Whole-Genome Sequence for Methicillin-Resistant Staphylococcus aureus Strain ATCC BAA-1680.
ABSTRACT: We report here the whole-genome sequence of the USA300 strain of methicillin-resistant Staphylococcus aureus (MRSA), designated ATCC BAA-1680, and commonly referred to as community-associated MRSA (CA-MRSA). This clinical MRSA isolate is commercially available from the American Type Culture Collection (ATCC) and is widely utilized as a control strain for research applications and clinical diagnosis. The isolate was propagated in ATCC medium 18, tryptic soy agar, and has been utilized as a model S. aureus strain in several studies, including MRSA genetic analysis after irradiation with 470-nm blue light.
Project description:Staphylococcus aureus ATCC BAA-39 is the reference organism for a multidrug-resistant Staphylococcus aureus (MRSA) strain that was used to study drug-induced resistance reversion by an iodine-containing nanomolecular complex, FS-1. PacBio sequencing was performed on both the experimental and control strains, followed by genome assembly, variant calling, and DNA modification profiling.
Project description:Application of supplementary drugs which increase susceptibility of pathogenic bacteria to antibiotics is a promising yet unexplored approach to overcome the global problem of multidrug-resistant infections. The discovery of a new drug, an iodine-containing nano-molecular complex FS-1, which has proven to improve susceptibility to antibiotics in various pathogens, including MRSA strain Staphylococcus aureus ATCC BAA-39TM, allowed studying this phenomenon. Chromosomal DNA and total RNA samples extracted from the FS-1 treated strain (FS) and from the negative control (NC) cultures were sequenced by PacBio SMRT and Ion Torrent technologies, respectively. PacBio DNA reads were used to assemble chromosomal DNA of the NC and FS variants of S. aureus BAA-39 and to perform profiling of epigenetically modified nucleotides. Results of transcriptional profiling, variant calling and detection of epigenetic modifications in the FS variant were compared to the NC variant. Additionally, the genetic alterations caused by the treatment of S. aureus BAA-39 with FS-1 were compared to the results of a similar experiment conducted with another model organism, E. coli ATCC BAA-196. Several commonalities in responses of these phylogenetically distant microorganisms to the treatment with FS-1 were discovered, which included metabolic transition toward anaerobiosis and oxidative/osmotic stress response. S. aureus culture appeared to be more sensitive to FS-1 due to a higher penetrability of cells by iodine bound compounds, which caused carbonyl stress associated with nucleotide damaging by FS-1, abnormal epigenetic modifications and an increased rate of mutations. It was hypothesized that the disrupted pattern of adenine methylated loci within methicillin-resistance chromosome cassettes (SCCmec) may promote excision of this antibiotic resistance determinant from chromosomes while the altered pattern of cytosine methylation was behind the adaptive gene regulation in the culture FS. The selection against the antibiotic resistance in bacterial populations caused by abnormal epigenetic modifications exemplifies possible mechanisms of antibiotic resistance reversion induced by iodine-containing compounds. These finding will facilitate development of therapeutic agents against multidrug-resistant infections.
Project description:BACKGROUND: Community acquired (CA) methicillin-resistant Staphylococcus aureus (MRSA) increasingly causes disease worldwide. USA300 has emerged as the predominant clone causing superficial and invasive infections in children and adults in the USA. Epidemiological studies suggest that USA300 is more virulent than other CA-MRSA. The genetic determinants that render virulence and dominance to USA300 remain unclear. RESULTS: We sequenced the genomes of two pediatric USA300 isolates: one CA-MRSA and one CA-methicillin susceptible (MSSA), isolated at Texas Children's Hospital in Houston. DNA sequencing was performed by Sanger dideoxy whole genome shotgun (WGS) and 454 Life Sciences pyrosequencing strategies. The sequence of the USA300 MRSA strain was rigorously annotated. In USA300-MRSA 2658 chromosomal open reading frames were predicted and 3.1 and 27 kilobase (kb) plasmids were identified. USA300-MSSA contained a 20 kb plasmid with some homology to the 27 kb plasmid found in USA300-MRSA. Two regions found in US300-MRSA were absent in USA300-MSSA. One of these carried the arginine deiminase operon that appears to have been acquired from S. epidermidis. The USA300 sequence was aligned with other sequenced S. aureus genomes and regions unique to USA300 MRSA were identified. CONCLUSION: USA300-MRSA is highly similar to other MRSA strains based on whole genome alignments and gene content, indicating that the differences in pathogenesis are due to subtle changes rather than to large-scale acquisition of virulence factor genes. The USA300 Houston isolate differs from another sequenced USA300 strain isolate, derived from a patient in San Francisco, in plasmid content and a number of sequence polymorphisms. Such differences will provide new insights into the evolution of pathogens.
