The vaginal bacterial strain AB70, belonging to the species Lactobacillus crispatus, was isolated from a vaginal swab from a healthy pregnant Korean woman. Here, we report the 2.37-Mb complete genome sequence of this strain. Copyright © 2019 Chang et al.
Background A high-quality reference genome is an essential tool for applied and basic research on arthropods. Long-read sequencing technologies may be used to generate more complete and contiguous genome assemblies than alternate technologies; however, long-read methods have historically had greater input DNA requirements and higher costs than next-generation sequencing, which are barriers to their use on many samples. Here, we present a 2.3 Gb de novo genome assembly of a field-collected adult female spotted lanternfly (Lycorma delicatula) using a single Pacific Biosciences SMRT Cell. The spotted lanternfly is an invasive species recently discovered in the northeastern United States that threatens to damage economically important crop plants in the region. Results The DNA from 1 individual was used to make 1 standard, size-selected library with an average DNA fragment size of ~20 kb. The library was run on 1 Sequel II SMRT Cell 8M, generating a total of 132 Gb of long-read sequences, of which 82 Gb were from unique library molecules, representing ~36× coverage of the genome. The assembly had high contiguity (contig N50 length = 1.5 Mb), completeness, and sequence level accuracy as estimated by conserved gene set analysis (96.8% of conserved genes both complete and without frame shift errors). Furthermore, it was possible to segregate more than half of the diploid genome into the 2 separate haplotypes. The assembly also recovered 2 microbial symbiont genomes known to be associated with L. delicatula, each microbial genome being assembled into a single contig. Conclusions We demonstrate that field-collected arthropods can be used for the rapid generation of high-quality genome assemblies, an attractive approach for projects on emerging invasive species, disease vectors, or conservation efforts of endangered species.
Enteropathogenic Escherichia coli (EPEC) is a leading cause of moderate to severe diarrhea among young children in developing countries, and EPEC isolates can be subdivided into two groups. Typical EPEC (tEPEC) bacteria are characterized by the presence of both the locus of enterocyte effacement (LEE) and the plasmid-encoded bundle-forming pilus (BFP), which are involved in adherence and translocation of type III effectors into the host cells. Atypical EPEC (aEPEC) bacteria also contain the LEE but lack the BFP. In the current report, we describe the complete genome of outbreak-associated aEPEC isolate E110019, which carries four plasmids. Comparative genomic analysis demonstrated that the type III secreted effector EspT gene, an autotransporter gene, a hemolysin gene, and putative fimbrial genes are all carried on plasmids. Further investigation of 65 espT-containing E. coli genomes demonstrated that different espT alleles are associated with multiple plasmids that differ in their overall gene content from the E110019 espT-containing plasmid. EspT has been previously described with respect to its role in the ability of E110019 to invade host cells. While other type III secreted effectors of E. coli have been identified on insertion elements and prophages of the chromosome, we demonstrated in the current study that the espT gene is located on multiple unique plasmids. These findings highlight a role of plasmids in dissemination of a unique E. coli type III secreted effector that is involved in host invasion and severe diarrheal illness.Copyright © 2019 American Society for Microbiology.
