July 7, 2019  |  

The genome of the toluene-degrading Pseudomonas veronii strain 1YdBTEX2 and its differential gene expression in contaminated sand.

The natural restoration of soils polluted by aromatic hydrocarbons such as benzene, toluene, ethylbenzene and m- and p-xylene (BTEX) may be accelerated by inoculation of specific biodegraders (bioaugmentation). Bioaugmentation mainly involves introducing bacteria that deploy their metabolic properties and adaptation potential to survive and propagate in the contaminated environment by degrading the pollutant. In order to better understand the adaptive response of cells during a transition to contaminated material, we analyzed here the genome and short-term (1 h) changes in genome-wide gene expression of the BTEX-degrading bacterium Pseudomonas veronii 1YdBTEX2 in non-sterile soil and liquid medium, both in presence or absence of toluene. We obtained a gapless genome sequence of P. veronii 1YdBTEX2 covering three individual replicons with a total size of 8 Mb, two of which are largely unrelated to current known bacterial replicons. One-hour exposure to toluene, both in soil and liquid, triggered massive transcription (up to 208-fold induction) of multiple gene clusters, such as toluene degradation pathway(s), chemotaxis and toluene efflux pumps. This clearly underlines their key role in the adaptive response to toluene. In comparison to liquid medium, cells in soil drastically changed expression of genes involved in membrane functioning (e.g., lipid composition, lipid metabolism, cell fatty acid synthesis), osmotic stress response (e.g., polyamine or trehalose synthesis, uptake of potassium) and putrescine metabolism, highlighting the immediate response mechanisms of P. veronii 1YdBTEX2 for successful establishment in polluted soil.


July 7, 2019  |  

Complete, closed genome sequences of 10 Salmonella enterica subsp. enterica serovar Typhimurium strains isolated from human and bovine sources.

Salmonella enterica is a leading cause of enterocolitis for humans and animals. S. enterica subsp. enterica serovar Typhimurium infects a broad range of hosts. To facilitate genomic comparisons among isolates from different sources, we present the complete genome sequences of 10 S Typhimurium strains, 5 each isolated from human and bovine sources. Copyright © 2016 Nguyen et al.


July 7, 2019  |  

Listeria monocytogenes in stone fruits linked to a multistate outbreak: enumeration of cells and whole-genome sequencing.

In 2014, the identification of stone fruits contaminated with Listeria monocytogenes led to the subsequent identification of a multistate outbreak. Simultaneous detection and enumeration of L. monocytogenes were performed on 105 fruits, each weighing 127 to 145 g, collected from 7 contaminated lots. The results showed that 53.3% of the fruits yielded L. monocytogenes (lower limit of detection, 5 CFU/fruit), and the levels ranged from 5 to 2,850 CFU/fruit, with a geometric mean of 11.3 CFU/fruit (0.1 CFU/g of fruit). Two serotypes, IVb-v1 and 1/2b, were identified by a combination of PCR- and antiserum-based serotyping among isolates from fruits and their packing environment; certain fruits contained a mixture of both serotypes. Single nucleotide polymorphism (SNP)-based whole-genome sequencing (WGS) analysis clustered isolates from two case-patients with the serotype IVb-v1 isolates and distinguished outbreak-associated isolates from pulsed-field gel electrophoresis (PFGE)-matched, but epidemiologically unrelated, clinical isolates. The outbreak-associated isolates differed by up to 42 SNPs. All but one serotype 1/2b isolate formed another WGS cluster and differed by up to 17 SNPs. Fully closed genomes of isolates from the stone fruits were used as references to maximize the resolution and to increase our confidence in prophage analysis. Putative prophages were conserved among isolates of each WGS cluster. All serotype IVb-v1 isolates belonged to singleton sequence type 382 (ST382); all but one serotype 1/2b isolate belonged to clonal complex 5.WGS proved to be an excellent tool to assist in the epidemiologic investigation of listeriosis outbreaks. The comparison at the genome level contributed to our understanding of the genetic diversity and variations among isolates involved in an outbreak or isolates associated with food and environmental samples from one facility. Fully closed genomes increased our confidence in the identification and comparison of accessory genomes. The diversity among the outbreak-associated isolates and the inclusion of PFGE-matched, but epidemiologically unrelated, isolates demonstrate the high resolution of WGS. The prevalence and enumeration data could contribute to our further understanding of the risk associated with Listeria monocytogenes contamination, especially among high-risk populations. Copyright © 2016 Chen et al.


