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September 22, 2019

Comparative genomics of Staphylococcus reveals determinants of speciation and diversification of antimicrobial defense.

The bacterial genus Staphylococcus comprises diverse species with most being described as colonizers of human and animal skin. A relational analysis of features that discriminate its species and contribute to niche adaptation and survival remains to be fully described. In this study, an interspecies, whole-genome comparative analysis of 21 Staphylococcus species was performed based on their orthologues. Three well-defined multi-species groups were identified: group A (including aureus/epidermidis); group B (including saprophyticus/xylosus) and group C (including pseudintermedius/delphini). The machine learning algorithm Random Forest was applied to prioritize orthologs that drive formation of the Staphylococcus species groups A-C. Orthologues driving staphylococcal intrageneric diversity comprised regulatory, metabolic and antimicrobial resistance proteins. Notably, the BraSR (NsaRS) two-component system (TCS) and its associated BraDE transporters that regulate antimicrobial resistance showed limited distribution in the genus and their presence was most closely associated with a subset of Staphylococcus species dominated by those that colonize human skin. Divergence of BraSR and GraSR antimicrobial peptide survival TCS and their associated transporters was observed across the staphylococci, likely reflecting niche specific evolution of these TCS/transporters and their specificities for AMPs. Experimental evolution, with selection for resistance to the lantibiotic nisin, revealed multiple routes to resistance and differences in the selection outcomes of the BraSR-positive species S. hominis and S. aureus. Selection supported a role for GraSR in nisin survival responses of the BraSR-negative species S. saprophyticus. Our study reveals diversification of antimicrobial-sensing TCS across the staphylococci and hints at differential relationships between GraSR and BraSR in those species positive for both TCS.

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September 22, 2019

Isolation, characterization, genomic sequencing, and GFP-marked insertional mutagenesis of a high-performance nitrogen-fixing bacterium, Kosakonia radicincitans GXGL-4A and visualization of bacterial colonization on cucumber roots.

A gram-negative bacterium GXGL-4A was originally isolated from maize roots. It displayed nitrogen-fixing (NF) ability under nitrogen-free culture condition, and had a significant promotion effect on cucumber growth in the pot inoculation test. The preliminary physiological and biochemical traits of GXGL-4A were characterized. Furthermore, a phylogenetic tree was constructed based on 16S ribosomal DNA (rDNA) sequences of genetically related species. To determine the taxonomic status of GXGL-4A and further utilize its nitrogen-fixing potential, genome sequence was obtained using PacBio RS II technology. The analyses of average nucleotide identity based on BLAST+ (ANIb) and correlation indexes of tetra-nucleotide signatures (Tetra) showed that the NF isolate GXGL-4A is closely related to the Kosakonia radicincitans type strain DSM 16656. Therefore, the isolate GXGL-4A was eventually classified into the species of Kosakonia radicincitans and designated K. radicincitans GXGL-4A. A high consistency in composition and gene arrangement of nitrogen-fixing gene cluster I (nif cluster I) was found between K. radicincitans GXGL-4A and other Kosakonia NF strains. The mutants tagged with green fluorescence protein (GFP) were obtained by transposon Tn5 mutagenesis, and then, the colonization of gfp-marked K. radicincitans GXGL-4A cells on cucumber seedling root were observed under fluorescence microscopy. The preferential sites of the labeled GXGL-4A cell population were the lateral root junctions, the differentiation zone, and the elongation zone. All these results should benefit for the deep exploration of nitrogen fixation mechanism of K. radicincitans GXGL-4A and will definitely facilitate the genetic modification process of this NF bacterium in sustainable agriculture.

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September 22, 2019

Genomic characterization reveals significant divergence within Chlorella sorokiniana (Chlorellales, Trebouxiophyceae)

Selection of highly productive algal strains is crucial for establishing economically viable biomass and biopro- duct cultivation systems. Characterization of algal genomes, including understanding strain-specific differences in genome content and architecture is a critical step in this process. Using genomic analyses, we demonstrate significant differences between three strains of Chlorella sorokiniana (strain 1228, UTEX 1230, and DOE1412). We found that unique, strain-specific genes comprise a substantial proportion of each genome, and genomic regions with> 80% local nucleotide identity constitute <15% of each genome among the strains, indicating substantial strain specific evolution. Furthermore, cataloging of meiosis and other sex-related genes in C. sor- okiniana strains suggests strategic breeding could be utilized to improve biomass and bioproduct yields if a sexual cycle can be characterized. Finally, preliminary investigation of epigenetic machinery suggests the pre- sence of potentially unique transcriptional regulation in each strain. Our data demonstrate that these three C. sorokiniana strains represent significantly different genomic content. Based on these findings, we propose in- dividualized assessment of each strain for potential performance in cultivation systems.

