolution of the technology used and the samples studied. Isolates of Escherichia coli O104:H4 from the outbreak centered in Germany in May-July 2011, and the much smaller outbreak in southwest France in June 2011, were indistinguishable by standard tests. We report a molecular epidemiological analysis using multiplatform whole-genome sequencing and analysis of multiple isolates from the German and French outbreaks. Isolates from the German outbreak showed remarkably little diversity, with only two single nucleotide polymorphisms (SNPs) found in isolates from four individuals. Surprisingly, we found much greater diversity (19 SNPs) in isolates from seven individuals infected in the French outbreak. The German isolates form a clade within the more diverse French outbreak strains. Moreover, five isolates derived from a single infected individual from the French outbreak had extremely limited diversity. The striking difference in diversity between the German and French outbreak samples is consistent with several hypotheses, including a bottleneck that purged diversity in the German isolates, variation in mutation rates in the two E. coli outbreak populations, or uneven distribution of diversity in the seed populations that led to each outbreak.
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http://dx.doi.org/10.1073/pnas.1121491109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3286951PMC
February 2012

Comparative genomic analysis of human fungal pathogens causing paracoccidioidomycosis.

PLoS Genet 2011 Oct 27;7(10):e1002345. Epub 2011 Oct 27.

Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

Paracoccidioides is a fungal pathogen and the cause of paracoccidioidomycosis, a health-threatening human systemic mycosis endemic to Latin America. Infection by Paracoccidioides, a dimorphic fungus in the order Onygenales, is coupled with a thermally regulated transition from a soil-dwelling filamentous form to a yeast-like pathogenic form. To better understand the genetic basis of growth and pathogenicity in Paracoccidioides, we sequenced the genomes of two strains of Paracoccidioides brasiliensis (Pb03 and Pb18) and one strain of Paracoccidioides lutzii (Pb01). These genomes range in size from 29.1 Mb to 32.9 Mb and encode 7,610 to 8,130 genes. To enable genetic studies, we mapped 94% of the P. brasiliensis Pb18 assembly onto five chromosomes. We characterized gene family content across Onygenales and related fungi, and within Paracoccidioides we found expansions of the fungal-specific kinase family FunK1. Additionally, the Onygenales have lost many genes involved in carbohydrate metabolism and fewer genes involved in protein metabolism, resulting in a higher ratio of proteases to carbohydrate active enzymes in the Onygenales than their relatives. To determine if gene content correlated with growth on different substrates, we screened the non-pathogenic onygenale Uncinocarpus reesii, which has orthologs for 91% of Paracoccidioides metabolic genes, for growth on 190 carbon sources. U. reesii showed growth on a limited range of carbohydrates, primarily basic plant sugars and cell wall components; this suggests that Onygenales, including dimorphic fungi, can degrade cellulosic plant material in the soil. In addition, U. reesii grew on gelatin and a wide range of dipeptides and amino acids, indicating a preference for proteinaceous growth substrates over carbohydrates, which may enable these fungi to also degrade animal biomass. These capabilities for degrading plant and animal substrates suggest a duality in lifestyle that could enable pathogenic species of Onygenales to transfer from soil to animal hosts.
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http://dx.doi.org/10.1371/journal.pgen.1002345DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3203195PMC
October 2011

Comparative functional genomics of the fission yeasts.

Science 2011 May 21;332(6032):930-6. Epub 2011 Apr 21.

Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605, USA.

The fission yeast clade--comprising Schizosaccharomyces pombe, S. octosporus, S. cryophilus, and S. japonicus--occupies the basal branch of Ascomycete fungi and is an important model of eukaryote biology. A comparative annotation of these genomes identified a near extinction of transposons and the associated innovation of transposon-free centromeres. Expression analysis established that meiotic genes are subject to antisense transcription during vegetative growth, which suggests a mechanism for their tight regulation. In addition, trans-acting regulators control new genes within the context of expanded functional modules for meiosis and stress response. Differences in gene content and regulation also explain why, unlike the budding yeast of Saccharomycotina, fission yeasts cannot use ethanol as a primary carbon source. These analyses elucidate the genome structure and gene regulation of fission yeast and provide tools for investigation across the Schizosaccharomyces clade.
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http://dx.doi.org/10.1126/science.1203357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3131103PMC
May 2011

A scalable, fully automated process for construction of sequence-ready human exome targeted capture libraries.

