Sponsor:  Procter & Gamble

Project goals:  1) Determine if fluorescent in situ hybridization (FISH) could be used to determine the location of S. aureus ex vivo in clinical vaginal washes and tampons during menstruation. 2) Determine if there is S. aureus biofilm formation in the samples.

Background:  Staphylococcus aureus is found in association with skin, sebaceous glands and mucous membranes and is usually a benign component of the human microflora. Under some conditions, however, S. aureus growth in the human vaginal environment can result in toxic shock syndrome. The ability to rapidly identify S. aureus with fluorescent in situ hybridization (FISH) within in vitro and ex vivo samples is a powerful research tool for determining microbial spatial relationships. In this project, microscopic analyses were used to characterize the physical association of S. aureus with other bacteria, eukaryotic cells and tampon fibers.

Special equipment or techniques used or developed:  FISH oligonucleotide probes

Results:   One hundred and seventeen women were screened by vaginal swab and traditional culture methods for the presence of S. aureus. Of the 117 women screened, 5 subjects vaginally-colonized with S. aureus and 7 non-colonized subjects were entered into the second phase of the trial. Tampons and vaginal washings were collected from the subjects during days 2-4 of their menstrual cycle. The colonization status was reconfirmed by swabbing and standard culture methods. The tampons and vaginal washes were then screened via two previously published rRNA targeting FISH probes for S. aureus. All tampon samples, including the samples identified as negative by standard culture methods, contained S. aureus and most of the visualized bacteria were predominantly in a biofilm phenotype. Analysis of the 12 vaginal washes indicated all of the subjects had S. aureus present in the vaginal fluid. The visualized bacteria in these samples were associated with red blood cells; some were adherent to desquamated epithelial cells. Use of the FISH probes demonstrated identification of S. aureus on tampon fibers and in vaginal washes from women who were identified by standard culture methods as both colonized and not colonized with S. aureus.