Heather Carleton

Complete genome sequence of USA300, an epidemic clone of community-acquired meticillin-resistant Staphylococcus aureus

BACKGROUND USA300, a clone of meticillin-resistant Staphylococcus aureus, is a major source of community-acquired infections in the USA, Canada, and Europe. Our aim was to sequence its genome and compare it with those of other strains of S aureus to try to identify genes responsible for its distinctive epidemiological and virulence properties. METHODS We ascertained the genome sequence of FPR3757, a multidrug resistant USA300 strain, by random shotgun sequencing, then compared it with the sequences of ten other staphylococcal strains. FINDINGS Compared with closely related S aureus, we noted that almost all of the unique genes in USA300 clustered in novel allotypes of mobile genetic elements. Some of the unique genes are involved in pathogenesis, including Panton-Valentine leucocidin and molecular variants of enterotoxin Q and K. The most striking feature of the USA300 genome is the horizontal acquisition of a novel mobile genetic element that encodes an arginine deiminase pathway and an oligopeptide permease system that could contribute to growth and survival of USA300. We did not detect this element, termed arginine catabolic mobile element (ACME), in other S aureus strains. We noted a high prevalence of ACME in S epidermidis, suggesting not only that ACME transfers into USA300 from S epidermidis, but also that this element confers a selective advantage to this ubiquitous commensal of the human skin. INTERPRETATION USA300 has acquired mobile genetic elements that encode resistance and virulence determinants that could enhance fitness and pathogenicity.

Roles of 34 Virulence Genes in the Evolution of Hospital- and Community-Associated Strains of Methicillin-Resistant Staphylococcus aureus

BACKGROUND The extent to which the horizontal transfer of virulence genes has contributed to the emergence of contemporary virulent strains of methicillin-resistant Staphylococcus aureus (MRSA) in hospital and community settings is poorly understood. METHODS Epidemiologically well-characterized MRSA isolates collected over 8.5 years were genotyped and tested for the presence of 34 virulence genes. RESULTS Six strain types accounted for 88.2% of all MRSA infections. The evolution of contemporary hospital and community phenotypes within the CC8 and CC30 lineages--2 background genomes that produced historical pandemic MRSA clones--were associated with multiple horizontal acquisitions of virulence genes. The epidemic community phenotype of a CC8 strain, designated ST8:USA300, was linked to the acquisition of staphylococcal cassette chromosome (SCC)mec type IV, the genes for Panton-Valentine leukocidin (PVL), and the enterotoxin Q and K genes. Similarly, the epidemic community phenotype of a CC30 strain, ST30:USA1100, was linked to the acquisition of SCCmec type IV and the pvl genes. In contrast, the epidemic hospital phenotype of another CC30 strain, ST36:USA200, was associated with the acquisition of SCCmec type II, the enterotoxin A gene, and the toxic shock syndrome toxin 1 gene. The pvl genes appear not to be essential for the evolution OF other community-associated strains of mrsa, including ST8:USA500 and ST59:USA1000. CONCLUSIONS The horizontal transfer of virulence genes, although infrequent, is epidemiologically associated with the emergence of new virulent strains of MRSA.

Widespread Skin and Soft-Tissue Infections Due to Two Methicillin-Resistant Staphylococcus aureus Strains Harboring the Genes for Panton-Valentine Leucocidin

ABSTRACT Infections caused by community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) are emerging as a major public health problem. CA-MRSA has been associated previously with skin and soft-tissue infection (SSTI) and with carriage of staphylococcal cassette chromosome mec (SCCmec) type IV and the Panton-Valentine leucocidin (PVL) virulence factor. To assess the clonal distribution of PVL-carrying strains and the association with SSTI in the San Francisco Bay area, we surveyed six collections of S. aureus isolates—671 isolates in all—collected between 1997 and 2002 originating from inpatient and outpatient clinical specimens and from a community-based sampling. Isolates were genotyped by pulsed-field gel electrophoresis, multilocus restriction fragment typing, and multilocus sequence typing and assayed for the PVL virulence factor. The S. aureus populations showed a high proportion of PVL-carrying strains, with frequencies ranging up to 70% in MRSA isolated from jail inmate patients and 69% in MRSA from patients receiving surgical treatment at an outpatient clinic specializing in treating SSTIs. PVL-carrying isolates were identified in nine clonal groups, but 88.5% of the PVL-carrying MRSA isolates belonged to only two clonal groups. These two clonal groups carried the SCCmec type IV resistance determinant and were more likely than other clonal groups to be recovered from SSTI sites than from other sites (P < 0.0001). There is evidence of clonal replacement over the period from 1999 to 2002, with one of these two clonal groups being supplanted by the other.

