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July 7, 2019

Complete genome sequence of Planococcus donghaensis JH1(T), a pectin-degrading bacterium.

The type strain Planococcus donghaensis JH1(T) is a psychrotolerant and halotolerant bacterium with starch-degrading ability. Here, we determine the carbon utilization profile of P. donghaensis JH1(T) and report the first complete genome of the strain. This study revealed the strain’s ability to utilize pectin and d-galacturonic acid, and identified genes responsible for degradation of the polysaccharides. The genomic information provided may serve as a fundamental resource for full exploration of the biotechnological potential of P. donghaensis JH1(T). Copyright © 2017. Published by Elsevier B.V.

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July 7, 2019

Whole genome characterization of a naturally occurring vancomycin-dependent Enterococcus faecium from a patient with bacteremia.

Vancomycin-dependent enterococci are a relatively uncommon phenotype recovered in the clinical laboratory. Recognition and recovery of these isolates are important, to provide accurate identification and susceptibility information to treating physicians. Herein, we describe the recovery of a vancomycin-dependent and revertant E. faecium isolates harboring vanB operon from a patient with bacteremia. Using whole genome sequencing, we found a unique single nucleotide polymorphism (S186N) in the D-Ala-D-Ala ligase (ddl) conferring vancomycin-dependency. Additionally, we found that a majority of in vitro revertants mutated outside ddl, with some strains harboring mutations in vanS, while others likely containing novel mechanisms of reversion. Copyright © 2017 Elsevier B.V. All rights reserved.

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July 7, 2019

Plasmid dynamics in Vibrio parahaemolyticus strains related to shrimp Acute Hepatopancreatic Necrosis Syndrome (AHPNS).

Vibrio parahaemolyticus is a causative agent of acute hapatopancreatic necrosis syndrome (AHPNS) which causes early mortality in white shrimp. Emergence of AHPNS has caused tremendous economic loss for aquaculture industry particularly in Asia since 2010. Previous studies reported that strains causing AHPNS harbor a 69-kb plasmid with possession of virulence genes, pirA and pirB. However, genetic variation of the 69-kb plasmid among AHPNS related strains has not been investigated. This study aimed to analyze genetic composition and diversity of the 69-kb plasmid in strains isolated from shrimps affected by AHPNS. Plasmids recovered from V. parahaemolyticus strain VPE61 which represented typical AHPNS pathogenicity, strain VP2HP which did not represent AHPNS pathogenicity but was isolated from AHPNS affected shrimp and other AHPNS V. parahaemolyticus isolates in Genbank were investigated. Protein coding genes of the 69-kb plasmid from the strain VPE61 were identical to that of AHPNS strain from Vietnam except the inverted complement 3.4-kb transposon covering pirA and pirB. The strain VP2HP possessed remarkable large 183-kb plasmid which shared similar protein coding genes to those of the 69-kb plasmid from strain VPE61. However, the 3.4-kb transposon covering pirA and pirB was absent from the 183-kb plasmid in strain VP2HP. A number of protein coding genes from the 183-kb plasmid were also detected in other AHPNS strains. In summary, this study identified a novel 183-kb plasmid that is related to AHPNS causing strains. Homologous recombination of the 69-kb AHPNS plasmid and other naturally occurring plasmids together with loss and gain of AHPNS virulence genes in V. parahaemolyticus were observed. The outcome of this research enables understanding of plasmid dynamics that possibly affect variable degrees of AHPNS pathogenicity. Copyright © 2017 Elsevier B.V. All rights reserved.

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July 7, 2019

Targeted disruption of the polyketide synthase gene pks15 affects virulence against insects and phagocytic survival in the fungus Beauveria bassiana.

