Molecular identification, virulence factors and antifungal susceptibility patterns of Candida parapsilosis complex species isolated from clinical samples.

AIMS: The aim of this study was to identify C. parapsilosis complex strains isolated from various clinical samples by sequence analysis and to investigate whether there are any differences between the species in terms of virulence factors and antifungal susceptibility. METHODS: The study included a total of 42 isolates identified as C. parapsilosis complex based on the color they formed in chromogenic medium, colony morphology, and microscopic appearance in Corn Meal-Tween 80 Agar and they were confirmed with API 20 C AUX. For the DNA sequence analysis of clinical isolates, V9G forward and LS reverse primers were used as well as internal transcribed spacers (ITS1 and ITS4). Biofilm formation, esterase, phospholipase, and protease activities were evaluated as virulence factors. Antifungal susceptibility was investigated via colorimetric microdilution method. RESULTS: A total of 75 non-C. albicans isolates were obtained from various clinical samples between 2016 and 2017 in a Turkish Tertiary Care Hospital. Of them, 42 were identified as members of the C. parapsilosis complex. Of the 42 strains, 41 were identified as C. parapsilosis sensu stricto (CpSS), while only one was identified as C. orthopsilosis. Of the CpSS strains, 31 (75.6%) were biofilm-positive, six (14.6%) were esterase-positive, nine (21.9%) were positive for phospholipase activity, and 31 (75.6%) were positive for protease formation, whereas all virulence factors of C. orthopsilosis strain were found to be negative. All CpSS strains were found susceptible to amphotericin B, echinocandins, and flucytosine. CONCLUSIONS: It has been concluded that the use of molecular methods to identify CpSS would not be effective in routine laboratory practices as it is the most commonly isolated species from the C. parapsilosis complex and there are no significant differences between species in terms of antifungal susceptibility.

Can serums be replaced by Mueller-Hinton agar in germ tube test?

BACKGROUND: The germ tube test (GTT) is inexpensive, easy, and well-defined test that differentiates Candida albicans (excluding Candida dubliniensis and Candida africana) from other species. The aim of this study was to evaluate various serums (i.e., human, rabbit, horse, and fetal bovine serum) used in the GTT and Mueller-Hinton agar (MHA). MATERIALS AND METHODS: Fifty species isolated from various clinical samples that were defined as C. albicans by both conventional and DNA sequence analysis methods were included in the study. One to two colonies of C. albicans were mixed into 0.5-1 ml of fetal bovine serum, horse serum, rabbit serum, and human serum. Serums and MHA were incubated at 37°C for GTT. They were removed from the incubator and evaluated after 30 min, 1 h, 2 h, and 3 h of incubation. The GTT was accepted to be positive only if germ tube was 1/2 the width and 3 times the length of the parent yeast cell and with no constriction at the point of origin. RESULTS: When the use of serums and MHA for GTT was statistically evaluated, according to the positive scoring, the best results were obtained with MHA and with rabbit, horse, and fetal bovine serum, respectively. The best definition over time statistically was the third hour. CONCLUSION: It is suggested that inexpensive MHA is a fast, appropriate, and reliable medium for the probable diagnosis of GTT and C. albicans; however, additional studies are still needed to define other Candida species.

Investigation of possible virulence factors in Candida strains isolated from blood cultures.

BACKGROUND: The Candida species, which are one of the most common causes of nosocomial bloodstream infections, present with high mortality and morbidity rates. This study aims to investigate the production of esterase, phospholipase, proteinase, and biofilm formation ability of the Candida strains isolated from the blood cultures. MATERIALS AND METHODS: Between June 2011 and July 2012, the Candida strains, which were isolated from blood cultures of a total of 50 patients, were studied. The esterase activity was analyzed in the Tween-80 agar, while phospholipase activity was studied in the egg yolk agar. The proteinase activity and biofilm formation were identified by using the petri dish method and microplate method, respectively. RESULTS: Of 50 specimens obtained from individual patients, 17 (34%) were identified as C. albicans, 14 (28%) as C. glabrata, 9 (18%) as C. parapsilosis, 5 (10%) as C. krusei, 4 (8%) as C. kefyr, and 1 (2%) as C. tropicalis. The rate of proteinase, phospholipase, and esterase positivity was higher in the C. albicans isolates. Biofilm formation was the highest in the C. parapsilosis strains. CONCLUSIONS: Higher rate of virulence factors in the most commonly isolated Candida species than other species indicates that these virulence factors play a crucial role in the pathogenesis.

Etiological agents of superficial mycoses in Kayseri, Turkey.