Project description:Widespread infections with community-associated (CA) methicillin-resistant Staphylococcus aureus (MRSA) have occurred in the United States with the dissemination of the USA300 strain beginning in 2000. We examined 105 isolates obtained from children treated at the University of Chicago from 1994 to 1997 (75 methicillin-susceptible S. aureus [MSSA] and 30 MRSA isolates) in order to investigate for possible evidence of USA300 during this period. Infections were defined epidemiologically based on medical record review. The isolates underwent multilocus sequence typing (MLST), as well as assays for the Panton-Valentine leukocidin (PVL) genes, the protein A gene (spa), and arcA and opp3, proxy markers for the arginine catabolic mobile element (ACME), characteristic of USA300 MRSA. MRSA isolates also underwent staphylococcal cassette chromosome mec (SCCmec) typing and pulsed-field gel electrophoresis (PFGE) subtyping. MSSA isolates belonged to 17 sequence type (ST) groups. The 12 epidemiologically defined CA-MRSA infection isolates were either ST1 (n = 4) or ST8 (n = 8). They belonged to 3 different PFGE types: USA100 (n = 1), USA400 (n = 5), and USA500 (n = 6). Among the CA-MRSA infection isolates, 8 (67%) were PVL(+). None of the MRSA or MSSA isolates contained arcA or opp3. Only one MRSA isolate was USA300 by PFGE. This was a health care-associated (HA) MRSA isolate, negative for PVL, that carried SCCmec type II. USA300 with its characteristic features was not identified in the collection from the years 1994 to 1997.
Project description:The cfr gene was identified in three linezolid-resistant USA300 methicillin-resistant Staphylococcus aureus (MRSA) isolates collected over a 3-day period at a New York City medical center in 2011 as part of a routine surveillance program. Each isolate possessed a plasmid containing a pSCFS3-like cfr gene environment. Transformation of the cfr-bearing plasmids into the S. aureus ATCC 29213 background recapitulated the expected Cfr antibiogram, including resistance to linezolid, tiamulin, clindamycin, and florfenicol and susceptibility to tedizolid.
Project description:Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) USA300 has spread rapidly across North America, and CA-MRSA is also increasing in Australia. However, the dominant Australian CA-MRSA strain, ST93-IV [2B] appears distantly related to USA300 despite strikingly similar clinical and epidemiological profiles. Here, we compared the virulence of a recent Australian ST93 isolate (JKD6159) to other MRSA, including USA300, and found that JKD6159 was the most virulent in a mouse skin infection model. We fully sequenced the genome of JKD6159 and confirmed that JKD6159 is a distinct clone with 7616 single nucleotide polymorphisms (SNPs) distinguishing this strain from all other S. aureus genomes. Despite its high virulence there were surprisingly few virulence determinants. However, genes encoding ?-hemolysin, Panton-Valentine leukocidin (PVL) and ?-type phenol soluble modulins were present. Genome comparisons revealed 32 additional CDS in JKD6159 but none appeared to encode new virulence factors, suggesting that this clone's enhanced pathogenicity could lie within subtler genome changes, such as SNPs within regulatory genes. To investigate the role of accessory genome elements in CA-MRSA epidemiology, we next sequenced three additional Australian non-ST93 CA-MRSA strains and compared them with JKD6159, 19 completed S. aureus genomes and 59 additional S. aureus genomes for which unassembled genome sequence data was publicly available (82 genomes in total). These comparisons showed that despite its distinctive genotype, JKD6159 and other CA-MRSA clones (including USA300) share a conserved repertoire of three notable accessory elements (SSCmecIV, PVL prophage, and pMW2). This study demonstrates that the genetically distinct ST93 CA-MRSA from Australia is highly virulent. Our comparisons of geographically and genetically diverse CA-MRSA genomes suggest that apparent convergent evolution in CA-MRSA may be better explained by the rapid dissemination of a highly conserved accessory genome from a common source.
Project description:Methicillin-resistant Staphylococcus aureus is increasingly responsible for staphylococcal infections in the community. A large percentage of the community-acquired methicillin-resistant (CA-MRSA) strains in the USA produce Panton-Valentine leukocidin (PVL), which is associated with severe infections. The virulence of the clinical CA-MRSA strain USA300 was compared to that of its isogenic pvl-deleted mutant, and it was shown that PVL contributes to lung and muscle tissue destruction, respectively, in murine necrotizing pneumonia and skin infection models. Mice infected with the USA300 strain developed a dominant anti-PVL response. The PVL subunits were therefore tested as vaccinogens against this isolate, and their vaccine efficacy correlated with both the route of vaccination and infection. These data suggest that PVL is a virulence factor in murine CA-MRSA infections.