The reducing clade III polyketide synthase genes, including pks15, are highly conserved among entomopathogenic fungi. To examine the function of pks15, we used targeted disruption to investigate the impact of Beauveria bassiana pks15 on insect pathogenesis. Southern analysis verified that the ?pks15 mutant was disrupted by a single integration of the transformation cassette at the pks15 locus. The ?pks15 mutant had a slight reduction in radial growth, and it produced fewer spores. Our insect bioassays indicated the ?pks15 mutant to be significantly reduced in virulence against beet armyworms compared to wild type (WT), which could be partially accounted for by its markedly decreased ability to survive phagocytosis. Total haemocyte count decreased sharply by 50-fold from days 1-3 post-inoculation in insects infected with WT, compared to a 5-fold decrease in the ?pks15 mutant. The mutant also produced fewer hemolymph hyphal bodies than WT by 3-fold. In co-culture studies with amoebae that have phagocytic ability similar to that of insect haemocytes, at 48 h the mortality rate of amoebae engulfing ?pks15 decreased by 72 %, and ?pks15 CFU decreased by 83 % compared to co-culture with WT. Thus, the ?pks15 mutant had a reduced ability to cope with phagocytosis and highly reduced virulence in an insect host. These data elucidate a mechanism of insect pathogenesis associated with polyketide biosynthesis. Copyright © 2017 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

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July 7, 2019

Comparison of pseudorabies virus China reference strain with emerging variants reveals independent virus evolution within specific geographic regions.

Pseudorabies virus (PRV) China reference strain Ea is genetically closely related to newly emerged variants; however, there is limited information about PRV Ea. Here, we compared PRV Ea with new variant strains by growth kinetics, genome sequencing, and protein expression analysis. Growth analysis showed that strain Ea forms smaller plaques than strain HNX. The full-length genome sequence of Ea revealed that it is clustered in the same subgroup as HNX. Ea and HNX strains exhibited similar extracellular virion protein polymorphisms, whereas strain Bartha expressed less VP26 and more GAPDH. In infected cells, strain Ea expressed high levels of IE180 protein, and Ea and HNX produced higher levels of UL21 protein than strain Bartha. These findings provide evidence that PRV China reference strain Ea is genetically closely related to the newly emerged variant strains, indicating that strain PRV China may have evolved independently leading to the emergence of a variant strain. Copyright © 2017 Elsevier Inc. All rights reserved.

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July 7, 2019

Complete genome sequence of Bacillus velezensis LM2303, a biocontrol strain isolated from the dung of wild yak inhabited Qinghai-Tibet plateau.

Bacillus velezensis LM2303 is a biocontrol strain with a broad inhibitory spectrum against plant pathogens, isolated from the dung of wild yak inhabited Qinghai-Tibet plateau, China. Here we present its complete genome sequence, which consists of a single, circular chromosome of 3,989,393bp with a 46.68% G+C content. Genome analysis revealed genes encoding specialized functions for the biosynthesis of antifungal metabolites and antibacterial metabolites, the promotion of plant growth, the alleviation of oxidative stress and nutrient utilization. And the biosynthesis of antimicrobial metabolites in strain LM2303 was confirmed by biochemical analysis, while its plant growth promoting traits were confirmed by inoculation tests. Our results will establish a better foundation for further studies and biocontrol application of B. velezensis LM2303. Copyright © 2017 Elsevier B.V. All rights reserved.

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July 7, 2019

Complete genome sequence of Bacillus sp. 275, producing extracellular cellulolytic, xylanolytic and ligninolytic enzymes.

Technologies for degradation of three major components of lignocellulose (e.g. cellulose, hemicellulose and lignin) are needed to efficiently utilize lignocellulose. Here, we report Bacillus sp. 275 isolated from a mudflat exhibiting various lignocellulolytic activities including cellulase, xylanase, laccase and peroxidase in the cell culture supernatant. The complete genome of Bacillus sp. 275 strain contains 3832 protein cording sequences and an average G+C content of 46.32% on one chromosome (4045,581bp) and one plasmid (6389bp). The genes encoding enzymes related to the degradation of cellulose, xylan and lignin were detected in the Bacillus sp. 275 genome. In addition, the genes encoding glucosidases that hydrolyze starch, mannan, galactoside and arabinan were also found in the genome, implying that Bacillus sp. 275 has potentially a wide range of uses in the degradation of polysaccharide in lignocellulosic biomasses. Copyright © 2017 Elsevier B.V. All rights reserved.

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July 7, 2019

Complete genome sequence and analysis of three kinds of ß-agarase of Cellulophaga lytica DAU203 isolated from marine sediment.

