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July 7, 2019

Quorum sensing activity of Aeromonas caviae strain YL12, a bacterium isolated from compost.

Quorum sensing is a well-studied cell-to-cell communication method that involves a cell-density dependent regulation of genes expression mediated by signalling molecules. In this study, a bacterium isolated from a plant material compost pile was found to possess quorum sensing activity based on bioassay screening. Isolate YL12 was identified using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry and molecular typing using rpoD gene which identified the isolate as Aeromonas caviae. High resolution tandem mass spectrometry was subsequently employed to identify the N-acyl homoserine lactone profile of Aeromonas caviae YL12 and confirmed that this isolate produced two short chain N-acyl homoserine lactones, namely C4-HSL and C6, and the production was observed to be cell density-dependent. Using the thin layer chromatography (TLC) bioassay, both AHLs were found to activate C. violaceum CV026, whereas only C6-HSL was revealed to induce bioluminescence expression of E. coli [pSB401]. The data presented in this study will be the leading steps in understanding the role of quorum sensing in Aeromonas caviae strain YL12.

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July 7, 2019

The Glanville fritillary genome retains an ancient karyotype and reveals selective chromosomal fusions in Lepidoptera.

Previous studies have reported that chromosome synteny in Lepidoptera has been well conserved, yet the number of haploid chromosomes varies widely from 5 to 223. Here we report the genome (393?Mb) of the Glanville fritillary butterfly (Melitaea cinxia; Nymphalidae), a widely recognized model species in metapopulation biology and eco-evolutionary research, which has the putative ancestral karyotype of n=31. Using a phylogenetic analyses of Nymphalidae and of other Lepidoptera, combined with orthologue-level comparisons of chromosomes, we conclude that the ancestral lepidopteran karyotype has been n=31 for at least 140?My. We show that fusion chromosomes have retained the ancestral chromosome segments and very few rearrangements have occurred across the fusion sites. The same, shortest ancestral chromosomes have independently participated in fusion events in species with smaller karyotypes. The short chromosomes have higher rearrangement rate than long ones. These characteristics highlight distinctive features of the evolutionary dynamics of butterflies and moths.

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July 7, 2019

Complete genome sequences of nitrofurantoin-sensitive and -resistant Escherichia coli ST540 and ST2747 strains.

Widespread multidrug resistance in Escherichia coli has necessitated the reintroduction of older antibiotics, such as nitrofurantoin. However, mechanisms by which resistance to nitrofurantoin emerges in E. coli are not well elucidated. Toward this aim, we sequenced two nitrofurantoin-sensitive E. coli sequence types (ST540 and ST2747) and their four nitrofurantoin-resistant derivatives generated in vitro under aerobic and anaerobic growth conditions.

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July 7, 2019

Whole-genome sequence of Burkholderia sp. strain RPE67, a bacterial gut symbiont of the bean bug Riptortus pedestris.

Burkholderia sp. strain RPE67 is a bacterial symbiont isolated from a field-collected bean bug, Riptortus pedestris. To understand the genetic basis of the insect-microbe symbiosis, we performed whole-genome sequencing of the Burkholderia strain, revealing an 8.69-Mb genome consisting of three chromosomes and three plasmids. Copyright © 2014 Takeshita et al.

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July 7, 2019

Complete genome sequence of Kosakonia sacchari type strain SP1(T.).

Kosakonia sacchari sp. nov. is a new species within the new genus Kosakonia, which was included in the genus Enterobacter. K sacchari is a nitrogen-fixing bacterium named for its association with sugarcane (Saccharum officinarum L.). K sacchari bacteria are Gram-negative, aerobic, non-spore-forming, motile rods. Strain SP1(T) (=CGMCC1.12102(T)=LMG 26783(T)) is the type strain of the K sacchari sp. nov and is able to colonize and fix N2 in association with sugarcane plants, thus promoting plant growth. Here we summarize the features of strain SP1(T) and describe its complete genome sequence. The genome contains a single chromosome and no plasmids, 4,902,024 nucleotides with 53.7% GC content, 4,460 protein-coding genes and 105 RNA genes including 22 rRNA genes, 82 tRNA genes, and 1 ncRNA gene.

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July 7, 2019

FASTQSim: platform-independent data characterization and in silico read generation for NGS datasets.

High-throughput next generation sequencing technologies have enabled rapid characterization of clinical and environmental samples. Consequently, the largest bottleneck to actionable data has become sample processing and bioinformatics analysis, creating a need for accurate and rapid algorithms to process genetic data. Perfectly characterized in silico datasets are a useful tool for evaluating the performance of such algorithms.Background contaminating organisms are observed in sequenced mixtures of organisms. In silico samples provide exact truth. To create the best value for evaluating algorithms, in silico data should mimic actual sequencer data as closely as possible.FASTQSim is a tool that provides the dual functionality of NGS dataset characterization and metagenomic data generation. FASTQSim is sequencing platform-independent, and computes distributions of read length, quality scores, indel rates, single point mutation rates, indel size, and similar statistics for any sequencing platform. To create training or testing datasets, FASTQSim has the ability to convert target sequences into in silico reads with specific error profiles obtained in the characterization step.FASTQSim enables users to assess the quality of NGS datasets. The tool provides information about read length, read quality, repetitive and non-repetitive indel profiles, and single base pair substitutions. FASTQSim allows the user to simulate individual read datasets that can be used as standardized test scenarios for planning sequencing projects or for benchmarking metagenomic software. In this regard, in silico datasets generated with the FASTQsim tool hold several advantages over natural datasets: they are sequencing platform independent, extremely well characterized, and less expensive to generate. Such datasets are valuable in a number of applications, including the training of assemblers for multiple platforms, benchmarking bioinformatics algorithm performance, and creating challenge datasets for detecting genetic engineering toolmarks, etc.

