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April 21, 2020

Genome Sequence of Bacillus Velezensis W1, A Strain with Strong Acaricidal Activity against Two-Spotted Spider Mite (Tetranychus Urticae)

Bacillus velezensis W1, isolated from two-spotted spider mites that had died naturally, is a patented strain with strong capability to cause mortality of the phytophagous mite Tetranychus urticae. The whole genome of W1 was completely sequenced with a combination of an Illumina Miseq platform (400-bp paired-end) with 2 × 250 bases and a Pacific Biosciences (PaBio) RS II Single Molecule Real Time (SMRT) sequencing platform using a 20 kb SMRTbellTM template library. Here, we report the complete genome sequence of B. velezensis W1, including one circular chromosome of 4,237,431 bp encoding 4,352 genes with GC content of 45.84%, providing insights into the genomic basis of its acaricidal activity and facilitating its application in red spider mite biocontrol.

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April 21, 2020

Complete chloroplast genome sequence of Amomum villosum

The first complete chloroplast genome of Amomum villosum (Zingiberaceae) was reported in this study. The A. villosum genome was 163,608bp in length, and comprised a pair of inverted repeat (IR) regions of 29,820bp each, a large single-copy (LSC) region of 88,680bp, and a small single-copy (SSC) region of 15,288bp. It encoded 141 genes, including 87 protein-coding genes (79 PCG species), 46 tRNA genes (28 tRNA species), and 8 rRNA genes (4 rRNA species). The overall AT content was 63.92%. Phylogenetic analysis showed that A. villosum was closely related to two species Amomum kravanh and Amomum compactum within the genus Amomum in family Zingiberaceae.

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April 21, 2020

Long-read sequencing identified intronic repeat expansions in SAMD12 from Chinese pedigrees affected with familial cortical myoclonic tremor with epilepsy.

The locus for familial cortical myoclonic tremor with epilepsy (FCMTE) has long been mapped to 8q24 in linkage studies, but the causative mutations remain unclear. Recently, expansions of intronic TTTCA and TTTTA repeat motifs within SAMD12 were found to be involved in the pathogenesis of FCMTE in Japanese pedigrees. We aim to identify the causative mutations of FCMTE in Chinese pedigrees.We performed genetic linkage analysis by microsatellite markers in a five-generation Chinese pedigree with 55 members. We also used array-comparative genomic hybridisation (CGH) and next-generation sequencing (NGS) technologies (whole-exome sequencing, capture region deep sequencing and whole-genome sequencing) to identify the causative mutations in the disease locus. Recently, we used low-coverage (~10×) long-read genome sequencing (LRS) on the PacBio Sequel and Oxford Nanopore platforms to identify the causative mutations, and used repeat-primed PCR for validation of the repeat expansions.Linkage analysis mapped the disease locus to 8q23.3-24.23. Array-CGH and NGS failed to identify causative mutations in this locus. LRS identified the intronic TTTCA and TTTTA repeat expansions in SAMD12 as the causative mutations, thus corroborating the recently published results in Japanese pedigrees.We identified the pentanucleotide repeat expansion in SAMD12 as the causative mutation in Chinese FCMTE pedigrees. Our study also suggested that LRS is an effective tool for molecular diagnosis of genetic disorders, especially for neurological diseases that cannot be positively diagnosed by conventional clinical microarray and NGS technologies. © Author(s) (or their employer(s)) 2019. No commercial re-use. See rights and permissions. Published by BMJ.

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April 21, 2020

Complete chloroplast genome sequence of Carthamus tinctorius L. from PacBio Sequel Platform

Carthamus tinctorius L, also known as safflower, is an important oil crop planted worldwide. The com- plete chloroplast (cp) genome was reported in this study using the PacBio Sequel Platform. The cp genome with a total size of 152,963bp consisted of two inverted repeats (25,128bp) separated by a large single-copy region (84,124bp) and a small single-copy region (18,583bp). Further annotation revealed the cp genome contains 112 genes, including 79 protein-coding genes, 29 tRNA genes, and 4 rRNA genes. The information of the cp genome will be useful for investigation of evolution and molecular breeding of safflower in the future.

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April 21, 2020

Carbohydrate catabolic capability of a Flavobacteriia bacterium isolated from hadal water.