Genome Biol 2011 4;12(1):R1. Epub 2011 Jan 4.

Genome Sequencing Platform, Broad Institute of MIT and Harvard, Cambridge, MA 02141, USA.

Genome targeting methods enable cost-effective capture of specific subsets of the genome for sequencing. We present here an automated, highly scalable method for carrying out the Solution Hybrid Selection capture approach that provides a dramatic increase in scale and throughput of sequence-ready libraries produced. Significant process improvements and a series of in-process quality control checkpoints are also added. These process improvements can also be used in a manual version of the protocol.
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http://dx.doi.org/10.1186/gb-2011-12-1-r1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3091298PMC
February 2012

High-quality draft assemblies of mammalian genomes from massively parallel sequence data.

Proc Natl Acad Sci U S A 2011 Jan 27;108(4):1513-8. Epub 2010 Dec 27.

Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.

Massively parallel DNA sequencing technologies are revolutionizing genomics by making it possible to generate billions of relatively short (~100-base) sequence reads at very low cost. Whereas such data can be readily used for a wide range of biomedical applications, it has proven difficult to use them to generate high-quality de novo genome assemblies of large, repeat-rich vertebrate genomes. To date, the genome assemblies generated from such data have fallen far short of those obtained with the older (but much more expensive) capillary-based sequencing approach. Here, we report the development of an algorithm for genome assembly, ALLPATHS-LG, and its application to massively parallel DNA sequence data from the human and mouse genomes, generated on the Illumina platform. The resulting draft genome assemblies have good accuracy, short-range contiguity, long-range connectivity, and coverage of the genome. In particular, the base accuracy is high (≥99.95%) and the scaffold sizes (N50 size = 11.5 Mb for human and 7.2 Mb for mouse) approach those obtained with capillary-based sequencing. The combination of improved sequencing technology and improved computational methods should now make it possible to increase dramatically the de novo sequencing of large genomes. The ALLPATHS-LG program is available at http://www.broadinstitute.org/science/programs/genome-biology/crd.
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http://dx.doi.org/10.1073/pnas.1017351108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3029755PMC
January 2011

Population genomic sequencing of Coccidioides fungi reveals recent hybridization and transposon control.

Genome Res 2010 Jul 1;20(7):938-46. Epub 2010 Jun 1.

Broad Institute, Cambridge, Massachusetts 02142, USA.

We have sequenced the genomes of 18 isolates of the closely related human pathogenic fungi Coccidioides immitis and Coccidioides posadasii to more clearly elucidate population genomic structure, bringing the total number of sequenced genomes for each species to 10. Our data confirm earlier microsatellite-based findings that these species are genetically differentiated, but our population genomics approach reveals that hybridization and genetic introgression have recently occurred between the two species. The directionality of introgression is primarily from C. posadasii to C. immitis, and we find more than 800 genes exhibiting strong evidence of introgression in one or more sequenced isolates. We performed PCR-based sequencing of one region exhibiting introgression in 40 C. immitis isolates to confirm and better define the extent of gene flow between the species. We find more coding sequence than expected by chance in the introgressed regions, suggesting that natural selection may play a role in the observed genetic exchange. We find notable heterogeneity in repetitive sequence composition among the sequenced genomes and present the first detailed genome-wide profile of a repeat-induced point mutation (RIP) process distinctly different from what has been observed in Neurospora. We identify promiscuous HLA-I and HLA-II epitopes in both proteomes and discuss the possible implications of introgression and population genomic data for public health and vaccine candidate prioritization. This study highlights the importance of population genomic data for detecting subtle but potentially important phenomena such as introgression.
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http://dx.doi.org/10.1101/gr.103911.109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2892095PMC
July 2010

A catalog of reference genomes from the human microbiome.