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July 7, 2019

Site-specific genetic engineering of the Anopheles gambiae Y chromosome.

Despite its function in sex determination and its role in driving genome evolution, the Y chromosome remains poorly understood in most species. Y chromosomes are gene-poor, repeat-rich and largely heterochromatic and therefore represent a difficult target for genetic engineering. The Y chromosome of the human malaria vector Anopheles gambiae appears to be involved in sex determination although very little is known about both its structure and function. Here, we characterize a transgenic strain of this mosquito species, obtained by transposon-mediated integration of a transgene construct onto the Y chromosome. Using meganuclease-induced homologous repair we introduce a site-specific recombination signal onto the Y chromosome and show that the resulting docking line can be used for secondary integration. To demonstrate its utility, we study the activity of a germ-line-specific promoter when located on the Y chromosome. We also show that Y-linked fluorescent transgenes allow automated sex separation of this important vector species, providing the means to generate large single-sex populations. Our findings will aid studies of sex chromosome function and enable the development of male-exclusive genetic traits for vector control.

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July 7, 2019

Genomic reconnaissance of clinical isolates of emerging human pathogen Mycobacterium abscessus reveals high evolutionary potential.

Mycobacterium abscessus (Ma) is an emerging human pathogen that causes both soft tissue infections and systemic disease. We present the first comparative whole-genome study of Ma strains isolated from patients of wide geographical origin. We found a high proportion of accessory strain-specific genes indicating an open, non-conservative pan-genome structure, and clear evidence of rapid phage-mediated evolution. Although we found fewer virulence factors in Ma compared to M. tuberculosis, our data indicated that Ma evolves rapidly and therefore should be monitored closely for the acquisition of more pathogenic traits. This comparative study provides a better understanding of Ma and forms the basis for future functional work on this important pathogen.

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July 7, 2019

Genomic insights into the taxonomic status of the three subspecies of Bacillus subtilis.

Bacillus subtilis contains three subspecies, i.e., subspecies subtilis, spizizenii, and inaquosorum. As these subspecies are phenotypically indistinguishable, their differentiation has relied on phylogenetic analysis of multiple protein-coding gene sequences. B. subtilis subsp. inaquosorum is a recently proposed taxon that encompasses strain KCTC 13429(T) and related strains, which were previously classified as members of subspecies spizizenii. However, DNA-DNA hybridization (DDH) values among the three subspecies raised a question as to their independence. Thus, we evaluated the taxonomic status of subspecies inaquosorum using genome-based comparative analysis. In contrast to the previous experimental values of DDH, the inter-genomic relatedness inferred by average nucleotide identity (ANI) values indicated that subspecies inaquosorum and spizizenii were sufficiently different from subspecies subtilis and hence raised the possibility that the former two could be classified as separate species from B. subtilis. The genome-based tree also supported the separation of the two subspecies from B. subtilis. The exclusive presence of a subtilin synthesis system in subspecies spizizenii was a remarkable genetic characteristic that could even distinguish subspecies spizizenii from subspecies inaquosorum in addition to the low ANI values (<95%). Conclusively, the genome-based data obtained in this study demonstrated that subspecies inaquosorum and spizizenii are clearly distinguished from subspecies subtilis, and raises the possibility that these two subspecies could be classified as separate species from B. subtilis. In addition, the low ANI values between subspecies inaquosorum and spizizenii and the exclusive presence of subtilin synthesis genes in subspecies spizizenii also suggest circumscription of these two subspecies at the species level. Copyright © 2013 Elsevier GmbH. All rights reserved.

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July 7, 2019

The effects of read length, quality and quantity on microsatellite discovery and primer development: from Illumina to PacBio.