Community-Adapted Methicillin-Resistant Staphylococcus aureus (MRSA): Population Dynamics of an Expanding Community Reservoir of MRSA

To define methicillin-resistant Staphylococcus aureus (MRSA) reservoirs in the community and their population dynamics, we studied the molecular epidemiology of a random sample (n=490) from a collection of 2154 inpatient and outpatient MRSA isolates during a 7-year period in San Francisco. We noted a progressive replacement of type II staphylococcal chromosomal cassette (SCC)mec-bearing isolates with type IV SCCmec-bearing isolates, which coincided with >4-fold increase in methicillin resistance between 1998 and 2002. Type IV SCCmec-bearing isolates involved in the increase in methicillin resistance belonged to 4 molecular genotypes. These 4 genotypes were associated predominantly with community-onset disease, rather than hospital- or long-term-care facility-onset disease (76.9% vs. 19.4% vs. 3.7%; P=.0005), suggesting that they are not feral descendants of hospital isolates. The longitudinal results linked the dramatic increase in MRSA infections to an expanding community reservoir of MRSA genotypes with intrinsic community survival advantage.

Increasing Prevalence of Methicillin-Resistant Staphylococcus aureus Infection in California Jails

Staphylococcus aureus clinical isolates obtained from patients who were inmates of the San Francisco County jail system showed an increase in the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) from 29%, in 1997, to 74%, in 2002; 91% of the MRSA isolates carried staphylococcal chromosomal cassette mec (SCCmec) type IV. Pulsed field gel electrophoresis and multilocus sequence typing demonstrated 2 major clonal groups. One of these clonal groups is genetically indistinguishable from the strain responsible for an outbreak of MRSA in the Los Angeles County jail system in 2002.

Population Dynamics of Nasal Strains of Methicillin-Resistant Staphylococcus aureus—and Their Relation to Community-Associated Disease Activity

BACKGROUND Nasal carriage of methicillin-resistant Staphylococcus aureus (MRSA) plays a key role in the epidemiology and pathogenesis of disease. The purpose of this study was to determine the characteristics and dynamics of nasal strains of MRSA, as well as their relation to community-associated disease activity. METHODS This study is a cross-sectional survey and molecular epidemiologic analysis of nasal colonization by S. aureus in homeless and runaway youths, an underserved population at high risk for staphylococcal disease. RESULTS Of the 308 study participants, 27.6% carried S. aureus, and 6.2% carried MRSA. Subgroups of individuals with increased MRSA carriage rates were also at highest risk for community-associated MRSA infection; these subgroups included individuals with either HIV infection or AIDS, injection drug users, patients with abscesses, and those recently hospitalized. Multilocus sequence typing and pulsed-field gel electrophoresis identified 2 genotypes--ST59:P (USA1000) and ST8:S (USA300)--that accounted for 84.2% (16/19) of the MRSA isolates carried. The genotypes were distinct from nosocomial genotypes endemic in the hospital, although they originated from individuals with prior exposure to health care. CONCLUSIONS Comparison of MRSA strains from asymptomatic carriers versus concurrently collected community-associated clinical strains from patients treated at local health-care facilities allowed for the identification of 3 population dynamics of nasal strains of MRSA: (1) endemic clones--for example, ST8:C and ST59:P--sustained asymptomatic carriage and infection over prolonged periods; (2) an epidemic clone, ST8:S, demonstrated enhanced capacity for rapid transmission and widespread infections; and (3) an outbreak clone, ST30:Z (USA1100), was highly infectious but exhibited poor asymptomatic transmission.