HIV elite controllers represent a remarkable minority of patients who maintain normal CD4+ T-cell counts and low or undetectable viral loads for decades in the absence of antiretroviral therapy. To examine the possible contribution of virus attenuation to elite control, we obtained a primary HIV-1 isolate from an elite controller who had been infected for 19?years, the last 10 of which were in the absence of antiretroviral therapy. Full-length sequencing of this isolate revealed a highly unusual V1 domain in Envelope (Env). The V1 domain in this HIV-1 strain was 49 amino acids, placing it in the top 1% of lengths among the 6,112 Env sequences in the Los Alamos National Laboratory online database. Furthermore, it included two additional N-glycosylation sites and a pair of cysteines suggestive of an extra disulfide loop. Virus with this Env retained good infectivity and replicative capacity; however, analysis of recombinant viruses suggested that other sequences in Env were adapted to accommodate the unusual V1 domain. While the long V1 domain did not confer resistance to neutralization by monoclonal antibodies of the V1/V2-glycan-dependent class, it did confer resistance to neutralization by monoclonal antibodies of the V3-glycan-dependent class. Our findings support results in the literature that suggest a role for long V1 regions in shielding HIV-1 from recognition by V3-directed broadly neutralizing antibodies. In the case of the elite controller described here, it seems likely that selective pressures from the humoral immune system were responsible for driving the highly unusual polymorphisms present in this HIV-1 Envelope.IMPORTANCE Elite controllers have long provided an avenue for researchers to reveal mechanisms underlying control of HIV-1. While the role of host genetic factors in facilitating elite control is well known, the possibility of infection by attenuated strains of HIV-1 has been much less studied. Here we describe an unusual viral feature found in an elite controller of HIV-1 infection and demonstrate its role in conferring escape from monoclonal antibodies of the V3-glycan class. Our results suggest that extreme variation may be needed by HIV-1 to escape neutralization by some antibody specificities. Copyright © 2019 Silver et al.
The Chinese chestnut (Castanea mollissima) is widely cultivated in China for nut production. This plant also plays an important ecological role in afforestation and ecosystem services. To facilitate and expand the use of C. mollissima for breeding and its genetic improvement, we report here the whole-genome sequence of C. mollissima.We produced a high-quality assembly of the C. mollissima genome using Pacific Biosciences single-molecule sequencing. The final draft genome is ~785.53 Mb long, with a contig N50 size of 944 kb, and we further annotated 36,479 protein-coding genes in the genome. Phylogenetic analysis showed that C. mollissima diverged from Quercus robur, a member of the Fagaceae family, ~13.62 million years ago.The high-quality whole-genome assembly of C. mollissima will be a valuable resource for further genetic improvement and breeding for disease resistance and nut quality. © The Author(s) 2019. Published by Oxford University Press.
Salmonella genomic island 3 (SGI3) was first described as a chromosomal island in Salmonella 4,[5],12:i:-, a monophasic variant of Salmonella enterica subsp. enterica serovar Typhimurium. The SGI3 DNA sequence detected from Salmonella 4,[5],12:i:- isolated in Japan was identical to that of a previously reported one across entire length of 81?kb. SGI3 consists of 86 open reading frames, including a copper homeostasis and silver resistance island (CHASRI) and an arsenic tolerance operon, in addition to genes related to conjugative transfer and DNA replication or partitioning, suggesting that the island is a mobile genetic element. We successfully selected transconjugants that acquired SGI3 after filter-mating experiments using the S. enterica serovars Typhimurium, Heidelberg, Hadar, Newport, Cerro, and Thompson as recipients. Southern blot analysis using I-CeuI-digested genomic DNA demonstrated that SGI3 was integrated into a chromosomal fragment of the transconjugants. PCR and sequencing analysis demonstrated that SGI3 was inserted into the 3′ end of the tRNA genes pheV or pheR The length of the target site was 52 or 55?bp, and a 55-bp attI sequence indicating generation of the circular form of SGI3 was also detected. The transconjugants had a higher MIC against CuSO4 compared to the recipient strains under anaerobic conditions. Tolerance was defined by the cus gene cluster in the CHASRI. The transconjugants also had distinctly higher MICs against Na2HAsO4 compared to recipient strains under aerobic conditions. These findings clearly demonstrate that SGI3 is an integrative and conjugative element and contributes to the copper and arsenic tolerance of S. enterica.Copyright © 2019 American Society for Microbiology.
