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Asset Tag: Whole Genome Sequencing,

April 21, 2020

The complete mitochondrial genome of the New Zealand parasitic roundworm Haemonchus contortus (Trichostrongyloidea: Haemonchidae) field strain NZ_Hco_NP

The complete mitochondrial genome of the New Zealand parasitic nematode Haemonchus contortus field strain NZ_Hco_NP was sequenced and annotated. The 14,001bp-long mitogenome contains 12 protein-coding genes (atp8 gene missing), two ribosomal RNAs, 22 transfer RNAs, and a 583bp non- coding region. Phylogenetic analysis showed that H. contortus NZ_Hco_NP forms a monophyletic clus- ter with the remaining three Haemonchidae species, and further illustrates the high levels of diversity and gene flow among Trichostrongylidae.

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April 21, 2020

Genome sequencing and CRISPR/Cas9 gene editing of an early flowering Mini-Citrus (Fortunella hindsii).

Hongkong kumquat (Fortunella hindsii) is a wild citrus species characterized by dwarf plant height and early flowering. Here, we identified the monoembryonic F. hindsii (designated as ‘Mini-Citrus’) for the first time and constructed its selfing lines. This germplasm constitutes an ideal model for the genetic and functional genomics studies of citrus, which have been severely hindered by the long juvenility and inherent apomixes of citrus. F. hindsii showed a very short juvenile period (~8 months) and stable monoembryonic phenotype under cultivation. We report the first de novo assembled 373.6 Mb genome sequences (Contig-N50 2.2 Mb and Scaffold-N50 5.2 Mb) for F. hindsii. In total, 32 257 protein-coding genes were annotated, 96.9% of which had homologues in other eight Citrinae species. The phylogenomic analysis revealed a close relationship of F. hindsii with cultivated citrus varieties, especially with mandarin. Furthermore, the CRISPR/Cas9 system was demonstrated to be an efficient strategy to generate target mutagenesis on F. hindsii. The modifications of target genes in the CRISPR-modified F. hindsii were predominantly 1-bp insertions or small deletions. This genetic transformation system based on F. hindsii could shorten the whole process from explant to T1 mutant to about 15 months. Overall, due to its short juvenility, monoembryony, close genetic background to cultivated citrus and applicability of CRISPR, F. hindsii shows unprecedented potentials to be used as a model species for citrus research. © 2019 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

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April 21, 2020

A chromosome-scale genome assembly reveals a highly dynamic effector repertoire of wheat powdery mildew.

Blumeria graminis f. sp. tritici (B.g. tritici) is the causal agent of the wheat powdery mildew disease. The highly fragmented B.g. tritici genome available so far has prevented a systematic analysis of effector genes that are known to be involved in host adaptation. To study the diversity and evolution of effector genes we produced a chromosome-scale assembly of the B.g. tritici genome. The genome assembly and annotation was achieved by combining long-read sequencing with high-density genetic mapping, bacterial artificial chromosome fingerprinting and transcriptomics. We found that the 166.6 Mb B.g. tritici genome encodes 844 candidate effector genes, over 40% more than previously reported. Candidate effector genes have characteristic local genomic organization such as gene clustering and enrichment for recombination-active regions and certain transposable element families. A large group of 412 candidate effector genes shows high plasticity in terms of copy number variation in a global set of 36 isolates and of transcription levels. Our data suggest that copy number variation and transcriptional flexibility are the main drivers for adaptation in B.g. tritici. The high repeat content may play a role in providing a genomic environment that allows rapid evolution of effector genes with selection as the driving force. © 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.

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April 21, 2020

Medaka Population Genome Structure and Demographic History Described via Genotyping-by-Sequencing.