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September 22, 2019

Phylogenomics of colistin-susceptible and resistant XDR Acinetobacter baumannii.

Acinetobacter baumannii is a healthcare-associated pathogen with high rates of carbapenem resistance. Colistin is now routinely used for treatment of infections by this pathogen. However, colistin use has been associated with development of resistance to this agent.To elucidate the phylogenomics of colistin-susceptible and -resistant A. baumannii strain pairs from a cohort of hospitalized patients at a tertiary medical centre in the USA.WGS data from 21 pairs of colistin-susceptible and -resistant, XDR clinical strains were obtained and compared using phylogeny of aligned genome sequences, assessment of pairwise SNP differences and gene content.Fourteen patients had colistin-resistant strains that were highly genetically related to their own original susceptible strain with a median pairwise SNP distance of 5.5 (range 1-40 SNPs), while seven other strain pairs were divergent with =84 SNP differences. In addition, several strains from different patients formed distinct clusters on the phylogeny in keeping with closely linked transmission chains. The majority of colistin-resistant strains contained non-synonymous mutations within the pmrAB locus suggesting a central role for pmrAB mutations in colistin resistance. Excellent genotype-phenotype correlation was also observed for carbapenems, aminoglycosides and tetracyclines.The findings suggest that colistin resistance in the clinical setting arises through both in vivo evolution from colistin-susceptible strains and reinfection by unrelated colistin-resistant strains, the latter of which may involve patient-to-patient transmission.

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September 22, 2019

The genomic architecture and molecular evolution of ant odorant receptors.

The massive expansions of odorant receptor (OR) genes in ant genomes are notable examples of rapid genome evolution and adaptive gene duplication. However, the molecular mechanisms leading to gene family expansion remain poorly understood, partly because available ant genomes are fragmentary. Here, we present a highly contiguous, chromosome-level assembly of the clonal raider ant genome, revealing the largest known OR repertoire in an insect. While most ant ORs originate via local tandem duplication, we also observe several cases of dispersed duplication followed by tandem duplication in the most rapidly evolving OR clades. We found that areas of unusually high transposable element density (TE islands) were depauperate in ORs in the clonal raider ant, and found no evidence for retrotransposition of ORs. However, OR loci were enriched for transposons relative to the genome as a whole, potentially facilitating tandem duplication by unequal crossing over. We also found that ant OR genes are highly AT-rich compared to other genes. In contrast, in flies, OR genes are dispersed and largely isolated within the genome, and we find that fly ORs are not AT-rich. The genomic architecture and composition of ant ORs thus show convergence with the unrelated vertebrate ORs rather than the related fly ORs. This might be related to the greater gene numbers and/or potential similarities in gene regulation between ants and vertebrates as compared to flies.© 2018 McKenzie and Kronauer; Published by Cold Spring Harbor Laboratory Press.

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September 22, 2019

Thermus sediminis sp. nov., a thiosulfate-oxidizing and arsenate-reducing organism isolated from Little Hot Creek in the Long Valley Caldera, California.

Thermus species are widespread in natural and artificial thermal environments. Two new yellow-pigmented strains, L198T and L423, isolated from Little Hot Creek, a geothermal spring in eastern California, were identified as novel organisms belonging to the genus Thermus. Cells are Gram-negative, rod-shaped, and non-motile. Growth was observed at temperatures from 45 to 75 °C and at salinities of 0-2.0% added NaCl. Both strains grow heterotrophically or chemolithotrophically by oxidation of thiosulfate to sulfate. L198T and L423 grow by aerobic respiration or anaerobic respiration with arsenate as the terminal electron acceptor. Values for 16S rRNA gene identity (=?97.01%), digital DNA-DNA hybridization (=?32.7%), OrthoANI (=?87.5%), and genome-to-genome distance (0.13) values to all Thermus genomes were less than established criteria for microbial species. The predominant respiratory quinone was menaquinone-8 and the major cellular fatty acids were iso-C15:0, iso-C17:0 and anteiso-C15:0. One unidentified phospholipid (PL1) and one unidentified glycolipid (GL1) dominated the polar lipid pattern. The new strains could be differentiated from related taxa by ß-galactosidase and ß-glucosidase activity and the presence of hydroxy fatty acids. Based on phylogenetic, genomic, phenotypic, and chemotaxonomic evidence, the novel species Thermus sediminis sp. nov. is proposed, with the type strain L198T (=?CGMCC 1.13590T?=?KCTC XXX).

