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July 7, 2019

Comparative analyses of clinical and environmental populations of Cryptococcus neoformans in Botswana.

Cryptococcus neoformans var. grubii (Cng) is the most common cause of fungal meningitis, and its prevalence is highest in sub-Saharan Africa. Patients become infected by inhaling airborne spores or desiccated yeast cells from the environment, where the fungus thrives in avian droppings, trees and soil. To investigate the prevalence and population structure of Cng in southern Africa, we analysed isolates from 77 environmental samples and 64 patients. We detected significant genetic diversity among isolates and strong evidence of geographic structure at the local level. High proportions of isolates with the rare MATa allele were observed in both clinical and environmental isolates; however, the mating-type alleles were unevenly distributed among different subpopulations. Nearly equal proportions of the MATa and MATa mating types were observed among all clinical isolates and in one environmental subpopulation from the eastern part of Botswana. As previously reported, there was evidence of both clonality and recombination in different geographic areas. These results provide a foundation for subsequent genomewide association studies to identify genes and genotypes linked to pathogenicity in humans. © 2015 The Authors. Molecular Ecology published by John Wiley & Sons Ltd.


July 7, 2019

Co-cultivation and transcriptome sequencing of two co-existing fish pathogens Moritella viscosa and Aliivibrio wodanis.

Aliivibrio wodanis and Moritella viscosa have often been isolated concurrently from fish with winter-ulcer disease. Little is known about the interaction between the two bacterial species and how the presence of one bacterial species affects the behaviour of the other.The impact on bacterial growth in co-culture was investigated in vitro, and the presence of A. wodanis has an inhibitorial effect on M. viscosa. Further, we have sequenced the complete genomes of these two marine Gram-negative species, and have performed transcriptome analysis of the bacterial gene expression levels from in vivo samples. Using bacterial implants in the fish abdomen, we demonstrate that the presence of A. wodanis is altering the gene expression levels of M. viscosa compared to when the bacteria are implanted separately.From expression profiling of the transcriptomes, it is evident that the presence of A. wodanis is altering the global gene expression of M. viscosa. Co-cultivation studies showed that A. wodanis is impeding the growth of M. viscosa, and that the inhibitorial effect is not contact-dependent.


July 7, 2019

Common cell shape evolution of two nasopharyngeal pathogens.

Respiratory infectious diseases are the third cause of worldwide death. The nasopharynx is the portal of entry and the ecological niche of many microorganisms, of which some are pathogenic to humans, such as Neisseria meningitidis and Moraxella catarrhalis. These microbes possess several surface structures that interact with the actors of the innate immune system. In our attempt to understand the past evolution of these bacteria and their adaption to the nasopharynx, we first studied differences in cell wall structure, one of the strongest immune-modulators. We were able to show that a modification of peptidoglycan (PG) composition (increased proportion of pentapeptides) and a cell shape change from rod to cocci had been selected for along the past evolution of N. meningitidis. Using genomic comparison across species, we correlated the emergence of the new cell shape (cocci) with the deletion, from the genome of N. meningitidis ancestor, of only one gene: yacF. Moreover, the reconstruction of this genetic deletion in a bacterium harboring the ancestral version of the locus together with the analysis of the PG structure, suggest that this gene is coordinating the transition from cell elongation to cell division. Accompanying the loss of yacF, the elongation machinery was also lost by several of the descendants leading to the change in the PG structure observed in N. meningitidis. Finally, the same evolution was observed for the ancestor of M. catarrhalis. This suggests a strong selection of these genetic events during the colonization of the nasopharynx. This selection may have been forced by the requirement of evolving permissive interaction with the immune system, the need to reduce the cellular surface exposed to immune attacks without reducing the intracellular storage capacity, or the necessity to better compete for adhesion to target cells.


July 7, 2019

Complete genome sequence of Piscirickettsia salmonis LF-89 (ATCC VR-1361) a major pathogen of farmed salmonid fish.

Piscirickettsia salmonis, the causative agent of salmonid rickettsial septicemia (SRS), is a significant threat to the healthy and sustainable production of salmonid farming industry. This Gram-negative bacterium, originally isolated from a coho salmon in Southern Chile, produces a systemic infection characterized by colonization of several fish organs. P. salmonis is able to infect, survive, and replicate inside salmonid macrophages however little is known about its mechanisms of pathogenesis. Here, we present the whole genome sequence and annotation of the P. salmonis reference strain LF-89 (ATCC VR-1361). The genome contains one circular chromosome of 3,184,851bp and three plasmids, pPSLF89-1 (180,124bp), pPSLF89-2 (33,516bp) and pPSLF89-3 (51,573bp). A total of 2850 protein-coding genes, 56 tRNAs and six copies of 5S-16S-23S rRNA. Copyright © 2015 Elsevier B.V. All rights reserved.