Cellulophaga lytica DAU203 (KACC 19187), isolated from the marine sediment in Korea, has a strong ability to degrade agar. The genome of C. lytica DAU203 contains a single chromosome that is 3,952,957bp in length, with 32.02% G+C contents. The genomic information predicted that the DAU203 has the potential to be utilized in various enzymatic industries. Copyright © 2017. Published by Elsevier B.V.

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July 7, 2019

The complete genome sequence of Bacillus velezensis 9912D reveals its biocontrol mechanism as a novel commercial biological fungicide agent.

A Bacillus sp. 9912 mutant, 9912D, was approved as a new biological fungicide agent by the Ministry of Agriculture of the People’s Republic of China in 2016 owing to its excellent inhibitory effect on various plant pathogens and being environment-friendly. Here, we present the genome of 9912D with a circular chromosome having 4436 coding DNA sequences (CDSs), and a circular plasmid encoding 59 CDSs. This strain was finally designated as Bacillus velezensis based on phylogenomic analyses. Genome analysis revealed a total of 19 candidate gene clusters involved in secondary metabolite biosynthesis, including potential new type II lantibiotics. The absence of fengycin biosynthetic gene cluster is noteworthy. Our data offer insights into the genetic, biological and physiological characteristics of this strain and aid in deeper understanding of its biocontrol mechanism. Copyright © 2017 Elsevier B.V. All rights reserved.

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July 7, 2019

Clostridium chauvoei, an evolutionary dead-end pathogen.

Full genome sequences of 20 strains of Clostridium chauvoei, the etiological agent of blackleg of cattle and sheep, isolated from four different continents over a period of 64 years (1951-2015) were determined and analyzed. The study reveals that the genome of the species C. chauvoei is highly homogeneous compared to the closely related species C. perfringens, a widespread pathogen that affects human and many animal species. Analysis of the CRISPR locus is sufficient to differentiate most C. chauvoei strains and is the most heterogenous region in the genome, containing in total 187 different spacer elements that are distributed as 30 – 77 copies in the various strains. Some genetic differences are found in the 3 allelic variants of fliC1, fliC2 and fliC3 genes that encode structural flagellin proteins, and certain strains do only contain one or two alleles. However, the major virulence genes including the highly toxic C.chauvoei toxin A, the sialidase and the two hyaluronidases are fully conserved as are the metabolic and structural genes of C. chauvoei. These data indicate that C. chauvoei is a strict ruminant-associated pathogen that has reached a dead end in its evolution.

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July 7, 2019

Geno- and phenotypic characteristics of a transfected Babesia bovis 6-Cys-E knockout clonal line.

Babesia bovis is an intra-erythrocytic tick-transmitted apicomplexan protozoan parasite. It has a complex lifestyle including asexual replication in the mammalian host and sexual replication occurring in the midgut of host tick vector, typically, Rhipicephalus microplus. Previous evidence showed that certain B. bovis genes, including members of 6-Cys gene family, are differentially expressed during tick and mammalian stages of the parasite’s life cycle. Moreover, the 6-Cys E gene is differentially expressed in the T3Bo strain of B. bovis tick stages, and anti 6-Cys E antibodies were shown to be able to inhibit in vitro growth of the phenotypically distinct B. bovis Mo7clonal line.In this study, the 6-Cys E gene of B. bovis T3Bo strain was disrupted by transfection using a plasmid containing 6-Cys gene E 5′ and 3′ regions to guide homologous recombination, and the egfp-bsd fusion gene under control of a ef-1a promoter, yielding a B. bovis clonal line designated 6-Cys EKO-cln. Full genome sequencing of 6-Cys EKO-cln parasites was performed and in vitro inhibition assays using anti 6-Cys E antibodies.Full genome sequencing of 6-Cys EKO-cln B. bovis demonstrated single insertion of egfp-bsd gene that disrupts the integrity of 6-Cys gene E. Undistinguishable growth rate of 6-Cys EKO-cln line compared to wild-type 6-Cys E intact T3Bo B. bovis strain in in vitro cultures indicates that expression of gene 6-Cys E is not essential for blood stage replication in this strain. In vitro inhibition assays confirmed the ability of anti-6 Cys E antibodies to inhibit the growth of the wild-type Mo7 and T3Bo B. bovis parasites, but no significant inhibition was found for 6-Cys EKO-cln line parasites.Overall, the data suggest that the anti-6 Cys E antibody neutralising effect on the wild type strains is likely due to mechanical hindrance, or cross-reactivity, rather than due to functional requirements of 6-Cys gene E product for survival and development of the erythrocyte stages. Further investigation is underway to determine if the 6-Cys E protein is required for replication and sexual stage development of B. bovis during tick stages.