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September 22, 2019

Genome mining of Streptomyces xinghaiensis NRRL B-24674T for the discovery of the gene cluster involved in anticomplement activities and detection of novel xiamycin analogs.

Marine actinobacterium Streptomyces xinghaiensis NRRL B-24674T has been characterized as a novel species, but thus far, its biosynthetic potential remains unexplored. In this study, the high-quality genome sequence of S. xinghaiensis NRRL B-24674T was obtained, and the production of anticomplement agents, xiamycin analogs, and siderophores was investigated by genome mining. Anticomplement compounds are valuable for combating numerous diseases caused by the abnormal activation of the human complement system. The biosynthetic gene cluster (BGC) nrps1 resembles that of complestatins, which are potent microbial-derived anticomplement agents. The identification of the nrps1 BGC revealed a core peptide that differed from that in complestatin; thus, we studied the anticomplement activity of this strain. The culture broth of S. xinghaiensis NRRL B-24674T displayed good anticomplement activity. Subsequently, the disruption of the genes in the nrps1 BGC resulted in the loss of anticomplement activity, confirming the involvement of this BGC in the biosynthesis of anticomplement agents. In addition, the mining of the BGC tep5, which resembles that of the antiviral pentacyclic indolosesquiterpene xiamycin, resulted in the discovery of nine xiamycin analogs, including three novel compounds. In addition to the BGCs responsible for desferrioxamine B, neomycin, ectoine, and carotenoid, 18 BGCs present in the genome are predicted to be novel. The results of this study unveil the potential of S. xinghaiensis as a producer of novel anticomplement agents and provide a basis for further exploration of the biosynthetic potential of S. xinghaiensis NRRL B-24674T for the discovery of novel bioactive compounds by genome mining.

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September 22, 2019

Genomic evidence for asymmetric introgression by sexual selection in the common wall lizard.

Strongly selected characters can be transferred from one lineage to another with limited genetic exchange, resulting in asymmetric introgression and a mosaic genome in the receiving population. However, systems are rarely sufficiently well studied to link the pattern of introgression to its underlying process. Male common wall lizards in western Italy exhibit exaggeration of a suite of sexually selected characters that make them outcompete males from a distantly related lineage that lack these characters. This results in asymmetric hybridization and adaptive introgression of the suite of characters following secondary contact. We developed genomewide markers to infer the demographic history of gene flow between different genetic lineages, identify the spread of the sexually selected syndrome, and test the prediction that introgression should be asymmetric and heterogeneous across the genome. Our results show that secondary contact was accompanied by gene flow in both directions across most of the genome, but with approximately 3% of the genome showing highly asymmetric introgression in the predicted direction. Demographic simulations reveal that this asymmetric gene flow is more recent than the initial secondary contact, and the data suggest that the exaggerated male sexual characters originated within the Italian lineage and subsequently spread throughout this lineage before eventually reaching the contact zone. These results demonstrate that sexual selection can cause a suite of characters to spread throughout both closely and distantly related lineages with limited gene flow across the genome at large.© 2018 John Wiley & Sons Ltd.

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September 22, 2019

Genomic discovery of the hypsin gene and biosynthetic pathways for terpenoids in Hypsizygus marmoreus.

Hypsizygus marmoreus (Beech mushroom) is a popular ingredient in Asian cuisine. The medicinal effects of its bioactive compounds such as hypsin and hypsiziprenol have been reported, but the genetic basis or biosynthesis of these components is unknown.In this study, we sequenced a reference strain of H. marmoreus (Haemi 51,987-8). We evaluated various assembly strategies, and as a result the Allpaths and PBJelly produced the best assembly. The resulting genome was 42.7 Mbp in length and annotated with 16,627 gene models. A putative gene (Hypma_04324) encoding the antifungal and antiproliferative hypsin protein with 75% sequence identity with the previously known N-terminal sequence was identified. Carbohydrate active enzyme analysis displayed the typical feature of white-rot fungi where auxiliary activity and carbohydrate-binding modules were enriched. The genome annotation revealed four terpene synthase genes responsible for terpenoid biosynthesis. From the gene tree analysis, we identified that terpene synthase genes can be classified into six clades. Four terpene synthase genes of H. marmoreus belonged to four different groups that implies they may be involved in the synthesis of different structures of terpenes. A terpene synthase gene cluster was well-conserved in Agaricomycetes genomes, which contained known biosynthesis and regulatory genes.Genome sequence analysis of this mushroom led to the discovery of the hypsin gene. Comparative genome analysis revealed the conserved gene cluster for terpenoid biosynthesis in the genome. These discoveries will further our understanding of the biosynthesis of medicinal bioactive molecules in this edible mushroom.

