[Investigation of the correlation between biofilm forming ability of urinary Candida isolates with the use of urinary catheters and change of antifungal susceptibility in the presence of biofilm]

Abstract

Frequency of Candida species causing urinary tract infections is increasing, and this increase is outstanding in nosocomial urinary tract infections especially in intensive care units. The ability of biofilm formation that is contributed to the virulence of the yeast, plays a role in the pathogenesis of biomaterial-related infections and also constitutes a risk for treatment failure. The aims of this study were to compare biofilm forming abilities of Candida strains isolated from urine cultures of patients with and without urinary catheters, and to investigate the change of antifungal susceptibility in the presence of biofilm. A total of 50 Candida strains isolated from urine cultures of 25 patients with urinary catheters (10 C.tropicalis, 6 C.glabrata, 4 C.albicans, 4 C.parapsilosis, 1 C.krusei) and 25 without urinary catheters (8 C.tropicalis, 6 C.albicans, 4 C.krusei, 3 C.parapsilosis, 2 C.kefyr, 1 C.glabrata, 1 C.lusitaniae) were included in the study. Biofilm forming ability was tested by Congo red agar (CRA) and microplate XTT [2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] reduction methods. Fluconazole (FLU) and amphotericin B (AMP-B) susceptibilities of the isolates were determined by reference microdilution method recommended by Clinical and Laboratory Standards Institute for planktonic cells and by XTT reduction assay in case of biofilm presence. Biofilm formation was detected in 12 (24%) by CRA and 50 (100%) of the isolates by XTT reduction method. None of the C.albicans (n= 10) and C.tropicalis (n= 18) strains were detected as biofilm positive by CRA, however, these strains were strongly positive by XTT reduction method. No statistically significant correlation was detected between the presence of urinary catheter and biofilm forming ability of the isolate (p> 0.05). This might be caused by the advantage of biofilm forming strains in adhesion to bladder mucosa at the initial stages of infection. For all of the isolates in planktonic form, minimum inhibitory concentration (MIC) values were below the clinical resistance breakpoints or epidemiological cut-off values. When tested in presence of biofilms, all the isolates became resistant aganist FLU. Similarly, 100% inhibition, which is recommended in the standard method to determine AMP-B MIC, could not be obtained in any of the isolates with the highest dilution 8 µg/ml AMP-B tested. When evaluation was performed according to 80% inhibition, only 14 (28%) of the isolates had an AMP-B MIC below species-specific epidemiological cut-off values in the presence of biofilm. As a result, no correlation between urinary catheters and biofilm formation ability of Candida isolates were detected. XTT reduction method was considered as more reliable than CRA for investigating biofilm formation of Candida species. In addition, CRA failed to detect biofilm formation in frequently isolated species such as C.albicans and C.tropicalis. Fluconazole activity was lost, while AMP-B could not provide 100% inhibition in presence of biofilm for all isolates tested. Even if 80% inhibition was taken into account, AMP-B activity was still variable according to strain.