A combined multi-virulence-locus sequence typing and Staphylococcal Cassette Chromosome mec typing scheme possesses enhanced discriminatory power for genotyping MRSA

Verghese, Bindhu; Schwalm, Nathan D.; Dudley, Edward G.; Knabel, Stephen J.

doi:10.1016/j.meegid.2012.07.026

Cited by 16 publications

(11 citation statements)

References 36 publications

(37 reference statements)

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“…Multi Locus Sequence Typing (MLST) was performed using consensus sequences from each sample for seven housekeeping genes (arc, aroE, glp, gmk, pta, tpi, yqiL) using standard programs available at (saureus.mlst.net) to assign allele and sequence types (STs). Similarly, all samples were analyzed for six virulence genes (alt, essC, geh, hlgA, htrA, and sdrC) as previously proposed for MLST [12]. Analysis of the virulence gene alleles in the study samples was performed via neighbor joining analysis with 10,000 bootstrap (the CLC bio Rapid Neighbor-Joining plugin).…”

Section: Methodsmentioning

confidence: 99%

“…It is possible that virulence genes, in isolation or combination, may play a role in the pathogenic behavior of specific strains, but this has been difficult to prove with traditional approaches to bacterial genetics using pulse field gel electrophoresis (PFGE) or multi-locus sequence typing (MLST) because they focus on macromolecular patterns or on core housekeeping genes which are common to all Staphylococcus aureus strains [11,12]. Bacterial isolates from persons with nasal carriage, community-acquired invasive disease, and hospital-acquired invasive disease appear to be evenly distributed among S .…”

Section: Introductionmentioning

confidence: 99%

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Genomic Heterogeneity of Methicillin Resistant Staphylococcus aureus Associated with Variation in Severity of Illness among Children with Acute Hematogenous Osteomyelitis

et al. 2015

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IntroductionThe association between severity of illness of children with osteomyelitis caused by Methicillin-resistant Staphylococcus aureus (MRSA) and genomic variation of the causative organism has not been previously investigated. The purpose of this study is to assess genomic heterogeneity among MRSA isolates from children with osteomyelitis who have diverse severity of illness.Materials and MethodsChildren with osteomyelitis were prospectively studied between 2010 and 2011. Severity of illness of the affected children was determined from clinical and laboratory parameters. MRSA isolates were analyzed with next generation sequencing (NGS) and optical mapping. Sequence data was used for multi-locus sequence typing (MLST), phylogenetic analysis by maximum likelihood (PAML), and identification of virulence genes and single nucleotide polymorphisms (SNP) relative to reference strains.ResultsThe twelve children studied demonstrated severity of illness scores ranging from 0 (mild) to 9 (severe). All isolates were USA300, ST 8, SCC mec IVa MRSA by MLST. The isolates differed from reference strains by 2 insertions (40 Kb each) and 2 deletions (10 and 25 Kb) but had no rearrangements or copy number variations. There was a higher occurrence of virulence genes among study isolates when compared to the reference strains (p = 0.0124). There were an average of 11 nonsynonymous SNPs per strain. PAML demonstrated heterogeneity of study isolates from each other and from the reference strains.DiscussionGenomic heterogeneity exists among MRSA isolates causing osteomyelitis among children in a single community. These variations may play a role in the pathogenesis of variation in clinical severity among these children.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Genomic Heterogeneity of Methicillin Resistant Staphylococcus aureus Associated with Variation in Severity of Illness among Children with Acute Hematogenous Osteomyelitis

et al. 2015

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“…Moreover, attempts have been undertaken to develop MLST schemes that are entirely based on virulence genes. Such approaches, termed multivirulence-locus sequence typing (MVLST), have been applied for the subtyping of pathogens like Listeria monocytogenes, V. cholerae, S. enterica and S. aureus [69][70][71][72]. Altogether, the currently available data suggest that MVLST is endowed with a higher discriminatory power than that of the 'classical' MLST.…”

Section: Multilocus Sequence Typingmentioning

confidence: 99%

Overview of molecular typing methods for outbreak detection and epidemiological surveillance

et al. 2013

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Typing methods for discriminating different bacterial isolates of the same species are essential epidemiological tools in infection prevention and control. Traditional typing systems based on phenotypes, such as serotype, biotype, phage-type, or antibiogram, have been used for many years. However, more recent methods that examine the relatedness of isolates at a molecular level have revolutionised our ability to differentiate among bacterial types and subtypes. Importantly, the development of molecular methods has provided new tools for enhanced surveillance and outbreak detection. This has resulted in better implementation of rational infection control programmes and efficient allocation of resources across Europe. The emergence of benchtop sequencers using next generation sequencing technology makes bacterial whole genome sequencing (WGS) feasible even in small research and clinical laboratories. WGS has already been used for the characterisation of bacterial isolates in several large outbreaks in Europe and, in the near future, is likely to replace currently used typing methodologies due to its ultimate resolution. However, WGS is still too laborious and time-consuming to obtain useful data in routine surveillance. Also, a largely unresolved question is how genome sequences must be examined for epidemiological characterisation. In the coming years, the lessons learnt from currently used molecular methods will allow us to condense the WGS data into epidemiologically useful information. On this basis, we have reviewed current and new molecular typing methods for outbreak detection and epidemiological surveillance of bacterial pathogens in clinical practice, aiming to give an overview of their specific advantages and disadvantages.

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“…It has been shown that multi-virulence-locus sequence typing (MVLST) is a more powerful typing approach than traditional MLST (76)(77)(78)(79). Addition of such a scheme to CRISPR sequencing (CRISPR-MVLST) in Salmonella (using fimH and sseL virulence genes) increased discriminatory power compared to either method used alone (50).…”

Section: Crispr-mvlstmentioning

confidence: 99%

CRISPRs: Molecular Signatures Used for Pathogen Subtyping

Shariat

Dudley

2014

Appl Environ Microbiol

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132

115

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Rapid and accurate strain identification is paramount in the battle against microbial outbreaks, and several subtyping approaches have been developed. One such method uses clustered regular interspaced short palindromic repeats (CRISPRs), DNA repeat elements that are present in approximately half of all bacteria. Though their signature function is as an adaptive immune system against invading DNA such as bacteriophages and plasmids, CRISPRs also provide an excellent framework for pathogen tracking and evolutionary studies. Analysis of the spacer DNA sequences that reside between the repeats has been tremendously useful for bacterial subtyping during molecular epidemiological investigations. Subtyping, or strain identification, using CRISPRs has been employed in diverse Gram-positive and Gram-negative bacteria, including Mycobacterium tuberculosis, Salmonella enterica, and the plant pathogen Erwinia amylovora. This review discusses the several ways in which CRISPR sequences are exploited for subtyping. This includes the well-established spoligotyping methodologies that have been used for 2 decades to type Mycobacterium species, as well as in-depth consideration of newer, higher-throughput CRISPR-based protocols.

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A combined multi-virulence-locus sequence typing and Staphylococcal Cassette Chromosome mec typing scheme possesses enhanced discriminatory power for genotyping MRSA

Cited by 16 publications

References 36 publications

Genomic Heterogeneity of Methicillin Resistant Staphylococcus aureus Associated with Variation in Severity of Illness among Children with Acute Hematogenous Osteomyelitis

Genomic Heterogeneity of Methicillin Resistant Staphylococcus aureus Associated with Variation in Severity of Illness among Children with Acute Hematogenous Osteomyelitis

Overview of molecular typing methods for outbreak detection and epidemiological surveillance

CRISPRs: Molecular Signatures Used for Pathogen Subtyping

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