Flavobacteriia are abundant in many marine environments including hadal waters, as demonstrated recently. However, it is unclear how this flavobacterial population adapts to hadal conditions. In this study, extensive comparative genomic analyses were performed for the flavobacterial strain Euzebyella marina RN62 isolated from the Mariana Trench hadal water in low abundance. The complete genome of RN62 possessed a considerable number of carbohydrate-active enzymes with a different composition. There was a predominance of GH family 13 proteins compared to closely related relatives, suggesting that RN62 has preserved a certain capacity for carbohydrate utilization and that the hadal ocean may hold an organic matter reservoir distinct from the surface ocean. Additionally, RN62 possessed potential intracellular cycling of the glycogen/starch pathway, which may serve as a strategy for carbon storage and consumption in response to nutrient pulse and starvation. Moreover, the discovery of higher glycoside hydrolase dissimilarities among Flavobacteriia, compared to peptidases and transporters, suggested variation in polysaccharide utilization related traits as an important ecophysiological factor in response to environmental alterations, such as decreased labile organic carbon in hadal waters. The presence of abundant toxin exporting, transcription and signal transduction related genes in RN62 may further help to survive in hadal conditions, including high pressure/low temperature.Copyright © 2019 Elsevier GmbH. All rights reserved.

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April 21, 2020

A new variant of mcr-1 identified from an extended-spectrum ß lactamase-producing Escherichia coli.

Plasmid-mediated colistin resistance gene, mcr-1, has been widely reported almost all over the world. The product of the gene, MCR-1, is one of the members of the phosphoethanolamine transferase enzyme family, which can add phosphoethanolamine to lipid A, thus reducing affinity to polymyxins. Isolates carrying mcr-1 gene are often multidrug resistant (MDR), including co-production of MCR-1 and extended spectrum B lactamases (ESBLs) or carbapenemases, resulting in great clinical concerns.

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April 21, 2020

Genetic characterization and potential molecular dissemination mechanism of tet(31) gene in Aeromonas caviae from an oxytetracycline wastewater treatment system.

Recently, the rarely reported tet(31) tetracycline resistance determinant was commonly found in Aeromonas salmonicida, Gallibacterium anatis, and Oblitimonas alkaliphila isolated from farming animals and related environment. However, its distribution in other bacteria and potential molecular dissemination mechanism in environment are still unknown. The purpose of this study was to investigate the potential mechanism underlying dissemination of tet(31) by analysing the tet(31)-carrying fragments in A. caviae strains isolated from an aerobic biofilm reactor treating oxytetracycline bearing wastewater. Twenty-three A. caviae strains were screened for the tet(31) gene by polymerase chain reaction (PCR). Three strains (two harbouring tet(31), one not) were subjected to whole genome sequencing using the PacBio RSII platform. Seventeen A. caviae strains carried the tet(31) gene and exhibited high resistance levels to oxytetracycline with minimum inhibitory concentrations (MICs) ranging from 256 to 512?mg/L. tet(31) was comprised of the transposon Tn6432 on the chromosome of A. caviae, and Tn6432 was also found in 15 additional tet(31)-positive A. caviae isolates by PCR. More important, Tn6432 was located on an integrative conjugative element (ICE)-like element, which could mediate the dissemination of the tet(31)-carrying transposon Tn6432 between bacteria. Comparative analysis demonstrated that Tn6432 homologs with the structure ISCR2-?phzF-tetR(31)-tet(31)-?glmM-sul2 were also carried by A. salmonicida, G. anatis, and O. alkaliphila, suggesting that this transposon can be transferred between species and even genera. This work provides the first report on the identification of the tet(31) gene in A. caviae, and will be helpful in exploring the dissemination mechanisms of tet(31) in water environment.Copyright © 2018. Published by Elsevier B.V.

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April 21, 2020

Iron-associated protein interaction networks reveal the key functional modules related to survival and virulence of Pasteurella multocida.

Pasteurella multocida causes respiratory infectious diseases in a multitude of birds and mammals. A number of virulence-associated genes were reported across different strains of P. multocida, including those involved in the iron transport and metabolism. Comparative iron-associated genes of P. multocida among different animal hosts towards their interaction networks have not been fully revealed. Therefore, this study aimed to identify the iron-associated genes from core- and pan-genomes of fourteen P. multocida strains and to construct iron-associated protein interaction networks using genome-scale network analysis which might be associated with the virulence. Results showed that these fourteen strains had 1587 genes in the core-genome and 3400 genes constituting their pan-genome. Out of these, 2651 genes associated with iron transport and metabolism were selected to construct the protein interaction networks and 361 genes were incorporated into the iron-associated protein interaction network (iPIN) consisting of nine different iron-associated functional modules. After comparing with the virulence factor database (VFDB), 21 virulence-associated proteins were determined and 11 of these belonged to the heme biosynthesis module. From this study, the core heme biosynthesis module and the core outer membrane hemoglobin receptor HgbA were proposed as candidate targets to design novel antibiotics and vaccines for preventing pasteurellosis across the serotypes or animal hosts for enhanced precision agriculture to ensure sustainability in food security. Copyright © 2018. Published by Elsevier Ltd.