July 7, 2019

The assembly and characterisation of two structurally distinct cattle MHC class I haplotypes point to the mechanisms driving diversity.

In cattle, there are six classical MHC class I genes that are variably present between different haplotypes. Almost all known haplotypes contain between one and three genes, with an allele of Gene 2 present on the vast majority. However, very little is known about the sequence and therefore structure and evolutionary history of this genomic region. To address this, we have refined the MHC class I region in the Hereford cattle genome assembly and sequenced a complete A14 haplotype from a homozygous Holstein. Comparison of the two haplotypes revealed extensive variation within the MHC class Ia region, but not within the flanking regions, with each gene contained within a conserved 63- to 68-kb sequence block. This variable region appears to have undergone block gene duplication and likely deletion at regular breakpoints, suggestive of a site-specific mechanism. Phylogenetic analysis using complete gene sequences provided evidence of allelic diversification via gene conversion, with breakpoints between each of the extracellular domains that were associated with high guanine-cytosine (GC) content. Advancing our knowledge of cattle MHC class I evolution will help inform investigations of cattle genetic diversity and disease resistance.


July 7, 2019

Essential roles of methionine and S-adenosylmethionine in the autarkic lifestyle of Mycobacterium tuberculosis.

Multidrug resistance, strong side effects, and compliance problems in TB chemotherapy mandate new ways to kill Mycobacterium tuberculosis (Mtb). Here we show that deletion of the gene encoding homoserine transacetylase (metA) inactivates methionine and S-adenosylmethionine (SAM) biosynthesis in Mtb and renders this pathogen exquisitely sensitive to killing in immunocompetent or immunocompromised mice, leading to rapid clearance from host tissues. Mtb ?metA is unable to proliferate in primary human macrophages, and in vitro starvation leads to extraordinarily rapid killing with no appearance of suppressor mutants. Cell death of Mtb ?metA is faster than that of other auxotrophic mutants (i.e., tryptophan, pantothenate, leucine, biotin), suggesting a particularly potent mechanism of killing. Time-course metabolomics showed complete depletion of intracellular methionine and SAM. SAM depletion was consistent with a significant decrease in methylation at the DNA level (measured by single-molecule real-time sequencing) and with the induction of several essential methyltransferases involved in biotin and menaquinone biosynthesis, both of which are vital biological processes and validated targets of antimycobacterial drugs. Mtb ?metA could be partially rescued by biotin supplementation, confirming a multitarget cell death mechanism. The work presented here uncovers a previously unidentified vulnerability of Mtb-the incapacity to scavenge intermediates of SAM and methionine biosynthesis from the host. This vulnerability unveils an entirely new drug target space with the promise of rapid killing of the tubercle bacillus by a new mechanism of action.


July 7, 2019

A multidrug resistance plasmid contains the molecular switch for type VI secretion in Acinetobacter baumannii.

Infections with Acinetobacter baumannii, one of the most troublesome and least studied multidrug-resistant superbugs, are increasing at alarming rates. A. baumannii encodes a type VI secretion system (T6SS), an antibacterial apparatus of Gram-negative bacteria used to kill competitors. Expression of the T6SS varies among different strains of A. baumannii, for which the regulatory mechanisms are unknown. Here, we show that several multidrug-resistant strains of A. baumannii harbor a large, self-transmissible resistance plasmid that carries the negative regulators for T6SS. T6SS activity is silenced in plasmid-containing, antibiotic-resistant cells, while part of the population undergoes frequent plasmid loss and activation of the T6SS. This activation results in T6SS-mediated killing of competing bacteria but renders A. baumannii susceptible to antibiotics. Our data show that a plasmid that has evolved to harbor antibiotic resistance genes plays a role in the differentiation of cells specialized in the elimination of competing bacteria.


July 7, 2019

Complete genome sequence of Acinetobacter baumannii strain B8300, which displays high twitching motility.

Acinetobacter baumannii has emerged as an important nosocomial pathogen causing health care-associated infections. In this study, we determined the genome of a twitching-positive clinical strain, B8300, isolated from a hospital in southern India. De novo assembly of PacBio long-read sequencing data generated the B8300 genome that consists of a chromosome of 3.82 Mbp and a plasmid of 25.15 kbp. Copyright © 2015 Vijaykumar et al.


July 7, 2019

Complete genome sequence of Achromobacter xylosoxidans MN001, a cystic fibrosis airway isolate.

The genome of Achromobacter xylosoxidans MN001, a strain isolated from sputum derived from an adult cystic fibrosis patient, was sequenced using combined single-molecule real-time and Illumina sequencing. Assembly of the complete genome resulted in a 5,876,039-bp chromosome, representing the smallest A. xylosoxidans genome sequenced to date. Copyright © 2015 Badalamenti and Hunter.


