Pulsed-Field Gel Electrophoresis Typing of Oxacillin-Resistant <i>Staphylococcus aureus</i> Isolates from the United States: Establishing a National Database

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ABSTRACT

Oxacillin-resistant Staphylococcus aureus (ORSA) is a virulent pathogen responsible for both health care-associated and community onset disease. We used Sma I-digested genomic DNA separated by pulsed-field gel electrophoresis (PFGE) to characterize 957 S. aureus isolates and establish a database of PFGE patterns. In addition to PFGE patterns of U.S. strains, the database contains patterns of representative epidemic-type strains from the United Kingdom, Canada, and Australia; previously described ORSA clonal-type isolates; 13 vancomycin-intermediate S. aureus (VISA) isolates, and two high-level vancomycin-resistant, vanA -positive strains (VRSA). Among the isolates from the United States, we identified eight lineages, designated as pulsed-field types (PFTs) USA100 through USA800, seven of which included both ORSA and oxacillin-susceptible S. aureus isolates. With the exception of the PFT pairs USA100 and USA800, and USA300 and USA500, each of the PFTs had a unique multilocus sequence type and spa type motif. The USA100 PFT, previously designated as the New York/Tokyo clone, was the most common PFT in the database, representing 44% of the ORSA isolates. USA100 isolates were typically multiresistant and included all but one of the U.S. VISA strains and both VRSA isolates. Multiresistant ORSA isolates from the USA200, -500, and -600 PFTs have PFGE patterns similar to those of previously described epidemic strains from Europe and Australia. The USA300 and -400 PFTs contained community isolates resistant only to β-lactam drugs and erythromycin. Noticeably absent from the U.S. database were isolates with the previously described Brazilian and EMRSA15 PFGE patterns. These data suggest that there are a limited number of ORSA genotypes present in the United States.

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The evolutionary history of methicillin-resistant Staphylococcus aureus (MRSA).

Mark C Enright, D. Ashley Robinson, Gaynor Randle … (2002)

Methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of hospital-acquired infections that are becoming increasingly difficult to combat because of emerging resistance to all current antibiotic classes. The evolutionary origins of MRSA are poorly understood, no rational nomenclature exists, and there is no consensus on the number of major MRSA clones or the relatedness of clones described from different countries. We resolve all of these issues and provide a more thorough and precise analysis of the evolution of MRSA clones than has previously been possible. Using multilocus sequence typing and an algorithm, BURST, we analyzed an international collection of 912 MRSA and methicillin-susceptible S. aureus (MSSA) isolates. We identified 11 major MRSA clones within five groups of related genotypes. The putative ancestral genotype of each group and the most parsimonious patterns of descent of isolates from each ancestor were inferred by using BURST, which, together with analysis of the methicillin resistance genes, established the likely evolutionary origins of each major MRSA clone, the genotype of the original MRSA clone and its MSSA progenitor, and the extent of acquisition and horizontal movement of the methicillin resistance genes. Major MRSA clones have arisen repeatedly from successful epidemic MSSA strains, and isolates with decreased susceptibility to vancomycin, the antibiotic of last resort, are arising from some of these major MRSA clones, highlighting a depressing progression of increasing drug resistance within a small number of ecologically successful S. aureus genotypes.

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Multilocus sequence typing for characterization of methicillin-resistant and methicillin-susceptible clones of Staphylococcus aureus.

M C Enright, Brian Spratt, Jeremy Day … (2000)

A multilocus sequence typing (MLST) scheme has been developed for Staphylococcus aureus. The sequences of internal fragments of seven housekeeping genes were obtained for 155 S. aureus isolates from patients with community-acquired and hospital-acquired invasive disease in the Oxford, United Kingdom, area. Fifty-three different allelic profiles were identified, and 17 of these were represented by at least two isolates. The MLST scheme was highly discriminatory and was validated by showing that pairs of isolates with the same allelic profile produced very similar SmaI restriction fragment patterns by pulsed-field gel electrophoresis. All 22 isolates with the most prevalent allelic profile were methicillin-resistant S. aureus (MRSA) isolates and had allelic profiles identical to that of a reference strain of the epidemic MRSA clone 16 (EMRSA-16). Four MRSA isolates that were identical in allelic profile to the other major epidemic MRSA clone prevalent in British hospitals (clone EMRSA-15) were also identified. The majority of isolates (81%) were methicillin-susceptible S. aureus (MSSA) isolates, and seven MSSA clones included five or more isolates. Three of the MSSA clones included at least five isolates from patients with community-acquired invasive disease and may represent virulent clones with an increased ability to cause disease in otherwise healthy individuals. The most prevalent MSSA clone (17 isolates) was very closely related to EMRSA-16, and the success of the latter clone at causing disease in hospitals may be due to its emergence from a virulent MSSA clone that was already a major cause of invasive disease in both the community and hospital settings. MLST provides an unambiguous method for assigning MRSA and MSSA isolates to known clones or assigning them as novel clones via the Internet.

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Association between Staphylococcus aureus strains carrying gene for Panton-Valentine leukocidin and highly lethal necrotising pneumonia in young immunocompetent patients.

Yves Gillet, Bertrand Issartel, Philippe Vanhems … (2002)

Between 1986 and 1998, eight cases of community-acquired pneumonia due to Staphylococcus aureus strains carrying the gene for the Panton-Valentine leukocidin (PVL) were recorded in France, six of which were fatal. We aimed to assess the clinical features of these eight cases, and those of other cases identified prospectively, and to compare them with the characteristics of patients with pneumonia caused by PVL-negative strains. We compared eight retrospective and eight prospective cases of PVL-positive S aureus pneumonia with 36 cases of PVL-negative S aureus pneumonia. For all patients, we recorded age, length of hospital stay, risk factors for infection, signs and symptoms, laboratory findings, antibiotic treatment, and serial radiological findings. Median age was 14.8 years (IQR 5.4-24.0) for the PVL-positive patients and 70.1 years (59.2-81.4) for the others (p=0.001). Influenza-like illness had occurred during the 2 days before admission in 12 of the 16 PVL-positive patients, but in only three of 33 PVL-negative patients (p<0.001). PVL-positive infections were more often marked by: temperature greater than 39 degrees C (p=0.01), heart rate above 140 beats per min (p=0.02), haemoptysis (p=0.005), onset of pleural effusion during hospital stay (p=0.004), and leucopenia (p=0.001). The survival rate 48 h after admission was 63% for the PVL-positive patients and 94% for PVL-negative individuals (p=0.007). Histopathological examination of lungs at necropsy from three cases of necrotising pneumonia associated with PVL-positive S aureus showed extensive necrotic ulcerations of the tracheal and bronchial mucosa and massive haemorrhagic necrosis of interalveolar septa. PVL-producing S aureus strains cause rapidly progressive, haemorrhagic, necrotising pneumonia, mainly in otherwise healthy children and young adults. The pneumonia is often preceded by influenza-like symptoms and has a high lethality rate.