Medaka is a model organism in medicine, genetics, developmental biology and population genetics. Lab stocks composed of more than 100 local wild populations are available for research in these fields. Thus, medaka represents a potentially excellent bioresource for screening disease-risk- and adaptation-related genes in genome-wide association studies. Although the genetic population structure should be known before performing such an analysis, a comprehensive study on the genome-wide diversity of wild medaka populations has not been performed. Here, we performed genotyping-by-sequencing (GBS) for 81 and 12 medakas captured from a bioresource and the wild, respectively. Based on the GBS data, we evaluated the genetic population structure and estimated the demographic parameters using an approximate Bayesian computation (ABC) framework. The genome-wide data confirmed that there were substantial differences between local populations and supported our previously proposed hypothesis on medaka dispersal based on mitochondrial genome (mtDNA) data. A new finding was that a local group that was thought to be a hybrid between the northern and the southern Japanese groups was actually an origin of the northern Japanese group. Thus, this paper presents the first population-genomic study of medaka and reveals its population structure and history based on chromosomal genetic diversity.Copyright © 2019 by the Genetics Society of America.

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April 21, 2020

Characterization of a novel, type II staphylococcal cassette chromosome mec element from an endemic oxacillin-resistant Staphylococcus lugdunensis clone in a hospital setting.

Staphylococcus lugdunensis is a significant pathogen that causes community-acquired and nosocomial infections. The high prevalence of oxacillin-resistant S. lugdunensis (ORSL) is of major concern. Resistance to ß-lactams is caused by acquisition of the staphylococcal cassette chromosome mec (SCCmec) element. The cassette is highly diverse, both structurally and genetically, among CoNS. Isolates carrying SCCmec II-ST6 are the major persistent clones in hospitals.To investigate the structure and evolutionary origin of a novel type II SCCmec element in an endemic ST6 S. lugdunensis clone.The structure of the SCCmec II element carried by ST6 strain CGMH-SL118 was determined by WGS and compared with those reported previously.A novel 39 kb SCCmec element, SCCmecCGMH-SL118, with a unique mosaic structure comprising 41 ORFs integrated into the 3′ end of the rlmH gene, was observed. Some regions of SCCmecCGMH-SL118 were homologous to SCCmec IIa of the prototype MRSA strain N315. The structure of SCCmecCGMH-SL118 was similar to that of SCCmec IIb of the MRSA strain, JCSC3063, mainly lacking the aminoglycoside resistance determinant pUB110 in the J3 region but containing the insertion sequence IS256 in the J2 region. Notably, SCCmecCGMH-SL118 deletions in the J1 region compared with SCCmec types IIa and IIb, and a high homology to SCCmec elements of Staphylococcus aureus JCSC4610 and Staphylococcus haemolyticus strain 621 were found.The genetic diversity of the type II SCCmec element in ORSL suggests that CoNS is a potential reservoir for interspecies transfer of SCCmec to S. aureus in hospitals. © The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.

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April 21, 2020

Multiple modes of convergent adaptation in the spread of glyphosate-resistant Amaranthus tuberculatus.

The selection pressure exerted by herbicides has led to the repeated evolution of herbicide resistance in weeds. The evolution of herbicide resistance on contemporary timescales in turn provides an outstanding opportunity to investigate key questions about the genetics of adaptation, in particular the relative importance of adaptation from new mutations, standing genetic variation, or geographic spread of adaptive alleles through gene flow. Glyphosate-resistant Amaranthus tuberculatus poses one of the most significant threats to crop yields in the Midwestern United States, with both agricultural populations and herbicide resistance only recently emerging in Canada. To understand the evolutionary mechanisms driving the spread of resistance, we sequenced and assembled the A. tuberculatus genome and investigated the origins and population genomics of 163 resequenced glyphosate-resistant and susceptible individuals from Canada and the United States. In Canada, we discovered multiple modes of convergent evolution: in one locality, resistance appears to have evolved through introductions of preadapted US genotypes, while in another, there is evidence for the independent evolution of resistance on genomic backgrounds that are historically nonagricultural. Moreover, resistance on these local, nonagricultural backgrounds appears to have occurred predominantly through the partial sweep of a single haplotype. In contrast, resistant haplotypes arising from the Midwestern United States show multiple amplification haplotypes segregating both between and within populations. Therefore, while the remarkable species-wide diversity of A. tuberculatus has facilitated geographic parallel adaptation of glyphosate resistance, more recently established agricultural populations are limited to adaptation in a more mutation-limited framework.Copyright © 2019 the Author(s). Published by PNAS.