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July 7, 2019

PipeCraft: Flexible open-source toolkit for bioinformatics analysis of custom high-throughput amplicon sequencing data.

High-throughput sequencing methods have become a routine analysis tool in environmental sciences as well as in public and private sector. These methods provide vast amount of data, which need to be analysed in several steps. Although the bioinformatics may be applied using several public tools, many analytical pipelines allow too few options for the optimal analysis for more complicated or customized designs. Here, we introduce PipeCraft, a flexible and handy bioinformatics pipeline with a user-friendly graphical interface that links several public tools for analysing amplicon sequencing data. Users are able to customize the pipeline by selecting the most suitable tools and options to process raw sequences from Illumina, Pacific Biosciences, Ion Torrent and Roche 454 sequencing platforms. We described the design and options of PipeCraft and evaluated its performance by analysing the data sets from three different sequencing platforms. We demonstrated that PipeCraft is able to process large data sets within 24 hr. The graphical user interface and the automated links between various bioinformatics tools enable easy customization of the workflow. All analytical steps and options are recorded in log files and are easily traceable.© 2017 John Wiley & Sons Ltd.

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July 7, 2019

Phylogeography of Burkholderia pseudomallei isolates, Western Hemisphere.

The bacterium Burkholderia pseudomallei causes melioidosis, which is mainly associated with tropical areas. We analyzed single-nucleotide polymorphisms (SNPs) among genome sequences from isolates of B. pseudomallei that originated in the Western Hemisphere by comparing them with genome sequences of isolates that originated in the Eastern Hemisphere. Analysis indicated that isolates from the Western Hemisphere form a distinct clade, which supports the hypothesis that these isolates were derived from a constricted seeding event from Africa. Subclades have been resolved that are associated with specific regions within the Western Hemisphere and suggest that isolates might be correlated geographically with cases of melioidosis. One isolate associated with a former World War II prisoner of war was believed to represent illness 62 years after exposure in Southeast Asia. However, analysis suggested the isolate originated in Central or South America.

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July 7, 2019

Critical points for an accurate human genome analysis.

Next-generation sequencing is radically changing how DNA diagnostic laboratories operate. What started as a single-gene profession is now developing into gene panel sequencing and whole-exome and whole-genome sequencing (WES/WGS) analyses. With further advances in sequencing technology and concomitant price reductions, WGS will soon become the standard and be routinely offered. Here, we focus on the critical steps involved in performing WGS, with a particular emphasis on points where WGS differs from WES, the important variables that should be taken into account, and the quality control measures that can be taken to monitor the process. The points discussed here, combined with recent publications on guidelines for reporting variants, will facilitate the routine implementation of WGS into a diagnostic setting.© 2017 Wiley Periodicals, Inc.

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July 7, 2019

Antibody-independent mechanisms regulate the establishment of chronic Plasmodium infection.

Malaria is caused by parasites of the genus Plasmodium. All human-infecting Plasmodium species can establish long-lasting chronic infections(1-5), creating an infectious reservoir to sustain transmission(1,6). It is widely accepted that the maintenance of chronic infection involves evasion of adaptive immunity by antigenic variation(7). However, genes involved in this process have been identified in only two of five human-infecting species: Plasmodium falciparum and Plasmodium knowlesi. Furthermore, little is understood about the early events in the establishment of chronic infection in these species. Using a rodent model we demonstrate that from the infecting population, only a minority of parasites, expressing one of several clusters of virulence-associated pir genes, establishes a chronic infection. This process occurs in different species of parasites and in different hosts. Establishment of chronicity is independent of adaptive immunity and therefore different from the mechanism proposed for maintenance of chronic P. falciparum infections(7-9). Furthermore, we show that the proportions of parasites expressing different types of pir genes regulate the time taken to establish a chronic infection. Because pir genes are common to most, if not all, species of Plasmodium(10), this process may be a common way of regulating the establishment of chronic infections.

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