Science 2010 May;328(5981):994-9

The human microbiome refers to the community of microorganisms, including prokaryotes, viruses, and microbial eukaryotes, that populate the human body. The National Institutes of Health launched an initiative that focuses on describing the diversity of microbial species that are associated with health and disease. The first phase of this initiative includes the sequencing of hundreds of microbial reference genomes, coupled to metagenomic sequencing from multiple body sites. Here we present results from an initial reference genome sequencing of 178 microbial genomes. From 547,968 predicted polypeptides that correspond to the gene complement of these strains, previously unidentified ("novel") polypeptides that had both unmasked sequence length greater than 100 amino acids and no BLASTP match to any nonreference entry in the nonredundant subset were defined. This analysis resulted in a set of 30,867 polypeptides, of which 29,987 (approximately 97%) were unique. In addition, this set of microbial genomes allows for approximately 40% of random sequences from the microbiome of the gastrointestinal tract to be associated with organisms based on the match criteria used. Insights into pan-genome analysis suggest that we are still far from saturating microbial species genetic data sets. In addition, the associated metrics and standards used by our group for quality assurance are presented.
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http://dx.doi.org/10.1126/science.1183605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2940224PMC
May 2010

Comparative genomics reveals mobile pathogenicity chromosomes in Fusarium.

Nature 2010 Mar;464(7287):367-73

The Broad Institute, Cambridge, Massachusetts 02141, USA.

Fusarium species are among the most important phytopathogenic and toxigenic fungi. To understand the molecular underpinnings of pathogenicity in the genus Fusarium, we compared the genomes of three phenotypically diverse species: Fusarium graminearum, Fusarium verticillioides and Fusarium oxysporum f. sp. lycopersici. Our analysis revealed lineage-specific (LS) genomic regions in F. oxysporum that include four entire chromosomes and account for more than one-quarter of the genome. LS regions are rich in transposons and genes with distinct evolutionary profiles but related to pathogenicity, indicative of horizontal acquisition. Experimentally, we demonstrate the transfer of two LS chromosomes between strains of F. oxysporum, converting a non-pathogenic strain into a pathogen. Transfer of LS chromosomes between otherwise genetically isolated strains explains the polyphyletic origin of host specificity and the emergence of new pathogenic lineages in F. oxysporum. These findings put the evolution of fungal pathogenicity into a new perspective.
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http://dx.doi.org/10.1038/nature08850DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3048781PMC
March 2010

Genome sequence and analysis of the Irish potato famine pathogen Phytophthora infestans.

Authors:
Brian J Haas Sophien Kamoun Michael C Zody Rays H Y Jiang Robert E Handsaker Liliana M Cano Manfred Grabherr Chinnappa D Kodira Sylvain Raffaele Trudy Torto-Alalibo Tolga O Bozkurt Audrey M V Ah-Fong Lucia Alvarado Vicky L Anderson Miles R Armstrong Anna Avrova Laura Baxter Jim Beynon Petra C Boevink Stephanie R Bollmann Jorunn I B Bos Vincent Bulone Guohong Cai Cahid Cakir James C Carrington Megan Chawner Lucio Conti Stefano Costanzo Richard Ewan Noah Fahlgren Michael A Fischbach Johanna Fugelstad Eleanor M Gilroy Sante Gnerre Pamela J Green Laura J Grenville-Briggs John Griffith Niklaus J Grünwald Karolyn Horn Neil R Horner Chia-Hui Hu Edgar Huitema Dong-Hoon Jeong Alexandra M E Jones Jonathan D G Jones Richard W Jones Elinor K Karlsson Sridhara G Kunjeti Kurt Lamour Zhenyu Liu Lijun Ma Daniel Maclean Marcus C Chibucos Hayes McDonald Jessica McWalters Harold J G Meijer William Morgan Paul F Morris Carol A Munro Keith O'Neill Manuel Ospina-Giraldo Andrés Pinzón Leighton Pritchard Bernard Ramsahoye Qinghu Ren Silvia Restrepo Sourav Roy Ari Sadanandom Alon Savidor Sebastian Schornack David C Schwartz Ulrike D Schumann Ben Schwessinger Lauren Seyer Ted Sharpe Cristina Silvar Jing Song David J Studholme Sean Sykes Marco Thines Peter J I van de Vondervoort Vipaporn Phuntumart Stephan Wawra Rob Weide Joe Win Carolyn Young Shiguo Zhou William Fry Blake C Meyers Pieter van West Jean Ristaino Francine Govers Paul R J Birch Stephen C Whisson Howard S Judelson Chad Nusbaum

Nature 2009 Sep 9;461(7262):393-8. Epub 2009 Sep 9.

Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02141, USA.