July 7, 2019  |  

Genomewide Dam methylation in Escherichia coli during long-term stationary phase.

DNA methylation in prokaryotes is widespread. The most common modification of the genome is the methylation of adenine at the N-6 position. In Escherichia coli K-12 and many gammaproteobacteria, this modification is catalyzed by DNA adenine methyltransferase (Dam) at the GATC consensus sequence and is known to modulate cellular processes including transcriptional regulation of gene expression, initiation of chromosomal replication, and DNA mismatch repair. While studies thus far have focused on the motifs associated with methylated adenine (meA), the frequency of meA across the genome, and temporal dynamics during early periods of incubation, here we conduct the first study on the temporal dynamics of adenine methylation in E. coli by Dam throughout all five phases of the bacterial life cycle in the laboratory. Using single-molecule real-time sequencing, we show that virtually all GATC sites are significantly methylated over time; nearly complete methylation of the chromosome was confirmed by mass spectroscopy analysis. However, we also detect 66 sites whose methylation patterns change significantly over time within a population, including three sites associated with sialic acid transport and catabolism, suggesting a potential role for Dam regulation of these genes; differential expression of this subset of genes was confirmed by quantitative real-time PCR. Further, we show significant growth defects of the dam mutant during long-term stationary phase (LTSP). Together these data suggest that the cell places a high premium on fully methylating the chromosome and that alterations in methylation patterns may have significant impact on patterns of transcription, maintenance of genetic fidelity, and cell survival. IMPORTANCE While it has been shown that methylation remains relatively constant into early stationary phase of E. coli, this study goes further through death phase and long-term stationary phase, a unique time in the bacterial life cycle due to nutrient limitation and strong selection for mutants with increased fitness. The absence of methylation at GATC sites can influence the mutation frequency within a population due to aberrant mismatch repair. Therefore, it is important to investigate the methylation status of GATC sites in an environment where cells may not prioritize methylation of the chromosome. This study demonstrates that chromosome methylation remains a priority even under conditions of nutrient limitation, indicating that continuous methylation at GATC sites could be under positive selection.


July 7, 2019  |  

MICADo – Looking for mutations in targeted PacBio cancer data: an alignment-free method.

Targeted sequencing is commonly used in clinical application of NGS technology since it enables generation of sufficient sequencing depth in the targeted genes of interest and thus ensures the best possible downstream analysis. This notwithstanding, the accurate discovery and annotation of disease causing mutations remains a challenging problem even in such favorable context. The difficulty is particularly salient in the case of third generation sequencing technology, such as PacBio. We present MICADo, a de Bruijn graph based method, implemented in python, that makes possible to distinguish between patient specific mutations and other alterations for targeted sequencing of a cohort of patients. MICADo analyses NGS reads for each sample within the context of the data of the whole cohort in order to capture the differences between specificities of the sample with respect to the cohort. MICADo is particularly suitable for sequencing data from highly heterogeneous samples, especially when it involves high rates of non-uniform sequencing errors. It was validated on PacBio sequencing datasets from several cohorts of patients. The comparison with two widely used available tools, namely VarScan and GATK, shows that MICADo is more accurate, especially when true mutations have frequencies close to backgound noise. The source code is available at http://github.com/cbib/MICADo.


July 7, 2019  |  

Spontaneous chloroplast mutants mostly occur by replication slippage and show a biased pattern in the plastome of Oenothera.

