Faculty Bibliography

Mutations in the staphylococcal virulence regulator gene agr frequently occur during Staphylococcus aureus infection. Whether agr-defective strains are fit for colonization, an important prerequisite for infection, is unknown. Screening by means of assays to detect delta-hemolysin activity and agr autoinducing peptide production indicated that 15 ( approximately 9%) of 160 healthy human subjects were colonized with an agr-defective strain or a mixture of agr-positive and -defective S. aureus strains. The presence of identical agr-defective strains in family members suggests that these strains are transmissible. Additionally, carriage of an agr-defective strain was associated with hospitalization, raising the possibility that such strains may be selected in a nosocomial setting

The accessory gene regulator (agr) of Staphylococcus aureus is a global regulator of the staphylococcal virulon, which includes secreted virulence factors and surface proteins. The agr locus is important for virulence in a variety of animal models of infection, and has been assumed by inference to have a major role in human infection. Although most human clinical S. aureus isolates are agr(+), there have been several reports of agr-defective mutants isolated from infected patients. Since it is well known that the agr locus is genetically labile in vitro, we have addressed the question of whether the reported agr-defective mutants were involved in the infection or could have arisen during post-isolation handling. We obtained a series of new staphylococcal isolates from local clinical infections and handled these with special care to avoid post-isolation mutations. Among these isolates, we found a number of strains with non-haemolytic phenotypes owing to mutations in the agr locus, and others with mutations elsewhere. We have also obtained isolates in which the population was continuously heterogeneous with respect to agr functionality, with agr(+) and agr(-) variants having otherwise indistinguishable chromosomal backgrounds. This finding suggested that the agr(-) variants arose by mutation during the course of the infection. Our results indicate that while most clinical isolates are haemolytic and agr(+), non-haemolytic and agr(-) strains are found in S. aureus infections, and that agr(+) and agr(-) variants may have a cooperative interaction in certain types of infections

The use of quinolones in the treatment of non-serious community-acquired methicillin-resistant Staphylococcus aureus is discussed. The new C8-modified quinolones may be suitable for such treatment but controlled trials should be carried out to ensure that the pharmacokinetics are such that there is little risk of resistance developing

PCR-based assays were used to evaluate agr locus nucleotide polymorphism for the identification of agr autoinducer receptor specificity groups within a population of Staphylococcus aureus isolates colonizing children and their guardians. All isolates could be assigned to one of three major agr groups that had similar prevalences, regardless of whether isolates were implicated in transmission of S. aureus within families. Among healthy carriers, agr groups I to III appear to be equally fit, which may reflect selection for the coexistence of S. aureus strains in a population

Subtyping methicillin- resistant Staphylococcus aureus (MRSA) isolates and tracking nosocomial infections have evolved from phenotypic to genotypic approaches; most laboratories now depend on pulsed-field gel electrophoresis (PFGE). We discuss the limitations of current image-based genotyping methods, including PFGE, and the advantages (including ease of entering data into a database) of using DNA sequence analysis to control MRSA infections in health-care facilities

Coagulase gene (coa) short sequence repeat region sequencing was used to measure relatedness among a collection of temporally and geographically diverse methicillin-resistant Staphylococcus aureus isolates. The results show that coa polymorphism is free of strong selective pressure and has a low index of variation that may be useful for long-term epidemiological investigations. coa typing is a useful addition to spa typing for analysis of S. aureus, including methicillin-resistant strains

The Mycobacterium tuberculosis strains H37Rv and H37Ra are the most commonly used controls for M. tuberculosis identification in the clinical and research laboratory setting. To reduce the likelihood of misidentification and possible cross-contamination with this laboratory neotype, it is important to be able to distinguish H37 from clinical isolates. To provide a reference for identifying H37, we used multiple molecular techniques to characterize H37 strains, including 18 of the most frequently used variants available through the American Type Culture Collection. Isolates were genotyped using gene probes to IS6110 and IS1085. In addition, we performed polymorphic GC-rich sequence typing (PGRS), spoligotyping, determination of variable number of tandem repeats (VNTR), and PCR amplification of the mtp40, msx4, and mpp8 polymorphic regions. Southern hybridization with IS6110 provided the most discrimination, differentiating the 18 H37 isolates into 10 discrete patterns made up of 9 H37Rv variants and 1 H37Ra variant. PGRS, IS1085, mpp8, and spoligotyping were not able to distinguish any H37 variants, while VNTR and msx4 discriminated two. Only IS6110 and spoligotyping could distinguish the H37 strain from clinical isolates. In summary, spoligotyping and IS6110 provide a rapid and accurate way to identify H37 contamination, though IS6110 can, in addition, classify many of the H37 variants that would otherwise require phenotypic segregation

Recent reports indicate that community-acquired methicillin-resistant Staphylococcus aureus (MRSA) infections are increasing and may now involve persons without risk factors predisposing for acquisition. To estimate the extent of community MRSA in New York City, the prevalence of S. aureus and MRSA nasal colonization in a well-patient population of 500 children and guardians was determined. The prevalence of S. aureus nasal carriage was 35% for children and 28% for guardians. One person with predisposing risk factors was colonized with an MRSA, which was identified as the predominant clone found in New York City hospitals. A high degree of methicillin-susceptible S. aureus strain diversity was noted, with no apparent selection for specific clonal types. Thus, MRSA colonization is not ubiquitous in persons without predisposing risk outside of the health care environment. Bacterial competition and a lack of strong selection may limit the community spread of MRSA and can account for its sporadic distribution

The population structure of methicillin-resistant Staphylococcus aureus (MRSA) is predominantly clonal, which may be related to the fitness of the genetic background of the methicillin-susceptible S. aureus (MSSA) into which the mecA chromosomal resistant determinant has inserted. To test this idea, we assessed whether the genotypes of New York MRSA are present in MSSA populations by using a combination of protein A gene sequence typing (spa typing) and pulsed-field gel electrophoresis (PFGE). Although about 16% of colonizing MSSA isolated from community subjects were related to MRSA, only one of the five predominant New York MRSA clonal types was found among the MSSA isolates. Similarly, among nosocomial MSSA, only four MRSA homologues were observed, two of which may have arisen through deletion of the mec element. Thus, MRSA clonal types represent a limited spectrum of the diversity seen in community and hospital S. aureus populations. The data are best explained by antibiotic selection pressure, as opposed to increased transmissibility or virulence, being responsible for the clonal dissemination of the resistance phenotype in MRSA genetic backgrounds, an in turn, the limited spread of these strains outside of the hospital environment