Triatoma rubrofasciata is a widespread pathogen vector for Chagas disease, an illness that affects approximately 7 million people worldwide. Despite its importance to human health, its evolutionary origin has not been conclusively determined. A reference genome for T. rubrofasciata is not yet available.We have sequenced the genome of a female individual with T. rubrofasciatausing a single molecular DNA sequencing technology (i.e., PacBio Sequel platform) and have successfully reconstructed a whole-genome (680-Mb) assembly that covers 90% of the nuclear genome (757 Mb). Through Hi-C analysis, we have reconstructed full-length chromosomes of this female individual that has 13 unique chromosomes (2n = 24 = 22 + X1 + X2) with a contig N50 of 2.72 Mb and a scaffold N50 of 50.7 Mb. This genome has achieved a high base-level accuracy of 99.99%. This platinum-grade genome assembly has 12,691 annotated protein-coding genes. More than 95.1% of BUSCO genes were single-copy completed, indicating a high level of completeness of the genome.The platinum-grade genome assembly and its annotation provide valuable information for future in-depth comparative genomics studies, including sexual determination analysis in T. rubrofasciata and the pathogenesis of Chagas disease. © The Author(s) 2019. Published by Oxford University Press.
The blood clam, Scapharca (Anadara) broughtonii, is an economically and ecologically important marine bivalve of the family Arcidae. Efforts to study their population genetics, breeding, cultivation, and stock enrichment have been somewhat hindered by the lack of a reference genome. Herein, we report the complete genome sequence of S. broughtonii, a first reference genome of the family Arcidae.A total of 75.79 Gb clean data were generated with the Pacific Biosciences and Oxford Nanopore platforms, which represented approximately 86× coverage of the S. broughtonii genome. De novo assembly of these long reads resulted in an 884.5-Mb genome, with a contig N50 of 1.80 Mb and scaffold N50 of 45.00 Mb. Genome Hi-C scaffolding resulted in 19 chromosomes containing 99.35% of bases in the assembled genome. Genome annotation revealed that nearly half of the genome (46.1%) is composed of repeated sequences, while 24,045 protein-coding genes were predicted and 84.7% of them were annotated.We report here a chromosomal-level assembly of the S. broughtonii genome based on long-read sequencing and Hi-C scaffolding. The genomic data can serve as a reference for the family Arcidae and will provide a valuable resource for the scientific community and aquaculture sector. © The Author(s) 2019. Published by Oxford University Press.
In the human gut, Clostridium scindens ATCC 35704 is a predominant bacterium and one of the major bile acid 7a-dehydroxylating anaerobes. While this organism is well-studied relative to bile acid metabolism, little is known about the basic nutrition and physiology of C. scindens ATCC 35704. To determine the amino acid and vitamin requirements of C. scindens, the leave-one-out (one amino acid group or vitamin) technique was used to eliminate the nonessential amino acids and vitamins. With this approach, the amino acid tryptophan and three vitamins (riboflavin, pantothenate, and pyridoxal) were found to be required for the growth of C. scindens In the newly developed defined medium, C. scindens fermented glucose mainly to ethanol, acetate, formate, and H2. The genome of C. scindens ATCC 35704 was completed through PacBio sequencing. Pathway analysis of the genome sequence coupled with transcriptome sequencing (RNA-Seq) under defined culture conditions revealed consistency with the growth requirements and end products of glucose metabolism. Induction with bile acids revealed complex and differential responses to cholic acid and deoxycholic acid, including the expression of potentially novel bile acid-inducible genes involved in cholic acid metabolism. Responses to toxic deoxycholic acid included expression of genes predicted to be involved in DNA repair, oxidative stress, cell wall maintenance/metabolism, chaperone synthesis, and downregulation of one-third of the genome. These analyses provide valuable insight into the overall biology of C. scindens which may be important in treatment of disease associated with increased colonic secondary bile acids.IMPORTANCEC. scindens is one of a few identified gut bacterial species capable of converting host cholic acid into disease-associated secondary bile acids such as deoxycholic acid. The current work represents an important advance in understanding the nutritional requirements and response to bile acids of the medically important human gut bacterium, C. scindens ATCC 35704. A defined medium has been developed which will further the understanding of bile acid metabolism in the context of growth substrates, cofactors, and other metabolites in the vertebrate gut. Analysis of the complete genome supports the nutritional requirements reported here. Genome-wide transcriptomic analysis of gene expression in the presence of cholic acid and deoxycholic acid provides a unique insight into the complex response of C. scindens ATCC 35704 to primary and secondary bile acids. Also revealed are genes with the potential to function in bile acid transport and metabolism.Copyright © 2019 American Society for Microbiology.
