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July 7, 2019

Complete genome sequencing of Arachidicoccus ginsenosidimutans sp. nov., and its application for production of minor ginsenosides by finding a novel ginsenoside-transforming beta-glucosidase

A novel bacterial strain (BS20T), which has ginsenoside-transforming ability, was whole genome sequenced for the identification of a target gene. After complete genome sequencing, phylogenetic, phenotypic and chemotaxonomic analyses, the strain BS20T (Arachidicoccus ginsenosidimutans sp. nov.) was placed within the genus Arachidicoccus of family Chitinophagaceae. The complete genome of strain BS20T comprised a circular chromosome of 4[thin space (1/6-em)]138[thin space (1/6-em)]017 bp. To find the target functional gene, 17 sets of four different glycoside hydrolases were cloned in E. coli BL21 (DE3) using the pGEX4T-1 vector and were characterized. Among these 17 sets of clones, only one, BglAg-762, exhibited ginsenoside-conversion ability. The BglAg-762 comprised 762 amino acid residues and belonged to the glycoside hydrolase family 3. The recombinant enzyme (GST-BglAg-762) was able to convert major ginsenosides Rb1 to F2 via gypenoside-XVII (Gyp-XVII), Rb2 to C-O, and Rb3, Rc, Rd, and Gyp-XVII to C-Mx1, C-Mc1, and F2, respectively. Finally, ginsenoside F2 was transformed into compound K (C-K). Besides, these pilot data demonstrate the identification of 17 sets of target/functional genes of 4 different glycoside hydrolases from a novel bacterial species via whole genome sequencing. Our results have shown that the recombinant BglAg-762 very quickly converts the major ginsenosides into minor ginsenosides, which can be used for the enhanced production of target minor ginsenosides. Furthermore, the web service of NCBI is suitable for any targeted gene identification, but based on our experimental analysis we concluded that the hypothetical protein present in NCBI should be considered as a putative or uncharacterized protein.

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July 7, 2019

Draft genomes of the fungal pathogen Phellinus noxius in Hong Kong

The fungal pathogen Phellinus noxius is the underlying cause of brown root rot, a disease with causing tree mortality globally, causing extensive damage in urban areas and crop plants. This disease currently has no cure, and despite the global epidemic, little is known about the pathogenesis and virulence of this pathogen. Using Ion Torrent PGM, Illumina MiSeq and PacBio RSII sequencing platforms with various genome assembly methods, we produced the draft genome sequences of four P. noxius strains isolated from infected trees in Hong Kong to further understand the pathogen and identify the mechanisms behind the aggressive nature and virulence of this fungus. The resulting genomes ranged from 30.8Mb to 31.8Mb in size, and of the four sequences, the YTM97 strain was chosen to produce a high-quality Hong Kong strain genome sequence, resulting in a 31Mb final assembly with 457 scaffolds, an N50 length of 275,889 bp and 96.2% genome completeness. RNA-seq of YTM97 using Illumina HiSeq400 was performed for improved gene prediction. AUGUSTUS and Genemark-ES prediction programs predicted 9,887 protein-coding genes which were annotated using GO and Pfam databases. The encoded carbohydrate active enzymes revealed large numbers of lignolytic enzymes present, comparable to those of other white-rot plant pathogens. In addition, P. noxius also possessed larger numbers of cellulose, xylan and hemicellulose degrading enzymes than other plant pathogens. Searches for virulence genes was also performed using PHI-Base and DFVF databases revealing a host of virulence-related genes and effectors. The combination of non-specific host range, unique carbohydrate active enzyme profile and large amount of putative virulence genes could explain the reasons behind the aggressive nature and increased virulence of this plant pathogen. The draft genome sequences presented here will provide references for strains found in Hong Kong. Together with emerging research, this information could be used for genetic diversity and epidemiology research on a global scale as well as expediting our efforts towards discovering the mechanisms of pathogenicity of this devastating pathogen.

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July 7, 2019

Comparative and population genomic landscape of Phellinus noxius: A hypervariable fungus causing root rot in trees.