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April 21, 2020

Integration of two pKPX-2-derived antibiotic resistance islands in the genome of an ESBL-producing Klebsiella pneumoniae ST3483 from Lebanon.

Contamination of fresh water with clinically important Gram-negative bacteria in Lebanon is being investigated in-depth, especially with evidence of dissemination into clinical settings. This study aimed to report the draft genome sequence of a Klebsiella pneumoniae strain with an integrated plasmid segment harbouring two antibiotic resistance islands (ARI). It is believed that this is the first report of plasmid antibiotic resistance islands integration in the genome of K. pneumoniae.Whole genome sequencing of the isolate was performed using Sequel platform. The genome was assembled using HGAP4. Analysis was conducted by uploading the sequence to the online databases from the Center for Genomic Epidemiology.The strain had a newly assigned ST 3483 with a genome size of 5385844 bp. The investigation of the antibiotic resistance islands suggested integration of two DNA segments from a previously identified IncFIA plasmid. The results revealed that the integration could have been accomplished either as a single-step integration event, with the two segments being integrated as a whole transposon mediated by the flanking IS26, or through two separate integration events involving the two segments, but independently.The sequenced genome revealed interesting aspects related to antibiotic resistance dissemination. The ARI are more stable in the genome and the chance of losing it is less probable, with the possibility of the described transposon to re-integrate in other plasmids, facilitating the dissemination of such resistance determinants.Copyright © 2019 International Society for Antimicrobial Chemotherapy. Published by Elsevier Ltd. All rights reserved.

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April 21, 2020

Genome Organization and Adaptive Potential of Archetypal Organophosphate Degrading Sphingobium fuliginis ATCC 27551.

Sphingobium fuliginis ATCC 27551, previously classified as Flavobacterium sp. ATCC 27551, degrades neurotoxic organophosphate insecticides and nerve agents through the activity of a membrane-associated organophosphate hydrolase. This study was designed to determine the complete genome sequence of S. fuliginis ATCC 27551 to unravel its degradative potential and adaptability to harsh environments. The 5,414,624?bp genome with a GC content of 64.4% is distributed between two chromosomes and four plasmids and encodes 5,557 proteins. Of the four plasmids, designated as pSF1, pSF2, pSF3, and pSF4, only two (pSF1 and pSF2) are self-transmissible and contained the complete genetic repertoire for a T4SS. The other two plasmids (pSF3 and pSF4) are mobilizable and both showed the presence of an oriT and relaxase-encoding sequences. The sequence of plasmid pSF3 coincided with the previously determined sequence of pPDL2 and included an opd gene encoding organophosphate hydrolase as a part of the mobile element. About 15,455 orthologous clusters were identified from among the cumulatively annotated genes of 49 Sphingobium species. Phylogenetic analysis done using the core genome consisting of 802 orthologous clusters revealed a close relationship between S. fuliginis ATCC 27551 and bacteria capable of degradation of polyaromatic hydrocarbon compounds. Genes coding for transposases, efflux pumps conferring resistance to heavy metals, and TonR-type outer membrane receptors are selectively enriched in the genome of S. fuliginis ATCC 27551 and appear to contribute to the adaptive potential of the organism to challenging and harsh environments. © 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

Toxin and genome evolution in a Drosophila defensive symbiosis.