Spontaneous plastome mutants have been used as a research tool since the beginning of genetics. However, technical restrictions have severely limited their contributions to research in physiology and molecular biology. Here, we used full plastome sequencing to systematically characterize a collection of 51 spontaneous chloroplast mutants in Oenothera (evening primrose). Most mutants carry only a single mutation. Unexpectedly, the vast majority of mutations do not represent single nucleotide polymorphisms but are insertions/deletions originating from DNA replication slippage events. Only very few mutations appear to be caused by imprecise double-strand break repair, nucleotide misincorporation during replication, or incorrect nucleotide excision repair following oxidative damage. U-turn inversions were not detected. Replication slippage is induced at repetitive sequences that can be very small and tend to have high A/T content. Interestingly, the mutations are not distributed randomly in the genome. The underrepresentation of mutations caused by faulty double-strand break repair might explain the high structural conservation of seed plant plastomes throughout evolution. In addition to providing a fully characterized mutant collection for future research on plastid genetics, gene expression, and photosynthesis, our work identified the spectrum of spontaneous mutations in plastids and reveals that this spectrum is very different from that in the nucleus.© 2016 American Society of Plant Biologists. All rights reserved.


July 7, 2019  |  

Efficient, cost-effective, high-throughput, Multilocus Sequencing Typing (MLST) method, NGMLST, and the analytical software program MLSTEZ.

Multilocus sequence typing (MLST) has become the preferred method for genotyping many biological species. It can be used to identify major phylogenetic clades, molecular groups, or subpopulations of a species, as well as individual strains or clones. However, conventional MLST is costly and time consuming, which limits its power for genotyping large numbers of samples. Here, we describe a new MLST method that uses next-generation sequencing, a multiplexing protocol, and appropriate analytical software to provide accurate, rapid, and economical MLST genotyping of 96 or more isolates in a single assay.


July 7, 2019  |  

Origins of the current seventh cholera pandemic.

Vibrio cholerae has caused seven cholera pandemics since 1817, imposing terror on much of the world, but bacterial strains are currently only available for the sixth and seventh pandemics. The El Tor biotype seventh pandemic began in 1961 in Indonesia, but did not originate directly from the classical biotype sixth-pandemic strain. Previous studies focused mainly on the spread of the seventh pandemic after 1970. Here, we analyze in unprecedented detail the origin, evolution, and transition to pandemicity of the seventh-pandemic strain. We used high-resolution comparative genomic analysis of strains collected from 1930 to 1964, covering the evolution from the first available El Tor biotype strain to the start of the seventh pandemic. We define six stages leading to the pandemic strain and reveal all key events. The seventh pandemic originated from a nonpathogenic strain in the Middle East, first observed in 1897. It subsequently underwent explosive diversification, including the spawning of the pandemic lineage. This rapid diversification suggests that, when first observed, the strain had only recently arrived in the Middle East, possibly from the Asian homeland of cholera. The lineage migrated to Makassar, Indonesia, where it gained the important virulence-associated elements Vibrio seventh pandemic island I (VSP-I), VSP-II, and El Tor type cholera toxin prophage by 1954, and it then became pandemic in 1961 after only 12 additional mutations. Our data indicate that specific niches in the Middle East and Makassar were important in generating the pandemic strain by providing gene sources and the driving forces for genetic events.