The order Hymenochaetales of white rot fungi contain some of the most aggressive wood decayers causing tree deaths around the world. Despite their ecological importance and the impact of diseases they cause, little is known about the evolution and transmission patterns of these pathogens. Here, we sequenced and undertook comparative genomic analyses of Hymenochaetales genomes using brown root rot fungus Phellinus noxius, wood-decomposing fungus Phellinus lamaensis, laminated root rot fungus Phellinus sulphurascens and trunk pathogen Porodaedalea pini. Many gene families of lignin-degrading enzymes were identified from these fungi, reflecting their ability as white rot fungi. Comparing against distant fungi highlighted the expansion of 1,3-beta-glucan synthases in P. noxius, which may account for its fast-growing attribute. We identified 13 linkage groups conserved within Agaricomycetes, suggesting the evolution of stable karyotypes. We determined that P. noxius has a bipolar heterothallic mating system, with unusual highly expanded ~60 kb A locus as a result of accumulating gene transposition. We investigated the population genomics of 60 P. noxius isolates across multiple islands of the Asia Pacific region. Whole-genome sequencing showed this multinucleate species contains abundant poly-allelic single nucleotide polymorphisms with atypical allele frequencies. Different patterns of intra-isolate polymorphism reflect mono-/heterokaryotic states which are both prevalent in nature. We have shown two genetically separated lineages with one spanning across many islands despite the geographical barriers. Both populations possess extraordinary genetic diversity and show contrasting evolutionary scenarios. These results provide a framework to further investigate the genetic basis underlying the fitness and virulence of white rot fungi.© 2017 John Wiley & Sons Ltd.

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July 7, 2019

Genomics of parallel adaptation at two timescales in Drosophila.

Two interesting unanswered questions are the extent to which both the broad patterns and genetic details of adaptive divergence are repeatable across species, and the timescales over which parallel adaptation may be observed. Drosophila melanogaster is a key model system for population and evolutionary genomics. Findings from genetics and genomics suggest that recent adaptation to latitudinal environmental variation (on the timescale of hundreds or thousands of years) associated with Out-of-Africa colonization plays an important role in maintaining biological variation in the species. Additionally, studies of interspecific differences between D. melanogaster and its sister species D. simulans have revealed that a substantial proportion of proteins and amino acid residues exhibit adaptive divergence on a roughly few million years long timescale. Here we use population genomic approaches to attack the problem of parallelism between D. melanogaster and a highly diverged conger, D. hydei, on two timescales. D. hydei, a member of the repleta group of Drosophila, is similar to D. melanogaster, in that it too appears to be a recently cosmopolitan species and recent colonizer of high latitude environments. We observed parallelism both for genes exhibiting latitudinal allele frequency differentiation within species and for genes exhibiting recurrent adaptive protein divergence between species. Greater parallelism was observed for long-term adaptive protein evolution and this parallelism includes not only the specific genes/proteins that exhibit adaptive evolution, but extends even to the magnitudes of the selective effects on interspecific protein differences. Thus, despite the roughly 50 million years of time separating D. melanogaster and D. hydei, and despite their considerably divergent biology, they exhibit substantial parallelism, suggesting the existence of a fundamental predictability of adaptive evolution in the genus.

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July 7, 2019

Genetic maps and whole genome sequences of radish

Radish, Raphanus sativus L., is a member of Brassicaceae, to which Arabidopsis thaliana, a model plant in plant biology, belongs, as do other Brassica species including important crops. However, genetic and genomic studies of radish have been behind those of Arabidopsis and Brassica. In this decade, much effort has been made to develop genetic resources for radish, e.g., DNA markers, genetic maps, and whole genome sequences. Studies using the obtained information have revealed the genome structure of radish in terms of ancestral karyotype and have also prompted the identification of genes for agronomically important traits in radish through a map-based cloning strategy and quantitative trait locus analysis. In this chapter, we review the evolving development of radish genetic map in the past 15 years and the current status of genome sequencing of radish. We also introduce the latest strategy for the construction of a high-density genetic map using next-generation sequencing technology and propose a prospective direction of genetics and genomics research in radish which would be helpful for researchers and breeders in their efforts to promote radish breeding programs efficiently.