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September 22, 2019

Comparative genomic and methylome analysis of non-virulent D74 and virulent Nagasaki Haemophilus parasuis isolates.

Haemophilus parasuis is a respiratory pathogen of swine and the etiological agent of Glässer’s disease. H. parasuis isolates can exhibit different virulence capabilities ranging from lethal systemic disease to subclinical carriage. To identify genomic differences between phenotypically distinct strains, we obtained the closed whole-genome sequence annotation and genome-wide methylation patterns for the highly virulent Nagasaki strain and for the non-virulent D74 strain. Evaluation of the virulence-associated genes contained within the genomes of D74 and Nagasaki led to the discovery of a large number of toxin-antitoxin (TA) systems within both genomes. Five predicted hemolysins were identified as unique to Nagasaki and seven putative contact-dependent growth inhibition toxin proteins were identified only in strain D74. Assessment of all potential vtaA genes revealed thirteen present in the Nagasaki genome and three in the D74 genome. Subsequent evaluation of the predicted protein structure revealed that none of the D74 VtaA proteins contain a collagen triple helix repeat domain. Additionally, the predicted protein sequence for two D74 VtaA proteins is substantially longer than any predicted Nagasaki VtaA proteins. Fifteen methylation sequence motifs were identified in D74 and fourteen methylation sequence motifs were identified in Nagasaki using SMRT sequencing analysis. Only one of the methylation sequence motifs was observed in both strains indicative of the diversity between D74 and Nagasaki. Subsequent analysis also revealed diversity in the restriction-modification systems harbored by D74 and Nagasaki. The collective information reported in this study will aid in the development of vaccines and intervention strategies to decrease the prevalence and disease burden caused by H. parasuis.

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September 22, 2019

A complete Leishmania donovani reference genome identifies novel genetic variations associated with virulence.

Leishmania donovani is responsible for visceral leishmaniasis, a neglected and lethal parasitic disease with limited treatment options and no vaccine. The study of L. donovani has been hindered by the lack of a high-quality reference genome and this can impact experimental outcomes including the identification of virulence genes, drug targets and vaccine development. We therefore generated a complete genome assembly by deep sequencing using a combination of second generation (Illumina) and third generation (PacBio) sequencing technologies. Compared to the current L. donovani assembly, the genome assembly reported within resulted in the closure over 2,000 gaps, the extension of several chromosomes up to telomeric repeats and the re-annotation of close to 15% of protein coding genes and the annotation of hundreds of non-coding RNA genes. It was possible to correctly assemble the highly repetitive A2 and Amastin virulence gene clusters. A comparative sequence analysis using the improved reference genome confirmed 70 published and identified 15 novel genomic differences between closely related visceral and atypical cutaneous disease-causing L. donovani strains providing a more complete map of genes associated with virulence and visceral organ tropism. Bioinformatic tools including protein variation effect analyzer and basic local alignment search tool were used to prioritize a list of potential virulence genes based on mutation severity, gene conservation and function. This complete genome assembly and novel information on virulence factors will support the identification of new drug targets and the development of a vaccine for L. donovani.

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September 22, 2019

Complete genome sequencing of Comamonas kerstersii 8943, a causative agent for peritonitis.

Because of poor differentiation among the members of genus Comamonas using phenotypic methods, human infections caused by C. kerstersii are sporadically reported in the literature. Here, we represent the first complete genome sequence of C. kerstersii 8943, which caused peritonitis in a patient with continuous ambulatory peritoneal dialysis (CAPD). The complete genome with no gaps was obtained using third-generation Pacific Biosciences (PacBio) RSII sequencing system with single-molecule real-time (SMRT) analysis. Protein-coding genes, rRNAs and tRNAs were predicted. Functional annotations of the genome using different databases revealed several genes related to pathogenicity including antibiotic resistance genes and prophages. Our work demonstrates that whole genome sequencing can enhance the resolution of clinical investigations and our data can be used as a reference genome during the rapid diagnosis of C. kerstersii infections in the future.