July 7, 2019  |  

Phylogeny of dermatophytes with genomic character evaluation of clinically distinct Trichophyton rubrum and T. áviolaceum

Trichophyton rubrum and T. violaceum are prevalent agents of human dermatophyte infections, the former being found on glabrous skin and nail, while the latter is confined to the scalp. The two species are phenotypically different but are highly similar phylogenetically. The taxonomy of dermatophytes is currently being reconsidered on the basis of molecular phylogeny. Molecular species definitions do not always coincide with existing concepts which are guided by ecological and clinical principles. In this article, we aim to bring phylogenetic and ecological data together in an attempt to develop new species concepts for anthropophilic dermatophytes. Focus is on the T. rubrum complex with analysis of rDNA ITS supplemented with LSU, TUB2, TEF3 and ribosomal protein L10 gene sequences. In order to explore genomic differences between T. rubrum and T. violaceum, one representative for both species was whole genome sequenced. Draft sequences were compared with currently available dermatophyte genomes. Potential virulence factors of adhesins and secreted proteases were predicted and compared phylogenetically. General phylogeny showed clear gaps between geophilic species of Arthroderma, but multilocus distances between species were often very small in the derived anthropophilic and zoophilic genus Trichophyton. Significant genome conservation between T. rubrum and T. violaceum was observed, with a high similarity at the nucleic acid level of 99.38 % identity. Trichophyton violaceum contains more paralogs than T. rubrum. About 30 adhesion genes were predicted among dermatophytes. Seventeen adhesins were common between T. rubrum and T. violaceum, while four were specific for the former and eight for the latter. Phylogenetic analysis of secreted proteases reveals considerable expansion and conservation among the analyzed species. Multilocus phylogeny and genome comparison of T. rubrum and T. violaceum underlined their close affinity. The possibility that they represent a single species exhibiting different phenotypes due to different localizations on the human body is discussed.


July 7, 2019  |  

Genome sequence resources for the wheat stripe rust pathogen (Puccinia striiformis f. sp. tritici) and the barley stripe rust pathogen (Puccinia striiformis f. sp. hordei)

Puccinia striiformis f. sp. tritici causes devastating stripe (yellow) rust on wheat and P. striiformis f. sp. hordei causes stripe rust on barley. Several P. striiformis f. sp. tritici genomes are available, but no P. striiformis f. sp. hordei genome is available. More genomes of P. striiformis f. sp. tritici and P. striiformis f. sp. hordei are needed to understand the genome evolution and molecular mechanisms of their pathogenicity. We sequenced P. striiformis f. sp. tritici isolate 93-210 and P. striiformis f. sp. hordei isolate 93TX-2, using PacBio and Illumina technologies and RNA sequencing. Their genomic sequences were assembled to contigs with high continuity and showed significant structural differences. The circular mitochondria genomes of both were complete. These genomes provide high-quality resources for deciphering the genomic basis of rapid evolution and host adaptation, identifying genes for avirulence and other important traits, and studying host-pathogen interactions.


July 7, 2019  |  

Complete and assembled genome sequence of an NDM-5- and CTX-M-15-producing Escherichia coli sequence type 617 isolated from wastewater in Switzerland.

Carbapenem-resistant Escherichia coli have emerged worldwide and represent a major challenge to effective healthcare management. Here we report the genome sequence of an NDM-5- and CTX-M-15-producing E. coli belonging to sequence type 617 isolated from wastewater treatment plant effluent in Switzerland.Whole-genome sequencing of E. coli 657SK2 was performed using Pacific Biosciences (PacBio) single-molecule real-time (SMRT) technology RS2 reads (C4/P6 chemistry). De novo assembly was carried out using Canu 1.6, and sequences were annotated using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP).The genome of E. coli 657SK2 consists of a 4.9-Mbp chromosome containing blaCTX-M-15, genes associated with virulence [fyuA, hlyE, the pyelonephritis-associated pili (pap) gene cluster and the yad gene cluster], the copper resistance gene pco, and genes associated with resistance to quaternary ammonium compound (QAC) disinfectants (emrA, mdfA and sugE). A 173.9-kb multidrug resistance IncFII-FIA-FIB plasmid was detected harbouring aadA2, aadA5, blaNDM-5, blaOXA-1, cat, drfA, drfA17, the mph(A)-mrx-mphR cluster, the tetA-tetC-tetR cluster, and the virulence genes iutA and ylpA.The genome sequence of E. coli 657SK2 provides information on resistance mechanisms and virulence characteristics of pathogenic E. coli harbouring blaNDM-5 and blaCTX-M-15 that are spreading into the environment via urban wastewater.Copyright © 2018 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.