OBJECTIVE: In this study, we aimed to determine the prevalence of agents that cause superficial mycoses and clinical types of superficial mycoses in terms of age and gender in our region were aimed. METHODS: Five hundred samples of nails, skin and skin with hair taken from 476 children and adult patients pre-diagnosed with superficial mycoses were examined by direct microscopy and cultural methods between October 2009 and October 2010. RESULTS: Fungal elements were determined in 212 (42.4%) of the samples by using direct microscopy. Fungal growth was detected in 111 (22.2%) cultures of the same samples. It was found that the most common agents in superficial mycoses were Trichophyton rubrum (43.7%), Candida spp. (28%) and less often, Aspergillus spp., Malassezia spp., Saccharomyces spp., Rhodotorula spp., Trichosporon spp. and Trichophyton verrucosum. CONCLUSION: The significance of diagnosis by using direct microscopy and culturing together was again shown for the diagnosis of superficial fungal infections that follow a chronic course and affect the quality of life of patients. The most common agents in the superficial mycoses were T. rubrum. With this study, defining the aetiological agents of superficial mycoses in the Kayseri region is expected to contribute to the literature in terms of epidemiological data.

What is the role of the (1→3)-ß-D-glucan assay in the screening of patients undergoing autologous haematopoietic stem-cell transplantation?

The aim of this study is to determine the clinical contribution of (1→3)-ß-d-glucan (BDG) screening in the case of patients undergoing autologous haematopoietic stem-cell transplantation (HSCT). The records at our stem-cell transplantation centre were reviewed to identify the patients who underwent autologous HSCT between April 2009 and December 2010. Patients were classified as having proven invasive aspergillosis (IA), probable IA, or possible IA on the basis of the criteria established by the European Organization for Research and Treatment of Cancer and Mycoses Study Group (independent of the BDG results). During the study period, the patients were screened for BDG twice a week from transplant (day 0) until engraftment. Three patients were diagnosed with probable IA and five were diagnosed with possible IA. A total of 354 serum samples from 79 patients who met the study inclusion criteria were used for statistical analysis. At the cut-off value of 80 pg ml(-1) , the sensitivity was 27.2% [95% confidence interval (CI); 7.3-60.6]; specificity, 94.4% (95% CI; 91.3-96.5); positive predictive value, 6.2%; and negative predictive, 93.7%. The clinical contribution of the BDG assay as a screening test was relatively limited in this cohort of patients undergoing autologous HSCT.

The interaction between piperacillin/tazobactam and assays for Aspergillus galactomannan and 1,3-beta-D-glucan in patients without risk factors for invasive fungal infections.

BACKGROUND: The aim of this study was to investigate the interaction between intravenous piperacillin/tazobactam treatment and Aspergillus galactomannan antigen (GM) and 1,3-beta-D: -glucan (BDG) test results in patients without known risk factors for invasive fungal infections (IFI). PATIENTS AND METHODS: Patients without known risk factors for IFI and who were to receive piperacillin/tazobactam monotherapy were considered eligible for the study. Serum samples were obtained both before and after antibiotic infusion on the first, third, seventh and tenth days of a piperacillin/tazobactam treatment course and 4 days after the last dose. GM was determined by Platelia Aspergillus ELISA (Bio-Rad Laboratories) and BDG was assayed using the Fungitell kit (Associates of Cape Cod, East Falmouth, MA) according to manufacturers' specifications. RESULTS: A total of 135 serum samples were collected from 15 patients. When a cut-off level of >or=0.7 was used for GM positivity, there were no false positive results. When a cut-off level of >or=0.5 was used, six serum samples were positive. There were no statistically significant differences between the median GM indices or median BDG levels of the various sampling times. However, 24 of 135 serum samples were positive for BDG for a threshold of 80 pg/mL. After ruling out fungal infections and all known potential causes of false BDG positivity, environmental contamination remained a possible cause of BDG reactivity. CONCLUSION: No significant interaction was observed between piperacillin/tazobactam administration and Aspergillus GM and BDG assays. Positive results for these tests should be evaluated cautiously in patients at high risk for IFI receiving piperacillin/tazobactam.

Comparative study of in vitro methods to analyse the antifungal activity of propolis against yeasts isolated from patients with superficial mycoses.

AIM: To test a total of 15 strains belonging to four species of yeasts by different in vitro methods against propolis and itraconazole (ITC). METHODS AND RESULTS: Three methods were compared for susceptibility testing of yeast isolates to propolis: disc diffusion method, agar dilution method and National Committee for Clinical Laboratory Standards (NCCLS, M27A) broth microdilution method. ITC was selected as the antifungal agent for comparison study. Using the broth microdilution method, the geometric mean for MIC (microg ml(-1)) with regard to all isolates was < or =0.06 for propolis and < or =0.35 for ITC. The broth microdilution and the agar dilution methods were in good agreement (75%) for propolis against yeasts isolated from patients with superficial mycoses. Using the diffusion method, all strains showed a broad zone of inhibition at the first available reading time (24 or 48 h). An increase of MIC values was accompanied by a decrease of growth inhibition zone diameter. A favourable correlation was found between MIC and inhibition zone around the disc for propolis sample and the correlation coefficient was: r = -0.626 (P < 0.01). CONCLUSIONS: This study suggests the potential value of the agar dilution and disc diffusion method as a convenient alternative method for testing of yeasts to propolis. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated that propolis and ITC were very active against yeasts from patients with superficial mycoses. The other prominent finding in this study is that RPMI 1640 with L-glutamine was the available broth for the in vitro susceptibility testing of yeasts.