Phytophthora infestans is the most destructive pathogen of potato and a model organism for the oomycetes, a distinct lineage of fungus-like eukaryotes that are related to organisms such as brown algae and diatoms. As the agent of the Irish potato famine in the mid-nineteenth century, P. infestans has had a tremendous effect on human history, resulting in famine and population displacement. To this day, it affects world agriculture by causing the most destructive disease of potato, the fourth largest food crop and a critical alternative to the major cereal crops for feeding the world's population. Current annual worldwide potato crop losses due to late blight are conservatively estimated at $6.7 billion. Management of this devastating pathogen is challenged by its remarkable speed of adaptation to control strategies such as genetically resistant cultivars. Here we report the sequence of the P. infestans genome, which at approximately 240 megabases (Mb) is by far the largest and most complex genome sequenced so far in the chromalveolates. Its expansion results from a proliferation of repetitive DNA accounting for approximately 74% of the genome. Comparison with two other Phytophthora genomes showed rapid turnover and extensive expansion of specific families of secreted disease effector proteins, including many genes that are induced during infection or are predicted to have activities that alter host physiology. These fast-evolving effector genes are localized to highly dynamic and expanded regions of the P. infestans genome. This probably plays a crucial part in the rapid adaptability of the pathogen to host plants and underpins its evolutionary potential.
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http://dx.doi.org/10.1038/nature08358DOI Listing
September 2009

Genome sequence, comparative analysis and haplotype structure of the domestic dog.

Authors:
Kerstin Lindblad-Toh Claire M Wade Tarjei S Mikkelsen Elinor K Karlsson David B Jaffe Michael Kamal Michele Clamp Jean L Chang Edward J Kulbokas Michael C Zody Evan Mauceli Xiaohui Xie Matthew Breen Robert K Wayne Elaine A Ostrander Chris P Ponting Francis Galibert Douglas R Smith Pieter J DeJong Ewen Kirkness Pablo Alvarez Tara Biagi William Brockman Jonathan Butler Chee-Wye Chin April Cook James Cuff Mark J Daly David DeCaprio Sante Gnerre Manfred Grabherr Manolis Kellis Michael Kleber Carolyne Bardeleben Leo Goodstadt Andreas Heger Christophe Hitte Lisa Kim Klaus-Peter Koepfli Heidi G Parker John P Pollinger Stephen M J Searle Nathan B Sutter Rachael Thomas Caleb Webber Jennifer Baldwin Adal Abebe Amr Abouelleil Lynne Aftuck Mostafa Ait-Zahra Tyler Aldredge Nicole Allen Peter An Scott Anderson Claudel Antoine Harindra Arachchi Ali Aslam Laura Ayotte Pasang Bachantsang Andrew Barry Tashi Bayul Mostafa Benamara Aaron Berlin Daniel Bessette Berta Blitshteyn Toby Bloom Jason Blye Leonid Boguslavskiy Claude Bonnet Boris Boukhgalter Adam Brown Patrick Cahill Nadia Calixte Jody Camarata Yama Cheshatsang Jeffrey Chu Mieke Citroen Alville Collymore Patrick Cooke Tenzin Dawoe Riza Daza Karin Decktor Stuart DeGray Norbu Dhargay Kimberly Dooley Kathleen Dooley Passang Dorje Kunsang Dorjee Lester Dorris Noah Duffey Alan Dupes Osebhajajeme Egbiremolen Richard Elong Jill Falk Abderrahim Farina Susan Faro Diallo Ferguson Patricia Ferreira Sheila Fisher Mike FitzGerald Karen Foley Chelsea Foley Alicia Franke Dennis Friedrich Diane Gage Manuel Garber Gary Gearin Georgia Giannoukos Tina Goode Audra Goyette Joseph Graham Edward Grandbois Kunsang Gyaltsen Nabil Hafez Daniel Hagopian Birhane Hagos Jennifer Hall Claire Healy Ryan Hegarty Tracey Honan Andrea Horn Nathan Houde Leanne Hughes Leigh Hunnicutt M Husby Benjamin Jester Charlien Jones Asha Kamat Ben Kanga Cristyn Kells Dmitry Khazanovich Alix Chinh Kieu Peter Kisner Mayank Kumar Krista Lance Thomas Landers Marcia Lara William Lee Jean-Pierre Leger Niall Lennon Lisa Leuper Sarah LeVine Jinlei Liu Xiaohong Liu Yeshi Lokyitsang Tashi Lokyitsang Annie Lui Jan Macdonald John Major Richard Marabella Kebede Maru Charles Matthews Susan McDonough Teena Mehta James Meldrim Alexandre Melnikov Louis Meneus Atanas Mihalev Tanya Mihova Karen Miller Rachel Mittelman Valentine Mlenga Leonidas Mulrain Glen Munson Adam Navidi Jerome Naylor Tuyen Nguyen Nga Nguyen Cindy Nguyen Thu Nguyen Robert Nicol Nyima Norbu Choe Norbu Nathaniel Novod Tenchoe Nyima Peter Olandt Barry O'Neill Keith O'Neill Sahal Osman Lucien Oyono Christopher Patti Danielle Perrin Pema Phunkhang Fritz Pierre Margaret Priest Anthony Rachupka Sujaa Raghuraman Rayale Rameau Verneda Ray Christina Raymond Filip Rege Cecil Rise Julie Rogers Peter Rogov Julie Sahalie Sampath Settipalli Theodore Sharpe Terrance Shea Mechele Sheehan Ngawang Sherpa Jianying Shi Diana Shih Jessie Sloan Cherylyn Smith Todd Sparrow John Stalker Nicole Stange-Thomann Sharon Stavropoulos Catherine Stone Sabrina Stone Sean Sykes Pierre Tchuinga Pema Tenzing Senait Tesfaye Dawa Thoulutsang Yama Thoulutsang Kerri Topham Ira Topping Tsamla Tsamla Helen Vassiliev Vijay Venkataraman Andy Vo Tsering Wangchuk Tsering Wangdi Michael Weiand Jane Wilkinson Adam Wilson Shailendra Yadav Shuli Yang Xiaoping Yang Geneva Young Qing Yu Joanne Zainoun Lisa Zembek Andrew Zimmer Eric S Lander