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July 7, 2019

Complete genome sequence analysis of Enterobacter sp. SA187, a plant multi-stress tolerance promoting endophytic bacterium

Enterobacter sp. SA187 is an endophytic bacterium that has been isolated from root nodules of the indigenous desert plant Indigofera argentea. SA187 could survive in the rhizosphere as well as in association with different plant species, and was able to provide abiotic stress tolerance to Arabidopsis thaliana. The genome sequence of SA187 was obtained by using Pacific BioScience (PacBio) single-molecule sequencing technology, with average coverage of 275X. The genome of SA187 consists of one single 4,429,597 bp chromosome, with an average 56% GC content and 4,347 predicted protein coding DNA sequences (CDS), 153 ncRNA, 7 rRNA, and 84 tRNA. Functional analysis of the SA187 genome revealed a large number of genes involved in uptake and exchange of nutrients, chemotaxis, mobilization and plant colonization. A high number of genes were also found to be involved in survival, defense against oxidative stress and production of antimicrobial compounds and toxins. Moreover, different metabolic pathways were identified that potentially contribute to plant growth promotion. The information encoded in the genome of SA187 reveals the characteristics of a dualistic lifestyle of a bacterium that can adapt to different environments and promote the growth of plants. This information provides a better understanding of the mechanisms involved in plant-microbe interaction and could be further exploited to develop SA187 as a biological agent to improve agricultural practices in marginal and arid lands.

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July 7, 2019

Genomic analysis of Bacillus licheniformis CBA7126 isolated from a human fecal sample.

Bacillus licheniformis is a Gram-positive, endospore-forming, saprophytic organism that occurs in plant and soil (Veith et al., 2004). A taxonomical approach shows that it is closely related to Bacillus subtilis (Lapidus et al., 2002; Xu and Côte, 2003; Rey et al., 2004). Generally, most bacilli are predominantly aerobic; however, B. licheniformis is a facultative anaerobe compared to other bacilli in ecological niches (Alexander, 1977). The commercial utility of the extracellular products of B. licheniformis makes this microorganism an economically interesting species (Kovács et al., 2009). For example, B. licheniformis is used industrially for manufacturing biochemicals, enzymes, antibiotics, and aminopeptidase. Several proteases such as a-amylase, penicillinase, pentosanase, cycloglucosyltransferase, ß-mannanase, and certain pectinolytic enzymes are synthesized industrially using B. licheniformis (Rodríguez-Absi and Prescott, 1978; Rey et al., 2004). The proteases are used in the detergent industry and the amylases are utilized for starch hydrolysis, desizing of textiles, and sizing of paper (Erickson, 1976). In addition, certain strains are utilized to produce peptide antibiotics, specialty chemicals, and poly-?-glutamic acid (Nierman and Maglott, 1989; Rey et al., 2004).

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July 7, 2019

Contributions of Zea mays subspecies mexicana haplotypes to modern maize.

Maize was domesticated from lowland teosinte (Zea mays ssp. parviglumis), but the contribution of highland teosinte (Zea mays ssp. mexicana, hereafter mexicana) to modern maize is not clear. Here, two genomes for Mo17 (a modern maize inbred) and mexicana are assembled using a meta-assembly strategy after sequencing of 10 lines derived from a maize-teosinte cross. Comparative analyses reveal a high level of diversity between Mo17, B73, and mexicana, including three Mb-size structural rearrangements. The maize spontaneous mutation rate is estimated to be 2.17?×?10-8 ~3.87?×?10-8 per site per generation with a nonrandom distribution across the genome. A higher deleterious mutation rate is observed in the pericentromeric regions, and might be caused by differences in recombination frequency. Over 10% of the maize genome shows evidence of introgression from the mexicana genome, suggesting that mexicana contributed to maize adaptation and improvement. Our data offer a rich resource for constructing the pan-genome of Zea mays and genetic improvement of modern maize varieties.

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July 7, 2019

Genome sequence of the small brown planthopper, Laodelphax striatellus.