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April 21, 2020

The first report of a novel IncHI1B blaSIM-1-carrying megaplasmid pSIM-1-BJ01 from a clinical Klebsiella pneumoniae isolate.

Background: A rare member of metallo-ß-lactamases genes, blaSIM-1, carried by a 316-kb plasmid designated pSIM-1-BJ01 was isolated from a clinical cephalosporins- and carbapenem-resistant Klebsiellapneumoniae 13624. This is the first sequence report of a transferable blaSIM-1-carrying conjugative plasmid isolated from K. pneumoniae. Purpose: The sequence analysis of pSIM-1-BJ01 will help us to identify genes responsible for conjugation, plasmid maintenance and drug resistance, to understand the evolution and control the dissemination of resistance plasmids. Patients and methods:K. pneumoniae 13624 was isolated from the urine specimen of a patient. Bacterial genomic DNA was sequenced with PacBio RSII platform. Results: Most of the pSIM-1-BJ01 backbone matches that of pRJA166a, which was isolated from a clinical hypervirulent K. pneumoniae ST23 strain at Shanghai, China, recently. The highly homologous backbones between the two plasmids imply the close relationship of evolution. Two different multidrug-resistant regions both carrying the class 1 integrons with different resistance genes have been assembled into the pSIM-1-BJ01. Besides, the other two resistance plasmids, pKP13624-1 carrying blaTEM-1 and blaCTX-M-15 and pKP13624-2 carrying blaCTX-M-14 and blaLAP-2 were also identified. Conclusion: The emergence of the blaSIM-1-carrying IncHI1B pSIM-1-BJ01 suggests the spread of blaSIM among Enterobacteriaceae is possible. We should pay more attention to supervise and control the dissemination of hypervirulent carbapenem-resistant K. pneumonia in public hospitals.

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April 21, 2020

Genome characterization of an extensively drug-resistant Streptococcus pneumoniae serotype 11A strain.

In this study, the whole genome sequences of two Streptococcus pneumoniae clinical isolates from South Korea were determined and compared. They were found to be the same serotype (11?A) and multilocus sequence typing analysis showed that they are single-locus variants (SLVs; ST8279 and ST166) of each other, differing at one allele (aroE). However, the ST8279 strain is extensively drug-resistant (XDR) whereas the ST166 strain is not. The genome of the XDR strain is very similar in structure to that of two previously reported genomes, AP200 (11?A:ST62) and 70585 (5:ST5803); however, some regions were inverted and there were some exogenous regions in the ST8279 strain. It was found that 6,502 single nucleotide polymorphisms are dispersed across the genome between the two serotype 11?A ST8279 and ST166 strains. Many of them are located in genes associated with antibiotic resistance. In addition, many amino acid differences were also identified in genes involved in DNA repair (mutL, uvrA and uvrC) and recombination (recU, recR and recA). On the basis of these results, it was inferred that the XDR strain did not evolve from its SLV via a single recombination event involving a large portion of the genome including the aroE gene. Rather, the strain likely evolved through many point mutations and recombination events involving small portions of the genome. © 2019 The Societies and John Wiley & Sons Australia, Ltd.

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April 21, 2020

The major histocompatibility complex of Old World camelids: Class I and class I-related genes.

The genomic structure of the Major Histocompatibility Complex (MHC) region and variation in selected MHC class I related genes in Old World camels, Camelus bactrianus and Camelus dromedaries were studied. The overall genomic organization of the camel MHC region follows a general pattern observed in other mammalian species and individual MHC loci appear to be well conserved. Selected MHC class I genes B-67 and BL3-7 exhibited unexpectedly low variability, even when compared to other camel MHC class I related genes MR1 and MICA. Interspecific SNP and allele sharing are relatively common, and frequencies of heterozygotes are usually low. Such a low variation in a genomic region generally considered as one of the most polymorphic in vertebrate genomes is unusual. Evolutionary relationships between MHC class I related genes and their counterparts from other species seem to be rather complex. Often, they do not follow the general evolutionary history of the species concerned. Close evolutionary relationships of individual MHC class I loci between camels, humans and dogs were observed. Based on the results of this study and on our data on MHC class II genes, the extent and the pattern of polymorphism of the MHC region of Old World camelids differed from most mammalian groups studied so far. Camels thus seem to be an important model for our understanding of the role of genetic diversity in immune functions, especially in the context of unique features of their immunoglobulin and T-cell receptor genes. © 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