Yellowhorn (Xanthoceras sorbifolium) is a species of the Sapindaceae family native to China and is an oil tree that can withstand cold and drought conditions. A pseudomolecule-level genome assembly for this species will not only contribute to understanding the evolution of its genes and chromosomes but also bring yellowhorn breeding into the genomic era.Here, we generated 15 pseudomolecules of yellowhorn chromosomes, on which 97.04% of scaffolds were anchored, using the combined Illumina HiSeq, Pacific Biosciences Sequel, and Hi-C technologies. The length of the final yellowhorn genome assembly was 504.2 Mb with a contig N50 size of 1.04 Mb and a scaffold N50 size of 32.17 Mb. Genome annotation revealed that 68.67% of the yellowhorn genome was composed of repetitive elements. Gene modelling predicted 24,672 protein-coding genes. By comparing orthologous genes, the divergence time of yellowhorn and its close sister species longan (Dimocarpus longan) was estimated at ~33.07 million years ago. Gene cluster and chromosome synteny analysis demonstrated that the yellowhorn genome shared a conserved genome structure with its ancestor in some chromosomes.This genome assembly represents a high-quality reference genome for yellowhorn. Integrated genome annotations provide a valuable dataset for genetic and molecular research in this species. We did not detect whole-genome duplication in the genome. The yellowhorn genome carries syntenic blocks from ancient chromosomes. These data sources will enable this genome to serve as an initial platform for breeding better yellowhorn cultivars. © The Author(s) 2019. Published by Oxford University Press.
The mycoparasitic fungus Hypomyces perniciosus causes wet bubble disease of mushrooms, particularly Agaricus bisporus. The genome of a highly virulent strain of H. perniciosus HP10 was sequenced and compared to three other fungi from the order Hypocreales that cause disease on A. bisporus. H. perniciosus genome is ~44 Mb, encodes 10,077 genes and enriched with transposable elements up to 25.3%. Phylogenetic analysis revealed that H. perniciosus is closely related to Cladobotryum protrusum and diverged from their common ancestor ~156.7 million years ago. H. perniciosus has few secreted proteins compared to C. protrusum and Trichoderma virens, but significantly expanded protein families of transporters, protein kinases, CAZymes (GH 18), peptidases, cytochrome P450, and SMs that are essential for mycoparasitism and adaptation to harsh environments. This study provides insights into H. perniciosus evolution and pathogenesis and will contribute to the development of effective disease management strategies to control wet bubble disease.
The pathogenic extended-spectrum-beta-lactamase (ESBL)-producing Escherichia coli lineage ST648 is increasingly reported from multiple origins. Our study of a large and global ST648 collection from various hosts (87 whole-genome sequences) combining core and accessory genomics with functional analyses and in vivo experiments suggests that ST648 is a nascent and generalist lineage, lacking clear phylogeographic and host association signals. By including large numbers of ST131 (n?=?107) and ST10 (n?=?96) strains for comparative genomics and phenotypic analysis, we demonstrate that the combination of multidrug resistance and high-level virulence are the hallmarks of ST648, similar to international high-risk clonal lineage ST131. Specifically, our in silico, in vitro, and in vivo results demonstrate that ST648 is well equipped with biofilm-associated features, while ST131 shows sophisticated signatures indicative of adaption to urinary tract infection, potentially conveying individual ecological niche adaptation. In addition, we used a recently developed NFDS (negative frequency-dependent selection) population model suggesting that ST648 will increase significantly in frequency as a cause of bacteremia within the next few years. Also, ESBL plasmids impacting biofilm formation aided in shaping and maintaining ST648 strains to successfully emerge worldwide across different ecologies. Our study contributes to understanding what factors drive the evolution and spread of emerging international high-risk clonal lineages.Copyright © 2019 American Society for Microbiology.