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September 22, 2019

Whole-genome sequencing of Chinese yellow catfish provides a valuable genetic resource for high-throughput identification of toxin genes.

Naturally derived toxins from animals are good raw materials for drug development. As a representative venomous teleost, Chinese yellow catfish (Pelteobagrus fulvidraco) can provide valuable resources for studies on toxin genes. Its venom glands are located in the pectoral and dorsal fins. Although with such interesting biologic traits and great value in economy, Chinese yellow catfish is still lacking a sequenced genome. Here, we report a high-quality genome assembly of Chinese yellow catfish using a combination of next-generation Illumina and third-generation PacBio sequencing platforms. The final assembly reached 714 Mb, with a contig N50 of 970 kb and a scaffold N50 of 3.65 Mb, respectively. We also annotated 21,562 protein-coding genes, in which 97.59% were assigned at least one functional annotation. Based on the genome sequence, we analyzed toxin genes in Chinese yellow catfish. Finally, we identified 207 toxin genes and classified them into three major groups. Interestingly, we also expanded a previously reported sex-related region (to ˜6 Mb) in the achieved genome assembly, and localized two important toxin genes within this region. In summary, we assembled a high-quality genome of Chinese yellow catfish and performed high-throughput identification of toxin genes from a genomic view. Therefore, the limited number of toxin sequences in public databases will be remarkably improved once we integrate multi-omics data from more and more sequenced species.

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September 22, 2019

The chromosome-level quality genome provides insights into the evolution of the biosynthesis genes for aroma compounds of Osmanthus fragrans.

Sweet osmanthus (Osmanthus fragrans) is a very popular ornamental tree species throughout Southeast Asia and USA particularly for its extremely fragrant aroma. We constructed a chromosome-level reference genome of O. fragrans to assist in studies of the evolution, genetic diversity, and molecular mechanism of aroma development. A total of over 118?Gb of polished reads was produced from HiSeq (45.1?Gb) and PacBio Sequel (73.35?Gb), giving 100× depth coverage for long reads. The combination of Illumina-short reads, PacBio-long reads, and Hi-C data produced the final chromosome quality genome of O. fragrans with a genome size of 727?Mb and a heterozygosity of 1.45 %. The genome was annotated using de novo and homology comparison and further refined with transcriptome data. The genome of O. fragrans was predicted to have?45,542 genes, of which 95.68 % were functionally annotated. Genome annotation found 49.35 % as the repetitive sequences, with long terminal repeats (LTR) being the richest (28.94 %). Genome evolution analysis indicated the evidence of whole-genome duplication 15 million years ago, which contributed to the current content of 45,242 genes. Metabolic analysis revealed that linalool, a monoterpene is the main aroma compound. Based on the genome and transcriptome, we further demonstrated the direct connection between terpene synthases (TPSs) and the rich aromatic molecules in O. fragrans. We identified three new flower-specific TPS genes, of which the expression coincided with the production of linalool. Our results suggest that the high number of TPS genes and the flower tissue- and stage-specific TPS genes expressions might drive the strong unique aroma production of O. fragrans.

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September 22, 2019

Genome sequence of the potato pathogenic fungus Alternaria solani HWC-168 reveals clues for its conidiation and virulence.

Alternaria solani is a known air-born deuteromycete fungus with a polycyclic life cycle and is the causal agent of early blight that causes significant yield losses of potato worldwide. However, the molecular mechanisms underlying the conidiation and pathogenicity remain largely unknown.We produced a high-quality genome assembly of A. solani HWC-168 that was isolated from a major potato-producing region of Northern China, which facilitated a comprehensive gene annotation, the accurate prediction of genes encoding secreted proteins and identification of conidiation-related genes. The assembled genome of A. solani HWC-168 has a genome size 32.8 Mb and encodes 10,358 predicted genes that are highly similar with related Alternaria species including Alternaria arborescens and Alternaria brassicicola. We identified conidiation-related genes in the genome of A. solani HWC-168 by searching for sporulation-related homologues identified from Aspergillus nidulans. A total of 975 secreted protein-encoding genes, which might act as virulence factors, were identified in the genome of A. solani HWC-168. The predicted secretome of A. solani HWC-168 possesses 261 carbohydrate-active enzymes (CAZy), 119 proteins containing RxLx[EDQ] motif and 27 secreted proteins unique to A. solani.Our findings will facilitate the identification of conidiation- and virulence-related genes in the genome of A. solani. This will permit new insights into understanding the molecular mechanisms underlying the A. solani-potato pathosystem and will add value to the global fungal genome database.

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