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April 21, 2020

Mate Selection in Self-Compatible Wild Tobacco Results from Coordinated Variation in Homologous Self-Incompatibility Genes.

In flowering plants, intraspecific mate preference is frequently related to mating systems: the rejection of self pollen in self-incompatible (SI) plants that prevents inbreeding is one of the best described examples. However, in other mating systems, more nuanced patterns of pollen rejection occur. In the self-compatible (SC) Nicotiana attenuata, in which SI is not found and all crosses are compatible, certain pollen genotypes are consistently selected in mixed pollinations. However, the molecular mechanisms of this polyandrous mate selection remain unknown. Style-expressed NaS-like-RNases and pollen-expressed NaSLF-like genes, homologous to SI factors in Solanaceae, were identified and examined for a role in N. attenuata’s mate selection. A comparison of two NaS-like-RNases and six NaSLF-like genes among 26 natural accessions revealed specific combinations of co-expression and direct protein-protein interactions. To evaluate their role in mate selection, we silenced the expression of specific NaS-like-RNases and NaSLF-like proteins and conducted diagnostic binary mixed pollinations and mixed pollinations with 14 different non-self pollen donors. Styles expressing particular combinations of NaS-like-RNases selected mates from plants with corresponding NaS-like-RNase expression patterns, while styles lacking NaS-like-RNase expression were non-selective in their fertilizations, which reflected the genotype ratios of pollen mixtures deposited on the stigmas. DNA methylation could account for some of the observed variation in stylar NaS-like-RNase patterns. We conclude that the S-RNase-SLF recognition mechanism plays a central role in polyandrous mate selection in this self-compatible species. These results suggest that after the SI-SC transition, natural variation of SI homologous genes was repurposed to mediate intraspecific mate selection. Copyright © 2019 Elsevier Ltd. All rights reserved.

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April 21, 2020

Genetic, structural, and functional diversity of low and high-affinity siderophores in strains of nitrogen fixing Azotobacter chroococcum.

To increase iron (Fe) bioavailability in surface soils, microbes secrete siderophores, chelators with widely varying Fe affinities. Strains of the soil bacterium Azotobacter chroococcum (AC), plant-growth promoting rhizobacteria used as agricultural inoculants, require high Fe concentrations for aerobic respiration and nitrogen fixation. Recently, A. chroococcum str. NCIMB 8003 was shown to synthesize three siderophore classes: (1) vibrioferrin, a low-affinity a-hydroxy carboxylate (pFe = 18.4), (2) amphibactins, high-affinity tris-hydroxamates, and (3) crochelin A, a high-affinity siderophore with mixed Fe-chelating groups (pFe = 23.9). The relevance and specific functions of these siderophores in AC strains remain unclear. We analyzed the genome and siderophores of a second AC strain, A. chroococcum str. B3, and found that it also produces vibrioferrin and amphibactins, but not crochelin A. Genome comparisons indicate that vibrioferrin production is a vertically inherited, conserved strategy for Fe uptake in A. chroococcum and other species of Azotobacter. Amphibactin and crochelin biosynthesis reflects a more complex evolutionary history, shaped by vertical gene transfer, gene gain and loss through recombination at a genomic hotspot. We found conserved patterns of low vs. high-affinity siderophore production across strains: the low-affinity vibrioferrin was produced by mildly Fe limited cultures. As cells became more severely Fe starved, vibrioferrin production decreased in favor of high-affinity amphibactins (str. B3, NCIMB 8003) and crochelin A (str. NCIMB 8003). Our results show the evolution of low and high-affinity siderophore families and conserved patterns for their production in response to Fe bioavailability in a common soil diazotroph.

