PCR-based ribotyping of Staphylococcus aureus

Abstract

Genotyping techniques are valuable tools for the epidemiologic study of Staphylococcus aureus infections in the hospital setting. Pulsedfield gel electrophoresis (PFGE) is the current method of choice for S. aureus strain typing. However, the method is laborious and requires expensive equipment. In the present study, we evaluated the natural polymorphism of the genomic 16S-23S rRNA region for genotyping purpose, by PCR-based ribotyping. Three primer pairs were tested to determine the size of amplicons produced and to obtain better discrimination with agar gel electrophoresis and ethidium bromide staining. The resolution of the typing system was determined using sets of bacteria obtained from clinical specimens from a large tertiary care hospital. These included DNA from three samples obtained from a bacteremic patient, six strains with known and diverse PGFE patterns, and 88 strains collected over a 3-month period in the same hospital. Amplification patterns obtained from samples from the same patient were identical, and PFGE from samples known to be different produced three genotypes. Amplification of DNA from 61 methicillinresistant isolates produced only one pattern. Methicillin-sensitive strains yielded a diversity of patterns, pointing to a true polyclonal distribution throughout the hospital (22 unique patterns from 27 strains). Computer-based software can be used to differentiate among identifiable strains, given the low number of bands and good characterization of PCR products. PCR-based ribotyping can be a useful technique for genotyping methicillin-sensitive S. aureus strains, but is of limited value for methicillin-resistant strains.

The occurrence and dissemination of methicillin and vancomycin-resistant Staphylococcus in samples from patients and health professionals of a university hospital in Recife, State of Pernambuco, Brazil

“…Pulsed-fi eld gel electrophoresis (PFGE) was used to analyze the presence of bacterial clones between patients and professionals, thus indicating the health professionals as a source outbreaks of nosocomial infections 7 . The ribotyping-PCR technique used here has the advantage of being easy to perform and less prone to variations, compared to other methods, because it characterizes a region that is essential for bacterial growth and therefore more stable [13][14][15] . However, this method cannot confi rm that health professionals are the source of transmission, as they are carriers of several ribotypes not found in patients.…”

“…The knowledge of the molecular epidemiology of the diseases caused by these bacteria may assist in developing more effi cient strategies for reducing the infection, as genetic relationships among the different clones can be inferred, the gene fl ow route can be detected and the spread of infection can be traced from the molecular profi les 9,13 . Appropriate systems for typing are needed to determine the genetic variability of the isolates, thus providing an effective epidemiological control.…”

“…The ribotyping-polymerase chain reaction (PCR) is often referenced and used because of its high taxonomic and epidemiological value. This technique is a valuable tool for identifying and differentiating isolates of the Staphylococcus genus [13][14][15] .…”

Staphylococcus aureus ampicillin-resistant from the odontological clinic environment

“…According to the phenotypic interpretation criterion by Etest ® assays such results suggest the inexistence of the mecA gene in chromosomal DNA of S. aureus isolated from odontological clinic environment. This gene codifies a protein (PPB2a) that confers resistance to synthetical penicillin (i.e., methicillin and oxacillin) 23 .…”