Laodelphax striatellus Fallén (Hemiptera: Delphacidae) is one of the most destructive rice pests. L. striatellus is different from 2 other rice planthoppers with a released genome sequence, Sogatella furcifera and Nilaparvata lugens, in many biological characteristics, such as host range, dispersal capacity, and vectoring plant viruses. Deciphering the genome of L. striatellus will further the understanding of the genetic basis of the biological differences among the 3 rice planthoppers.A total of 190 Gb of Illumina data and 32.4 Gb of Pacbio data were generated and used to assemble a high-quality L. striatellus genome sequence, which is 541 Mb in length and has a contig N50 of 118 Kb and a scaffold N50 of 1.08 Mb. Annotated repetitive elements account for 25.7% of the genome. A total of 17?736 protein-coding genes were annotated, capturing 97.6% and 98% of the BUSCO eukaryote and arthropoda genes, respectively. Compared with N. lugens and S. furcifera, L. striatellus has the smallest genome and the lowest gene number. Gene family expansion and transcriptomic analyses provided hints to the genomic basis of the differences in important traits such as host range, migratory habit, and plant virus transmission between L. striatellus and the other 2 planthoppers.We report a high-quality genome assembly of L. striatellus, which is an important genomic resource not only for the study of the biology of L. striatellus and its interactions with plant hosts and plant viruses, but also for comparison with other planthoppers.© The Authors 2017. Published by Oxford University Press.

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July 7, 2019

The genome sequence of Bipolaris cookei reveals mechanisms of pathogenesis underlying target leaf spot of sorghum.

Bipolaris cookei (=Bipolaris sorghicola) causes target leaf spot, one of the most prevalent foliar diseases of sorghum. Little is known about the molecular basis of pathogenesis in B. cookei, in large part due to a paucity of resources for molecular genetics, such as a reference genome. Here, a draft genome sequence of B. cookei was obtained and analyzed. A hybrid assembly strategy utilizing Illumina and Pacific Biosciences sequencing technologies produced a draft nuclear genome of 36.1?Mb, organized into 321 scaffolds with L50 of 31 and N50 of 378?kb, from which 11,189 genes were predicted. Additionally, a finished mitochondrial genome sequence of 135,790?bp was obtained, which contained 75 predicted genes. Comparative genomics revealed that B. cookei possessed substantially fewer carbohydrate-active enzymes and secreted proteins than closely related Bipolaris species. Novel genes involved in secondary metabolism, including genes implicated in ophiobolin biosynthesis, were identified. Among 37 B. cookei genes induced during sorghum infection, one encodes a putative effector with a limited taxonomic distribution among plant pathogenic fungi. The draft genome sequence of B. cookei provided novel insights into target leaf spot of sorghum and is an important resource for future investigation.

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July 7, 2019

Assembly of an early-matured japonica (Geng) rice genome, Suijing18, based on PacBio and Illumina sequencing.

The early-matured japonica (Geng) rice variety, Suijing18 (SJ18), carries multiple elite traits including durable blast resistance, good grain quality, and high yield. Using PacBio SMRT technology, we produced over 25?Gb of long-read sequencing raw data from SJ18 with a coverage of 62×. Using Illumina paired-end whole-genome shotgun sequencing technology, we generated 59?Gb of short-read sequencing data from SJ18 (23.6?Gb from a 200?bp library with a coverage of 59× and 35.4?Gb from an 800?bp library with a coverage of 88×). With these data, we assembled a single SJ18 genome and then generated a set of annotation data. These data sets can be used to test new programs for variation deep mining, and will provide new insights into the genome structure, function, and evolution of SJ18, and will provide essential support for biological research in general.

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July 7, 2019

Chromosome level assembly and secondary metabolite potential of the parasitic fungus Cordyceps militaris.