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April 21, 2020

Development of a Molecular Marker Linked to the A4 Locus and the Structure of HD Genes in Pleurotus eryngii

Allelic differences in A and B mating-type loci are a prerequisite for the progression of mating in the genus Pleurotus eryngii; thus, the crossing is hampered by this biological barrier in inbreeding. Molecular markers linked to mating types of P. eryngii KNR2312 were investigated with randomly amplified polymorphic DNA to enhance crossing efficiency. An A4-linked sequence was identified and used to find the adjacent genomic region with the entire motif of the A locus from a contig sequenced by PacBio. The sequence-characterized amplified region marker 7-2299 distinguished A4 mating-type monokaryons from KNR2312 and other strains. A BLAST search of flanked sequences revealed that the A4 locus had a general feature consisting of the putative HD1 and HD2 genes. Both putative HD transcription factors contain a homeodomain sequence and a nuclear localization sequence; however, valid dimerization motifs were found only in the HD1 protein. The ACAAT motif, which was reported to have relevance to sex determination, was found in the intergenic region. The SCAR marker could be applicable in the classification of mating types in the P. eryngii breeding program, and the A4 locus could be the basis for a multi-allele detection marker.

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April 21, 2020

The complete mitochondrial genome of the tartar Sand Boa Eryx tataricus

Eryx is a genus of snakes belonging to the family Boidae. In this study, the mitochondrial genome sequence of Eryx tataricus was generated using a PacBio RSII DNA sequencer employing the single mol- ecule, real-time sequencing technology. A maximum-likelihood (ML) phylogenetic tree of 26 snakes was re-constructed based on the 13 protein-coding genes for convincing the mitochondrial DNA sequences.

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April 21, 2020

Ancestral Admixture Is the Main Determinant of Global Biodiversity in Fission Yeast.

Mutation and recombination are key evolutionary processes governing phenotypic variation and reproductive isolation. We here demonstrate that biodiversity within all globally known strains of Schizosaccharomyces pombe arose through admixture between two divergent ancestral lineages. Initial hybridization was inferred to have occurred ~20-60 sexual outcrossing generations ago consistent with recent, human-induced migration at the onset of intensified transcontinental trade. Species-wide heritable phenotypic variation was explained near-exclusively by strain-specific arrangements of alternating ancestry components with evidence for transgressive segregation. Reproductive compatibility between strains was likewise predicted by the degree of shared ancestry. To assess the genetic determinants of ancestry block distribution across the genome, we characterized the type, frequency, and position of structural genomic variation using nanopore and single-molecule real-time sequencing. Despite being associated with double-strand break initiation points, over 800 segregating structural variants exerted overall little influence on the introgression landscape or on reproductive compatibility between strains. In contrast, we found strong ancestry disequilibrium consistent with negative epistatic selection shaping genomic ancestry combinations during the course of hybridization. This study provides a detailed, experimentally tractable example that genomes of natural populations are mosaics reflecting different evolutionary histories. Exploiting genome-wide heterogeneity in the history of ancestral recombination and lineage-specific mutations sheds new light on the population history of S. pombe and highlights the importance of hybridization as a creative force in generating biodiversity. © The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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April 21, 2020

The Genome Sequence of the Anthelmintic-Susceptible New Zealand Haemonchus contortus.

Internal parasitic nematodes are a global animal health issue causing drastic losses in livestock. Here, we report a H. contortus representative draft genome to serve as a genetic resource to the scientific community and support future experimental research of molecular mechanisms in related parasites. A de novo hybrid assembly was generated from PCR-free whole genome sequence data, resulting in a chromosome-level assembly that is 465 Mb in size encoding 22,341 genes. The genome sequence presented here is consistent with the genome architecture of the existing Haemonchus species and is a valuable resource for future studies regarding population genetic structures of parasitic nematodes. Additionally, comparative pan-genomics with other species of economically important parasitic nematodes have revealed highly open genomes and strong collinearities within the phylum Nematoda. © The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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