FREQUENCY-DISTRIBUTION OF CANDIDA-ALBICANS BLASTOSPORES ADHERED TO MUCOSAL EPITHELIAL-CELLS IN-VITRO

Although several methods are available for examination of microbial ad herence to epithelial cells, these do not distinguish between adherenc e of viable and non-viable micro-organisms. This study reports the use of acridine orange-stained blastospores of Candida albicans in conjun ction with direct epifluorescence microscopy to determine viable (oran ge-fluorescing) and non-viable (green-fluorescing) blastospore adheren ce to buccal epithelial cells. The method was also employed to examine the effects of chlorhexidine treatment at sub-minimum inhibitory conc entrations on the adherence of viable and non-viable blastospores. The re was good correlation in the assessment of blastospore viability bet ween the direct epifluorescence microscopy technique and the standard serial dilution and plating method for viable counting, confirming the reliability of direct epifluorescence microscopy. Chlorhexidine treat ment before acridine orange staining did not alter this assessment of viability. Blastospore adherence to buccal epithelial cells resulted i n a similarly skewed distribution whether examined using a crystal vio let stain in conjunction with light microscopy or using direct epifluo rescence microscopy, therefore validating the direct epifluorescence m icroscopy technique for the enumeration of blastospore adherence. Chlo rhexidine treatment (0.0005% v/v, 30 min) of either blastospores or bu ccal epithelial cells altered the distribution of adherent blastospore s per epithelial cell by increasing the number of epithelial cells hav ing no adherent blastospores. No differences in adherence were, howeve r, observed between blastospore or epithelial cells after treatment wi th this agent. Examination of the adherence of viable and non-viable b lastospores to buccal epithelial cells using direct epifluorescence mi croscopy revealed a greater adherence capacity of non-viable than viab le blastospores for buccal epithelial cells. Treatment of blastospores with chlorhexidine altered the frequency distributions of viable and non-viable blastospores with lower numbers of blastospores adherent pe r epithelial cell. The larger reduction in adherent viable blastospore s in comparison with their non-viable counterparts is, however, an imp ortant observation which might have clinical relevance. Microbial cell s adhere to epithelial cells resulting in a skewed distribution; study of this distribution gives useful information about the adherence pro cess. Viable and non-viable components of a microbial population have different adherence capabilities and treatment of such populations wit h an antimicrobial agent exerting anti-adherent activity at sub-minimu m inhibitory concentrations reduces the amount of adherence of these v iable/non-viable components to different extents.