July 7, 2019  |  

Genome sequence of Halomonas hydrothermalis Y2, an efficient ectoine-producer isolated from pulp mill wastewater.

Halophilic microorganisms have great potentials towards biotechnological applications. Halomonas hydrothermalis Y2 is a halotolerant and alkaliphilic strain that isolated from the Na+-rich pulp mill wastewater. The strain is dominant in the bacterial community of pulp mill wastewater and exhibits metabolic diversity in utilizing various substrates. Here we present the genome sequence of this strain, which comprises a circular chromosome 3,933,432 bp in size and a GC content of 60.2%. Diverse genes that encoding proteins for compatible solutes synthesis and transport were identified from the genome. With a complete pathway for ectoine synthesis, the strain could produce ectoine from monosodium glutamate and further partially secreted into the medium. In addition, around 20% ectoine was increased by deleting the ectoine hydroxylase (EctD). The genome sequence we report here will provide genetic information regarding adaptive mechanisms of strain Y2 to its harsh habitat, as well as facilitate exploration of metabolic strategies for diverse compatible solutes, e.g., ectoine production. Copyright © 2018 Elsevier B.V. All rights reserved.


July 7, 2019  |  

Genome analysis of Vallitalea guaymasensis strain L81 isolated from a deep-sea hydrothermal vent system.

Abyssivirga alkaniphila strain L81T, recently isolated from a black smoker biofilm at the Loki’s Castle hydrothermal vent field, was previously described as a mesophilic, obligately anaerobic heterotroph able to ferment carbohydrates, peptides, and aliphatic hydrocarbons. The strain was classified as a new genus within the family Lachnospiraceae. Herein, its genome is analyzed and A. alkaniphila is reassigned to the genus Vallitalea as a new strain of V. guaymasensis, designated V. guaymasensis strain L81. The 6.4 Mbp genome contained 5651 protein encoding genes, whereof 4043 were given a functional prediction. Pathways for fermentation of mono-saccharides, di-saccharides, peptides, and amino acids were identified whereas a complete pathway for the fermentation of n-alkanes was not found. Growth on carbohydrates and proteinous compounds supported methane production in co-cultures with Methanoplanus limicola. Multiple confurcating hydrogen-producing hydrogenases, a putative bifurcating electron-transferring flavoprotein—butyryl-CoA dehydrogenase complex, and a Rnf-complex form a basis for the observed hydrogen-production and a putative reverse electron-transport in V. guaymasensis strain L81. Combined with the observation that n-alkanes did not support growth in co-cultures with M. limicola, it seemed more plausible that the previously observed degradation patterns of crude-oil in strain L81 are explained by unspecific activation and may represent a detoxification mechanism, representing an interesting ecological function. Genes encoding a capacity for polyketide synthesis, prophages, and resistance to antibiotics shows interactions with the co-occurring microorganisms. This study enlightens the function of the fermentative microorganisms from hydrothermal vents systems and adds valuable information on the bioprospecting potential emerging in deep-sea hydrothermal systems.


July 7, 2019  |  

Complete genome sequence of WM99c, an antibiotic-resistant Acinetobacter baumannii global clone 2 (GC2) strain representing an Australian GC2 lineage.

The extensively antibiotic-resistant Acinetobacter baumannii isolate WM99c recovered in Sydney, Australia, in 1999 is an early representative of a distinct lineage of global clone 2 (GC2) seen on the east coast of Australia. We present the complete 4.121-Mbp genome sequence (chromosome plus 2 plasmids), generated via long-read sequencing (PacBio).


Talk with an expert

If you have a question, need to check the status of an order, or are interested in purchasing an instrument, we're here to help.