Defenses conferred by microbial symbionts play a vital role in the health and fitness of their animal hosts. An important outstanding question in the study of defensive symbiosis is what determines long term stability and effectiveness against diverse natural enemies. In this study, we combine genome and transcriptome sequencing, symbiont transfection and parasite protection experiments, and toxin activity assays to examine the evolution of the defensive symbiosis between Drosophila flies and their vertically transmitted Spiroplasma bacterial symbionts, focusing in particular on ribosome-inactivating proteins (RIPs), symbiont-encoded toxins that have been implicated in protection against both parasitic wasps and nematodes. Although many strains of Spiroplasma, including the male-killing symbiont (sMel) of Drosophila melanogaster, protect against parasitic wasps, only the strain (sNeo) that infects the mycophagous fly Drosophila neotestacea appears to protect against parasitic nematodes. We find that RIP repertoire is a major differentiating factor between strains that do and do not offer nematode protection, and that sMel RIPs do not show activity against nematode ribosomes in vivo. We also discovered a strain of Spiroplasma infecting a mycophagous phorid fly, Megaselia nigra. Although both the host and its Spiroplasma are distantly related to D. neotestacea and its symbiont, genome sequencing revealed that the M. nigra symbiont encodes abundant and diverse RIPs, including plasmid-encoded toxins that are closely related to the RIPs in sNeo. Our results suggest that distantly related Spiroplasma RIP toxins may perform specialized functions with regard to parasite specificity and suggest an important role for horizontal gene transfer in the emergence of novel defensive phenotypes.

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April 21, 2020

Confident phylogenetic identification of uncultured prokaryotes through long read amplicon sequencing of the 16S-ITS-23S rRNA operon.

Amplicon sequencing of the 16S rRNA gene is the predominant method to quantify microbial compositions and to discover novel lineages. However, traditional short amplicons often do not contain enough information to confidently resolve their phylogeny. Here we present a cost-effective protocol that amplifies a large part of the rRNA operon and sequences the amplicons with PacBio technology. We tested our method on a mock community and developed a read-curation pipeline that reduces the overall read error rate to 0.18%. Applying our method on four environmental samples, we captured near full-length rRNA operon amplicons from a large diversity of prokaryotes. The method operated at moderately high-throughput (22286-37,850 raw ccs reads) and generated a large amount of putative novel archaeal 23S rRNA gene sequences compared to the archaeal SILVA database. These long amplicons allowed for higher resolution during taxonomic classification by means of long (~1000 bp) 16S rRNA gene fragments and for substantially more confident phylogenies by means of combined near full-length 16S and 23S rRNA gene sequences, compared to shorter traditional amplicons (250 bp of the 16S rRNA gene). We recommend our method to those who wish to cost-effectively and confidently estimate the phylogenetic diversity of prokaryotes in environmental samples at high throughput. © 2019 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

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April 21, 2020

The complete mitochondrial genome sequence of Schisandra chinensis (Austrobaileyales: Schisandraceae)

Chinese magnolia vine (Schisandra chinensis) is an economically important oriental medicinal plant that belongs to the Schisandraceae family. The complete mitochondrial genome sequence of S. chinensis was 946,141bp in length. A total of 45 genes was annotated, including 30 protein-coding genes, 12 tRNA genes, and 3 rRNA genes. A phylogenetic tree based on the mitochondrial genome demonstrated that S. chinensis was most closely related to Schisandra sphenanthera of the Schisandraceae family.

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April 21, 2020

Long-read sequence capture of the haemoglobin gene clusters across codfish species.

Combining high-throughput sequencing with targeted sequence capture has become an attractive tool to study specific genomic regions of interest. Most studies have so far focused on the exome using short-read technology. These approaches are not designed to capture intergenic regions needed to reconstruct genomic organization, including regulatory regions and gene synteny. Here, we demonstrate the power of combining targeted sequence capture with long-read sequencing technology for comparative genomic analyses of the haemoglobin (Hb) gene clusters across eight species separated by up to 70 million years. Guided by the reference genome assembly of the Atlantic cod (Gadus morhua) together with genome information from draft assemblies of selected codfishes, we designed probes covering the two Hb gene clusters. Use of custom-made barcodes combined with PacBio RSII sequencing led to highly continuous assemblies of the LA (~100 kb) and MN (~200 kb) clusters, which include syntenic regions of coding and intergenic sequences. Our results revealed an overall conserved genomic organization of the Hb genes within this lineage, yet with several, lineage-specific gene duplications. Moreover, for some of the species examined, we identified amino acid substitutions at two sites in the Hbb1 gene as well as length polymorphisms in its regulatory region, which has previously been linked to temperature adaptation in Atlantic cod populations. This study highlights the use of targeted long-read capture as a versatile approach for comparative genomic studies by generation of a cross-species genomic resource elucidating the evolutionary history of the Hb gene family across the highly divergent group of codfishes. © 2018 The Authors. Molecular Ecology Resources Published by John Wiley & Sons Ltd.

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