Nature 2005 Dec;438(7069):803-19

Broad Institute of Harvard and MIT, 320 Charles Street, Cambridge, Massachusetts 02141, USA.

Here we report a high-quality draft genome sequence of the domestic dog (Canis familiaris), together with a dense map of single nucleotide polymorphisms (SNPs) across breeds. The dog is of particular interest because it provides important evolutionary information and because existing breeds show great phenotypic diversity for morphological, physiological and behavioural traits. We use sequence comparison with the primate and rodent lineages to shed light on the structure and evolution of genomes and genes. Notably, the majority of the most highly conserved non-coding sequences in mammalian genomes are clustered near a small subset of genes with important roles in development. Analysis of SNPs reveals long-range haplotypes across the entire dog genome, and defines the nature of genetic diversity within and across breeds. The current SNP map now makes it possible for genome-wide association studies to identify genes responsible for diseases and traits, with important consequences for human and companion animal health.
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http://dx.doi.org/10.1038/nature04338DOI Listing
December 2005

Nucleotide sequence based characterizations of two cryptic plasmids from the marine bacterium Ruegeria isolate PR1b.

Plasmid 2003 May;49(3):233-52

Division of Biology, University of California, San Diego, La Jolla, CA 92093-0322, USA.

Two plasmids, 76 and 148 kb in size, isolated from Ruegeria strain PR1b were entirely sequenced. These are the first plasmids to be characterized from this genus of marine bacteria. Sequence analysis revealed a biased distribution of function among the putative proteins encoded on the two plasmids. The smaller plasmid, designated pSD20, encodes a large number of putative proteins involved in polysaccharide biosynthesis and export. The larger plasmid, designated pSD25, primarily encodes putative proteins involved in the transport of small molecules and in DNA mobilization. Sequence analysis revealed uncommon potential replication systems on both plasmids. pSD25, the first repABC-type replicon isolated from the marine environment, actually contains two repABC-type replicons. pSD20 contains a complex replication region, including a replication origin and initiation protein similar to iteron-containing plasmids (such as pSW500 from the plant pathogen Erwinia stewartii) linked to putative RepA and RepB stabilization proteins of a repABC-type replicon and is highly homologous to a plasmid from the phototrophic bacterium Rhodobacter sphaeroides. Given the nature of the putative proteins encoded by both plasmids it is possible that these plasmids enhance the metabolic and physiological flexibility of the host bacterium, and thus its adaptation to the marine sediment environment.
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http://dx.doi.org/10.1016/s0147-619x(03)00014-3DOI Listing
May 2003