What defines the quality of patient care in tinea pedis?

OBJECTIVES: The aim of this study has been to evaluate patients with tinea pedis for their demographic data and attitudes affecting the treatment of disease, and to compare the in vitro activity of 10 antifungal agents and to relate them to their in vivo activity. METHODS: Patients with positive mycological examination were enrolled in the study, and a questionnaire comprised of 22 questions was administered. A mycological culture was carried out for each specimen. The antifungal susceptibility of the subcultured species was determined for griseofulvin, terbinafine, ciclopiroxolamine, fluconazole, ketoconazole, itraconazole, bifonazole, sulconazole, oxiconazole and miconazole with microdilution. RESULTS: Mycological cultures were carried out from 59 patients and there were 35 positive cultures (59.3%). The dermatophytes were Trichophyton rubrum (n = 25) and Trichophyton mentagrophytes (n = 3). The yeasts were Candida albicans (n = 7), Candida glabrata (n = 1) and Trichosporon (n = 2). In the minimum inhibitory concentration (MIC) study, the mean +/- standard error of the mean (SEM) MICs of the antifungals for T. rubrum were as follows: terbinafine 0.01 +/- 0.003, oxiconazole 0.16 +/- 0.05, sulkonazole 0.31 +/- 0.05, miconazole 0.45 +/- 0.15, itraconazole 0.74 +/- 0.01, ketokonazole 1.03 +/- 0.17, ciclopiroxolamine 1.30 +/- 0.12, bifonazole 1.94 +/- 0.51, griseofulvin 4.87 +/- 0.61, and fluconazole 17.91 +/- 3.67 microg/mL. CONCLUSION: Our study supports that azoles could be used as first-line treatment, as oxiconazole is very effective for both dermatophytes and C. albicans. Correlation between in vitro results and clinical outcomes of cases of dermatophytes is still to be established and interpretive breakpoints defined, in order to increase the quality of patient care in tinea pedis.

Comparison of in vitro activities of antifungal drugs and ethanolic extract of propolis against Trichophyton rubrum and T. mentagrophytes by using a microdilution assay.

The in vitro activities of propolis against 29 strains of dermatophytes were compared with those of terbinafine, itraconazole, ketoconazole, and fluconazole. Minimal inhibitory concentrations (MICs) were determined according to a National Committee for Clinical Laboratory Standards broth microdilution method. Among the systemic antifungals tested, terbinafine was the most potent. Propolis showed important antifungal activity and it merits further investigation as a potentially useful agent for the treatment of dermatophytosis.

Fatal peritonitis due to Trichoderma sp. in a patient undergoing continuous ambulatory peritoneal dialysis.

We describe a rare case of peritonitis caused by an unusual fungus, Trichoderma sp., in a patient on continuous ambulatory peritoneal dialysis. Management of the patient consisted of Tenckhoff catheter removal and antifungal chemotherapy, but the patient died.

Case report. Successfully treated subcutaneous infection with Sporothrix schenckii in Turkey.

Sporotrichosis is rare in Turkey. We report a 48-year-old man who had subcutaneous sporotrichosis caused by Sporothrix schenckii that was successfully treated with short-term itraconazole and potassium iodide. The isolate was susceptible to itraconazole also in vitro. Short-term itraconazole and potassium iodide should be agents of choice for treatment of subcutaneous sporotrichosis. However, treatment is controversial both in choice of agent used and in duration of therapy.

Comparison of Etest with the broth microdilution method in susceptibility testing of yeast isolates against four antifungals.

A comparative evaluation of the Etest and the broth microdilution methods for antifungal susceptibility testing of 102 clinical yeast isolates against amphotericin B, fluconazole, itraconazole, and ketoconazole was conducted. The agreements between the Etest and the broth microdilution methods were 93.1% for amphotericin B 85.2% for ketoconazole, 82.3% for itraconazole and 79.4% for fluconazole. These results suggest that the Etest approach to antifungal susceptibility testing may be a viable alternative to the NCCLS reference methods for testing yeasts, but that further evaluations are needed.

Case report. Fatal Aspergillus flavus pericarditis in a patient with acute myeloblastic leukaemia.

We report a case of Aspergillus flavus pericarditis treated with fluconazole for oral candidosis. The patient with acute myeloblastic leukaemia developed tachypnoea after antileukaemic chemotherapy. Pericardial effusion was seen in the echocardiogram. Aspergillus flavus was isolated from the pericardial fluid. The patient died from aspergillosis, before the antimycotic treatment could be changed to amphotericin B.