Haemophilus influenzae exclusively colonizes the human nasopharynx and can cause a variety of respiratory infections as well as invasive diseases, including meningitis and sepsis. A key virulence determinant of H. influenzae is the polysaccharide capsule, of which six serotypes are known, each encoded by a distinct variation of the capsule biosynthesis locus (cap-a to cap-f). H. influenzae type b (Hib) was historically responsible for the majority of invasive H. influenzae disease, and its prevalence has been markedly reduced in countries that have implemented vaccination programs targeting this serotype. In the postvaccine era, nontypeable H. influenzae emerged as the most dominant group causing disease, but in recent years a resurgence of encapsulated H. influenzae strains has also been observed, most notably serotype a. Given the increasing incidence of encapsulated strains and the high frequency of Hib in countries without vaccination programs, there is growing interest in genomic epidemiology of H. influenzae Here we present hicap, a software tool for rapid in silico serotype prediction from H. influenzae genome sequences. hicap is written using Python3 and is freely available at https://github.com/scwatts/hicap under the GNU General Public License v3 (GPL3). To demonstrate the utility of hicap, we used it to investigate the cap locus diversity and distribution in 691 high-quality H. influenzae genomes from GenBank. These analyses identified cap loci in 95 genomes and confirmed the general association of each serotype with a unique clonal lineage, and they also identified occasional recombination between lineages that gave rise to hybrid cap loci (2% of encapsulated strains).Copyright © 2019 Watts and Holt.
Anopheles funestus is one of the 3 most consequential and widespread vectors of human malaria in tropical Africa. However, the lack of a high-quality reference genome has hindered the association of phenotypic traits with their genetic basis in this important mosquito.Here we present a new high-quality A. funestus reference genome (AfunF3) assembled using 240× coverage of long-read single-molecule sequencing for contigging, combined with 100× coverage of short-read Hi-C data for chromosome scaffolding. The assembled contigs total 446 Mbp of sequence and contain substantial duplication due to alternative alleles present in the sequenced pool of mosquitos from the FUMOZ colony. Using alignment and depth-of-coverage information, these contigs were deduplicated to a 211 Mbp primary assembly, which is closer to the expected haploid genome size of 250 Mbp. This primary assembly consists of 1,053 contigs organized into 3 chromosome-scale scaffolds with an N50 contig size of 632 kbp and an N50 scaffold size of 93.811 Mbp, representing a 100-fold improvement in continuity versus the current reference assembly, AfunF1.This highly contiguous and complete A. funestus reference genome assembly will serve as an improved basis for future studies of genomic variation and organization in this important disease vector. © The Author(s) 2019. Published by Oxford University Press.
Several emerging pathogens have arisen as a result of selection pressures exerted by modern health care. Klebsiella quasipneumoniae was recently defined as a new species, yet its prevalence, niche, and propensity to acquire antimicrobial resistance genes are not fully described. We have been tracking inter- and intraspecies transmission of the Klebsiella pneumoniae carbapenemase (KPC) gene, blaKPC, between bacteria isolated from a single institution. We applied a combination of Illumina and PacBio whole-genome sequencing to identify and compare K. quasipneumoniae from patients and the hospital environment over 10- and 5-year periods, respectively. There were 32 blaKPC-positive K. quasipneumoniae isolates, all of which were identified as K. pneumoniae in the clinical microbiology laboratory, from 8 patients and 11 sink drains, with evidence for seven separate blaKPC plasmid acquisitions. Analysis of a single subclade of K. quasipneumoniae subsp. quasipneumoniae (n?=?23 isolates) from three patients and six rooms demonstrated seeding of a sink by a patient, subsequent persistence of the strain in the hospital environment, and then possible transmission to another patient. Longitudinal analysis of this strain demonstrated the acquisition of two unique blaKPC plasmids and then subsequent within-strain genetic rearrangement through transposition and homologous recombination. Our analysis highlights the apparent molecular propensity of K. quasipneumoniae to persist in the environment as well as acquire carbapenemase plasmids from other species and enabled an assessment of the genetic rearrangements which may facilitate horizontal transmission of carbapenemases. Copyright © 2019 Mathers et al.
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