Project description:BACKGROUND:The spread of USA300 methicillin-resistant Staphylococcus aureus (MRSA) across the United States resulted in an epidemic of infections. In Europe, only sporadic cases or small clusters of USA300 infections are described, and its prevalence in England is unknown. We conducted prospective surveillance for USA300 in the east of England. METHODS:We undertook a 12-month prospective observational cohort study of all individuals with MRSA isolated from community and hospital samples submitted to a microbiology laboratory. At least 1 MRSA isolate from each individual underwent whole-genome sequencing. USA300 was identified on the basis of sequence analysis, and phylogenetic comparisons were made between these and USA300 genomes from the United States. RESULTS:Between April 2012 and April 2013, we sequenced 2283 MRSA isolates (detected during carriage screening and in clinical samples) from 1465 individuals. USA300 was isolated from 24 cases (1.6%). Ten cases (42%) had skin and soft tissue infection, and 2 cases had invasive disease. Phylogenetic analyses identified multiple introductions and household transmission of USA300. CONCLUSIONS:Use of a diagnostic laboratory as a sentinel for surveillance has identified repeated introductions of USA300 in eastern England in 2012-2013, with evidence for limited transmission. Our results show how systematic surveillance could provide an early warning of strain emergence and dissemination.
Project description:UNLABELLED:Methicillin-resistant Staphylococcus aureus (MRSA) USA300 is a successful S. aureus clone in the United States and a common cause of skin and soft tissue infections (SSTIs). We performed whole-genome sequencing (WGS) of 146 USA300 MRSA isolates from SSTIs and colonization cultures obtained from an investigation conducted from 2008 to 2010 in Chicago and Los Angeles households that included an index case with an S. aureus SSTI. Identifying unique single nucleotide polymorphisms (SNPs) and analyzing whole-genome phylogeny, we characterized isolates to understand transmission dynamics, genetic relatedness, and microevolution of USA300 MRSA within the households. We also compared the 146 USA300 MRSA isolates from our study with the previously published genome sequences of the USA300 MRSA isolates from San Diego (n = 35) and New York City (n = 277). We found little genetic variation within the USA300 MRSA household isolates from Los Angeles (mean number of SNPs ± standard deviation, 17.6 ± 35; π nucleotide diversity, 3.1 × 10(-5)) or from Chicago (mean number of SNPs ± standard deviation, 12 ± 19; π nucleotide diversity, 3.1 × 10(-5)). The isolates within a household clustered into closely related monophyletic groups, suggesting the introduction into and transmission within each household of a single common USA300 ancestral strain. From a Bayesian evolutionary reconstruction, we inferred that USA300 persisted within households for 2.33 to 8.35 years prior to sampling. We also noted that fluoroquinolone-resistant USA300 clones emerged around 1995 and were more widespread in Los Angeles and New York City than in Chicago. Our findings strongly suggest that unique USA300 MRSA isolates are transmitted within households that contain an individual with an SSTI. Decolonization of household members may be a critical component of prevention programs to control USA300 MRSA spread in the United States. IMPORTANCE:USA300, a virulent and easily transmissible strain of methicillin-resistant Staphylococcus aureus (MRSA), is the predominant community-associated MRSA clone in the United States. It most commonly causes skin infections but also causes necrotizing pneumonia and endocarditis. Strategies to limit the spread of MRSA in the community can only be effective if we understand the most common sources of transmission and the microevolutionary processes that provide a fitness advantage to MRSA. We performed a whole-genome sequence comparison of 146 USA300 MRSA isolates from Chicago and Los Angeles. We show that households represent a frequent site of transmission and a long-term reservoir of USA300 strains; individuals within households transmit the same USA300 strain among themselves. Our study also reveals that a large proportion of the USA300 isolates sequenced are resistant to fluoroquinolone antibiotics. The significance of this study is that if households serve as long-term reservoirs of USA300, household MRSA eradication programs may result in a uniquely effective control method.
Project description:Unlike USA300, a strain of community-acquired methicillin-resistant Staphylococcus aureus (MRSA), commensal Staphylococcus aureus (S. aureus) bacteria isolated from human skin demonstrated the ability to mediate the glycerol fermentation to produce short-chain fatty acids (SCFAs). Quantitative proteomic analysis of enzymes involved in glycerol fermentation demonstrated that the expression levels of six enzymes, including glycerol-3-phosphate dehydrogenase (GPDH) and phosphoglycerate mutase (PGM), in commensal S. aureus are more than three-fold higher than those in USA300. Western blotting validated the low expression levels of GPDH in USA300, MRSA252 (a strain of hospital-acquired MRSA), and invasive methicillin-susceptible S. aureus (MSSA). In the presence of glycerol, commensal S. aureus effectively suppressed the growth of USA300 in vitro and in vivo. Active immunization of mice with lysates or recombinant α-hemolysin of commensal S. aureus or passive immunization with neutralizing sera provided immune protection against the skin infection of USA300. Our data illustrate for the first time that commensal S. aureus elicits both innate and adaptive immunity via glycerol fermentation and systemic antibody production, respectively, to fight off the skin infection of pathogenic MRSA.