Implementation of Nationwide Real-time Whole-genome Sequencing to Enhance Listeriosis Outbreak Detection and Investigation

Listeria monocytogenes (Lm) causes severe foodborne illness (listeriosis). Previous molecular subtyping methods, such as pulsed-field gel electrophoresis (PFGE), were critical in detecting outbreaks that led to food safety improvements and declining incidence, but PFGE provides limited genetic resolution. A multiagency collaboration began performing real-time, whole-genome sequencing (WGS) on all US Lm isolates from patients, food, and the environment in September 2013, posting sequencing data into a public repository. Compared with the year before the project began, WGS, combined with epidemiologic and product trace-back data, detected more listeriosis clusters and solved more outbreaks (2 outbreaks in pre-WGS year, 5 in WGS year 1, and 9 in year 2). Whole-genome multilocus sequence typing and single nucleotide polymorphism analyses provided equivalent phylogenetic relationships relevant to investigations; results were most useful when interpreted in context of epidemiological data. WGS has transformed listeriosis outbreak surveillance and is being implemented for other foodborne pathogens.

Whole genome-based population biology and epidemiological surveillance of Listeria monocytogenes

Listeria monocytogenes (Lm) is a major human foodborne pathogen. Numerous Lm outbreaks have been reported worldwide and associated with a high case fatality rate, reinforcing the need for strongly coordinated surveillance and outbreak control. We developed a universally applicable genome-wide strain genotyping approach and investigated the population diversity of Lm using 1,696 isolates from diverse sources and geographical locations. We define, with unprecedented precision, the population structure of Lm, demonstrate the occurrence of international circulation of strains and reveal the extent of heterogeneity in virulence and stress resistance genomic features among clinical and food isolates. Using historical isolates, we show that the evolutionary rate of Lm from lineage I and lineage II is low (∼2.5 × 10−7 substitutions per site per year, as inferred from the core genome) and that major sublineages (corresponding to so-called ‘epidemic clones’) are estimated to be at least 50–150 years old. This work demonstrates the urgent need to monitor Lm strains at the global level and provides the unified approach needed for global harmonization of Lm genome-based typing and population biology.

Intermediate vancomycin susceptibility in a community-associated MRSA clone.

We describe a case of treatment failure caused by a strain of USA300 community-associated methicillin-resistant Staphylococcus aureus (MRSA) with intermediate susceptibility to vancomycin and reduced susceptibility to daptomycin. The strain was isolated from the bone of a 56-year-old man with lumbar osteomyelitis after a 6-week treatment course of vancomycin for catheter-associated septic thrombophlebitis.

Differences in Clinical and Molecular Characteristics of Skin and Soft Tissue Methicillin-Resistant Staphylococcus aureus Isolates between Two Hospitals in Northern California

ABSTRACT Community-onset methicillin-resistant Staphylococcus aureus (CO-MRSA) skin and soft tissue infections (SSTI) are associated with SCCmec IV and Panton-Valentine leukocidin (PVL) genes. CO-MRSA epidemiologic studies suggest that genotypic variation exists within one geographic region. We compared MRSA genotypes and demographic and clinical characteristics of patients with CO-MRSA SSTI between two regional medical centers. We also examined factors associated with SCCmec IV and PVL carriage. A total of 279 MRSA SSTI isolates from 2000 to 2002 at San Francisco General Hospital (SFGH) and Stanford University Hospital (SUH) were genotyped by pulsed-field gel electrophoresis and PCR for SCCmec and PVL genes. Medical records were reviewed for clinical characteristics. Ninety-three percent and 69% of MRSA SSTI were caused by CO-MRSA at SFGH and SUH, respectively. Patients with CO-MRSA SSTI at SFGH were more likely to be nonwhite, younger, homeless, and have no previous exposure to health care (P < 0.01). SFGH CO-MRSA strains were more likely to carry SCCmec type IV and PVL genes (90% and 55%, respectively) than SUH strains (29% and 16%, respectively). In multivariate analyses, nonwhite ethnicity was associated with both SCCmec type IV and PVL carriage (odds ratio [OR] of 2.65 and 95% confidence interval [CI] of 1.19 to 5.95 and OR of 1.94 and 95% CI of 1.03 to 3.65, respectively). ST8:USA300:IV became the dominant clone at SFGH, but not at SUH, by 2002. Despite geographic proximity, CO-MRSA SSTI exhibited differing SCCmec types, PVL carriage, and clonal dynamics. CO-MRSA SSTI at SUH were more likely to represent feral isolates of nosocomial origin. Clinicians should assess for nosocomial and community risk factors, realizing that different populations with CO-MRSA SSTI may require separate antimicrobial strategies.