The advent of next-generation sequencing (NGS) technologies has transformed the way microsatellites are isolated for ecological and evolutionary investigations. Recent attempts to employ NGS for microsatellite discovery have used the 454, Illumina, and Ion Torrent platforms, but other methods including single-molecule real-time DNA sequencing (Pacific Biosciences or PacBio) remain viable alternatives. We outline a workflow from sequence quality control to microsatellite marker validation in three plant species using PacBio circular consensus sequencing (CCS). We then evaluate the performance of PacBio CCS in comparison with other NGS platforms for microsatellite isolation, through simulations that focus on variations in read length, read quantity and sequencing error rate. Although quality control of CCS reads reduced microsatellite yield by around 50%, hundreds of microsatellite loci that are expected to have improved conversion efficiency to functional markers were retrieved for each species. The simulations quantitatively validate the advantages of long reads and emphasize the detrimental effects of sequencing errors on NGS-enabled microsatellite development. In view of the continuing improvement in read length on NGS platforms, sequence quality and the corresponding strategies of quality control will become the primary factors to consider for effective microsatellite isolation. Among current options, PacBio CCS may be optimal for rapid, small-scale microsatellite development due to its flexibility in scaling sequencing effort, while platforms such as Illumina MiSeq will provide cost-efficient solutions for multispecies microsatellite projects. © 2014 John Wiley & Sons Ltd.

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July 7, 2019

A fault-tolerant method for HLA typing with PacBio data.

Human leukocyte antigen (HLA) genes are critical genes involved in important biomedical aspects, including organ transplantation, autoimmune diseases and infectious diseases. The gene family contains the most polymorphic genes in humans and the difference between two alleles is only a single base pair substitution in many cases. The next generation sequencing (NGS) technologies could be used for high throughput HLA typing but in silico methods are still needed to correctly assign the alleles of a sample. Computer scientists have developed such methods for various NGS platforms, such as Illumina, Roche 454 and Ion Torrent, based on the characteristics of the reads they generate. However, the method for PacBio reads was less addressed, probably owing to its high error rates. The PacBio system has the longest read length among available NGS platforms, and therefore is the only platform capable of having exon 2 and exon 3 of HLA genes on the same read to unequivocally solve the ambiguity problem caused by the “phasing” issue.We proposed a new method BayesTyping1 to assign HLA alleles for PacBio circular consensus sequencing reads using Bayes’ theorem. The method was applied to simulated data of the three loci HLA-A, HLA-B and HLA-DRB1. The experimental results showed its capability to tolerate the disturbance of sequencing errors and external noise reads.The BayesTyping1 method could overcome the problems of HLA typing using PacBio reads, which mostly arise from sequencing errors of PacBio reads and the divergence of HLA genes, to some extent.

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July 7, 2019

Improved draft genome sequence of Clostridium pasteurianum strain ATCC 6013 (DSM 525) using a hybrid next-generation sequencing approach.

We present an improved draft genome sequence for Clostridium pasteurianum strain ATCC 6013 (DSM 525), the type strain of the species and an important solventogenic bacterium with industrial potential. Availability of a near-complete genome sequence will enable strain engineering of this promising bacterium. Copyright © 2014 Pyne et al.

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July 7, 2019

Compact genome of the Antarctic midge is likely an adaptation to an extreme environment.

The midge, Belgica antarctica, is the only insect endemic to Antarctica, and thus it offers a powerful model for probing responses to extreme temperatures, freeze tolerance, dehydration, osmotic stress, ultraviolet radiation and other forms of environmental stress. Here we present the first genome assembly of an extremophile, the first dipteran in the family Chironomidae, and the first Antarctic eukaryote to be sequenced. At 99 megabases, B. antarctica has the smallest insect genome sequenced thus far. Although it has a similar number of genes as other Diptera, the midge genome has very low repeat density and a reduction in intron length. Environmental extremes appear to constrain genome architecture, not gene content. The few transposable elements present are mainly ancient, inactive retroelements. An abundance of genes associated with development, regulation of metabolism and responses to external stimuli may reflect adaptations for surviving in this harsh environment.

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July 7, 2019

Automated ensemble assembly and validation of microbial genomes.

The continued democratization of DNA sequencing has sparked a new wave of development of genome assembly and assembly validation methods. As individual research labs, rather than centralized centers, begin to sequence the majority of new genomes, it is important to establish best practices for genome assembly. However, recent evaluations such as GAGE and the Assemblathon have concluded that there is no single best approach to genome assembly. Instead, it is preferable to generate multiple assemblies and validate them to determine which is most useful for the desired analysis; this is a labor-intensive process that is often impossible or unfeasible.To encourage best practices supported by the community, we present iMetAMOS, an automated ensemble assembly pipeline; iMetAMOS encapsulates the process of running, validating, and selecting a single assembly from multiple assemblies. iMetAMOS packages several leading open-source tools into a single binary that automates parameter selection and execution of multiple assemblers, scores the resulting assemblies based on multiple validation metrics, and annotates the assemblies for genes and contaminants. We demonstrate the utility of the ensemble process on 225 previously unassembled Mycobacterium tuberculosis genomes as well as a Rhodobacter sphaeroides benchmark dataset. On these real data, iMetAMOS reliably produces validated assemblies and identifies potential contamination without user intervention. In addition, intelligent parameter selection produces assemblies of R. sphaeroides comparable to or exceeding the quality of those from the GAGE-B evaluation, affecting the relative ranking of some assemblers.Ensemble assembly with iMetAMOS provides users with multiple, validated assemblies for each genome. Although computationally limited to small or mid-sized genomes, this approach is the most effective and reproducible means for generating high-quality assemblies and enables users to select an assembly best tailored to their specific needs.

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