Cordyceps militaris is an insect pathogenic fungus that is prized for its use in traditional medicine. This and other entomopathogenic fungi are understudied sources for the discovery of new bioactive molecules. In this study, PacBio SMRT long read sequencing technology was used to sequence the genome of C. militaris with a focus on the genetic potential for secondary metabolite production in the genome assembly of this fungus.This is first chromosome level assembly of a species in the Cordyceps genera. In this seven chromosome assembly of 33.6 Mba there were 9371 genes identified. Cordyceps militaris was determined to have the MAT 1-1-1 and MAT 1-1-2 mating type genes. Secondary metabolite analysis revealed the potential for at least 36 distinct metabolites from a variety of classes. Three of these gene clusters had homology with clusters producing desmethylbassianin, equisetin and emericellamide that had been studied in other fungi.Our assembly and analysis has revealed that C. militaris has a wealth of gene clusters for secondary metabolite production distributed among seven chromosomes. The identification of these gene clusters will facilitate the future study and identification of the secondary metabolites produced by this entomopathogenic fungus.

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July 7, 2019

Draft genome of Paraburkholderia caballeronis TNe-841T, a free-living, nitrogen-fixing, tomato plant-associated bacterium.

10.1601/nm.26956 caballeronis is a plant-associated bacterium. Strain TNe-841T was isolated from the rhizosphere of tomato (Solanum lycopersicum L. var. lycopersicum) growing in Nepantla Mexico State. Initially this bacterium was found to effectively nodulate Phaseolus vulgaris L. However, from an analysis of the genome of strain TNe-841T and from repeat inoculation experiments, we found that this strain did not nodulate bean and also lacked nodulation genes, suggesting that the genes were lost. The genome consists of 7,115,141 bp with a G?+?C content of 67.01%. The sequence includes 6251 protein-coding genes and 87 RNA genes.

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July 7, 2019

Streptococcal toxic shock syndrome caused by the dissemination of an invasive emm3/ST15 strain of Streptococcus pyogenes.

Streptococcus pyogenes (group A Streptococcus [GAS]) is a major human pathogen that causes a wide spectrum of clinical manifestations. Although invasive GAS (iGAS) infections are relatively uncommon, emm3/ST15 GAS is a highly virulent, invasive, and pathogenic strain. Global molecular epidemiology analysis has suggested that the frequency of emm3 GAS has been recently increasing.A 14-year-old patient was diagnosed with streptococcal toxic shock syndrome and severe pneumonia, impaired renal function, and rhabdomyolysis. GAS was isolated from a culture of endotracheal aspirates and designated as KS030. Comparative genome analysis suggested that KS030 is classified as emm3 (emm-type) and ST15 (multilocus sequencing typing [MLST]), which is similar to iGAS isolates identified in the UK (2013) and Switzerland (2015).We conclude that the global dissemination of emm3/ST15 GAS strain has the potential to cause invasive disease.

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July 7, 2019

Draft sequencing of the heterozygous diploid genome of Satsuma (Citrus unshiu Marc.) using a hybrid assembly approach.

Satsuma (Citrus unshiu Marc.) is one of the most abundantly produced mandarin varieties of citrus, known for its seedless fruit production and as a breeding parent of citrus. De novo assembly of the heterozygous diploid genome of Satsuma (“Miyagawa Wase”) was conducted by a hybrid assembly approach using short-read sequences, three mate-pair libraries, and a long-read sequence of PacBio by the PLATANUS assembler. The assembled sequence, with a total size of 359.7 Mb at the N50 length of 386,404 bp, consisted of 20,876 scaffolds. Pseudomolecules of Satsuma constructed by aligning the scaffolds to three genetic maps showed genome-wide synteny to the genomes of Clementine, pummelo, and sweet orange. Gene prediction by modeling with MAKER-P proposed 29,024 genes and 37,970 mRNA; additionally, gene prediction analysis found candidates for novel genes in several biosynthesis pathways for gibberellin and violaxanthin catabolism. BUSCO scores for the assembled scaffold and predicted transcripts, and another analysis by BAC end sequence mapping indicated the assembled genome consistency was close to those of the haploid Clementine, pummel, and sweet orange genomes. The number of repeat elements and long terminal repeat retrotransposon were comparable to those of the seven citrus genomes; this suggested no significant failure in the assembly at the repeat region. A resequencing application using the assembled sequence confirmed that both kunenbo-A and Satsuma are offsprings of Kishu, and Satsuma is a back-crossed offspring of Kishu. These results illustrated the performance of the hybrid assembly approach and its ability to construct an accurate heterozygous diploid genome.

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