July 7, 2019

Complete genome sequences of low-passage virulent and high-passage avirulent variants of pathogenic Leptospira interrogans serovar Manilae strain UP-MMC-NIID, originally isolated from a patient with severe leptospirosis, determined using PacBio Single-Molecule Real-Time technology.

Here, we report the complete genome sequences of low-passage virulent and high-passage avirulent variants of pathogenic Leptospira interrogans serovar Manilae strain UP-MMC-NIID, a major causative agent of leptospirosis. While there were no major differences between the genome sequences, the levels of base modifications were higher in the avirulent variant. Copyright © 2015 Satou et al.


July 7, 2019

Complete genome sequence of Acinetobacter baumannii strain B8342, a motility-positive clinical isolate.

Acinetobacter baumannii is an emerging Gram-negative pathogen responsible for health care-associated infections. In this study, we determined the genome of a motility-positive clinical strain, B8342, isolated from a hospital in southern India. The B8342 genome, which is 3.94 Mbp, was generated by de novo assembly of PacBio long-read sequencing data. Copyright © 2015 Vijaykumar et al.


July 7, 2019

Comparative genomics and characterization of hybrid Shigatoxigenic and enterotoxigenic Escherichia coli (STEC/ETEC) strains.

Shigatoxigenic Escherichia coli (STEC) and enterotoxigenic E. coli (ETEC) cause serious foodborne infections in humans. These two pathogroups are defined based on the pathogroup-associated virulence genes: stx encoding Shiga toxin (Stx) for STEC and elt encoding heat-labile and/or est encoding heat-stable enterotoxin (ST) for ETEC. The study investigated the genomics of STEC/ETEC hybrid strains to determine their phylogenetic position among E. coli and to define the virulence genes they harbor.The whole genomes of three STEC/ETEC strains possessing both stx and est genes were sequenced using PacBio RS sequencer. Two of the strains were isolated from the patients, one with hemolytic uremic syndrome, and one with diarrhea. The third strain was of bovine origin. Core genome analysis of the shared chromosomal genes and comparison with E. coli and Shigella spp. reference genomes was performed to determine the phylogenetic position of the STEC/ETEC strains. In addition, a set of virulence genes and ETEC colonization factors were extracted from the genomes. The production of Stx and ST were studied.The human STEC/ETEC strains clustered with strains representing ETEC, STEC, enteroaggregative E. coli, and commensal and laboratory-adapted E. coli. However, the bovine STEC/ETEC strain formed a remote cluster with two STECs of bovine origin. All three STEC/ETEC strains harbored several other virulence genes, apart from stx and est, and lacked ETEC colonization factors. Two STEC/ETEC strains produced both toxins and one strain Stx only.This study shows that pathogroup-associated virulence genes of different E. coli can co-exist in strains originating from different phylogenetic lineages. The possibility of virulence genes to be associated with several E. coli pathogroups should be taken into account in strain typing and in epidemiological surveillance. Development of novel hybrid E. coli strains may cause a new public health risk, which challenges the traditional diagnostics of E. coli infections.


July 7, 2019

Complete genome sequences of three Neisseria gonorrhoeae laboratory reference Strains, determined using PacBio Single-Molecule Real-Time technology.

Neisseria gonorrhoeae, the etiological agent that causes the sexually transmitted infection gonorrhea, is a significant public health concern due to the emergence of antimicrobial resistance. We report the complete genome sequences of three reference isolates with varied antimicrobial susceptibility that will aid in elucidating the genetic mechanisms that confer resistance. Copyright © 2015 Abrams et al.


July 7, 2019

Genome sequences of two multidrug-resistant Acinetobacter baumannii clinical strains isolated from Southern India

Acinetobacter baumannii is an emerging nosocomial pathogen causing infections worldwide. In this study, we determined the genome sequences of two multidrug-resistant A. baumannii clinical strains isolated from a hospital in southern India. Genome analyses indicate that both the strains harbor numerous horizontally transferred genetic elements and antibiotic resistance cassettes. Copyright © 2015 Balaji et al.


July 7, 2019

Advances in DNA sequencing technologies for high resolution HLA typing.

This communication describes our experience in large-scale G group-level high resolution HLA typing using three different DNA sequencing platforms – ABI 3730 xl, Illumina MiSeq and PacBio RS II. Recent advances in DNA sequencing technologies, so-called next generation sequencing (NGS), have brought breakthroughs in deciphering the genetic information in all living species at a large scale and at an affordable level. The NGS DNA indexing system allows sequencing multiple genes for large number of individuals in a single run. Our laboratory has adopted and used these technologies for HLA molecular testing services. We found that each sequencing technology has its own strengths and weaknesses, and their sequencing performances complement each other. HLA genes are highly complex and genotyping them is quite challenging. Using these three sequencing platforms, we were able to meet all requirements for G group-level high resolution and high volume HLA typing. Copyright © 2015 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.


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