Yasemin Oz

Diagnostic methods for fungal infections in pediatric patients: microbiological, serological and molecular methods

Although invasive fungal infections (IFIs) are relatively rare, they are important causes of morbidity and mortality in immunocompromised pediatric patients. Early and precise diagnosis of IFI is important to allow antifungal treatment to be started in time and to reduce the unnecessary use of toxic antifungal agents. Although traditional approaches such as direct microscopic examination, histopathological evaluation and cultivation are still gold standard, the diagnosis of IFI is generally difficult because of inadequate sensitivity and specificity with these tests. Commercial systems detecting the Aspergillus cell wall antigen galactomannan and 1,3-β-D-glucan are seen as the most convenient nonculture methods for the diagnosis of the IFI and monitoring of antifungal treatment. Several molecular methods have been described for the diagnosis of opportunistic mycoses. However, they have not been standardized and have only been used in experimental studies.

Correlation between broth microdilution and disk diffusion methods for antifungal susceptibility testing of caspofungin, voriconazole, amphotericin B, itraconazole and fluconazole against Candida glabrata

Candida glabrata is one of the most frequent organisms isolated from superficial and invasive fungal infections, after Candida albicans. This organism also exhibits intrinsically low susceptibility to azole antifungals and treatment often fails. The microdilution method is not very practical for use in routine susceptibility testing in the clinical laboratory, thus necessitating the use of other methods. In this study, we compared the in vitro activity of five antifungal agents in three different groups (echinocandin, polyene and azole) against 50 C. glabrata isolates by broth microdilution and disk diffusion methods recommended by Clinical Laboratory Standards Institute CLSI M27-A3 and CLSI M44-A, respectively. All the isolates were susceptible to amphotericin B (100%) and 98% of the isolates were susceptible to caspofungin by the broth microdilution method. Within the azole group drugs, voriconazole was the most active followed by fluconazole and itraconazole in vitro. The highest rate of resistance was obtained against itraconazole with a high number of isolates defined as susceptible-dose dependent or resistant. Although the disk diffusion method is easy to use in clinical laboratories, it shows very poor agreement with the reference method for fluconazole and itraconazole against C. glabrata (8% and 14%, respectively).

Synergistic Activities of Three Triazoles with Caspofungin against Candida glabrata Isolates Determined by Time-Kill, Etest, and Disk Diffusion Methods

ABSTRACT Combinations of voriconazole, fluconazole, and itraconazole with caspofungin were evaluated against 50 Candida glabrata isolates by the time-kill, disk diffusion, and Etest methods. The majority of antifungal combinations were indifferent. By the time-kill method, synergistic activity was detected with eight (16%) of the caspofungin-voriconazole and seven (14%) of the caspofungin-fluconazole combinations, but synergy was not seen with the caspofungin-itraconazole combination. Further comparisons of the Etest and disk diffusion synergy techniques with the time-kill method are warranted.

Antifungal susceptibility of ocular fungal pathogens recovered from around the world against itraconazole, voriconazole, amphotericin B, and caspofungin.

Although fungal infections of the eye are rare, they create an intractable clinical problem in ophthalmology because of the limited number of intravitreal and systemic therapeutic options. In this investigation, the in vitro efficacies of itraconazole (ITR), voriconazole (VOR), amphotericin B (AMB), and caspofungin (CAS) against 29 globally-collected ocular fungal isolates were assessed, following the standards that are outlined in the Clinical and Laboratory Standards Institute (CLSI) M38-A2 document. AMB [Geometric Mean (GM) MIC (Minimum Inhibitory Concentration): 0.49 μg/ml] was the most active drug, followed by VOR, CAS, and ITR (GM MICs: 0.52, 1.07, and 2.86 μg/ml, respectively). For the Exophiala strains (n = 8), VOR was the most active drug, followed by AMB, ITR, and CAS (GM MICS: 0.21, 0.27, and 1.09 μg/ml, respectively). ITR had no activity against Fusarium spp. (n = 9; GM MIC: 32 μg/ml), but AMB was found to be the most effective antifungal against the tested members of this genus, followed by CAS and VOR (GM MICs: 0.86, 1.59, and 2.72 μg/ml, respectively). These data should be used to design future targeted clinical efficacy trials. We also report on several fungal species that are rarely encountered in the clinical laboratory, for which little information about drug sensitivities was previously available.

Species distribution and in vitro antifungal susceptibility of clinical Candida isolates from a university hospital in Turkey over a 5-year period

We retrospectively evaluated the distribution of clinical Candida spp. isolated over a 5-year period in our hospital relative to year, specimen types, hospital departments and their antifungal susceptibility patterns. Overall 3,756 Candida spp. were recovered from 10,857 specimens. In vitro antifungal susceptibility tests were conducted with 2,068 isolates against amphotericin B, fluconazole and itraconazole using the Etest method. C. albicans was isolated frequently from non-sterile body specimens while non-C. albicans Candida spp. were commonly recovered from sterile body specimens. Isolation rates of C. albicans were 83%, 61.2% and 49% in non-sterile body specimens, sterile body specimens and blood-sterile body fluids, respectively. C. krusei was an important isolate from specimens of patients in the Haematology and Bone Marrow Transplantation units and its rate of recovery increased in these departments. Amphotericin B resistance was detected in only seven C. krusei isolates, whereas 80% (n = 1,653), 76% (n = 1,572) and 99% (n = 2,061) of all isolates were susceptible to fluconazole, itraconazole and amphotericin B, respectively. In conclusion, the distribution of Candida species was variable among hospital departments and among body sites. These results may be useful in predicting potential fungal pathogens and the choice of antifungal treatment.

Prevalence and epidemiology of tinea pedis and toenail onychomycosis and antifungal susceptibility of the causative agents in patients with type 2 diabetes in Turkey

Diabetes patients are particularly susceptible to fungal infections because their vascular and immunological systems are compromised.

The effect of clinical characteristics on the performance of galactomannan and PCR for the diagnosis of invasive aspergillosis in febrile neutropenic patients

Rapid diagnosis and early treatment of invasive aspergillosis is crucial for the management of the patients with haematological malignancy. We evaluated 358 sera from 78 febrile neutropenic episodes in patient with invasive aspergillosis (IA) (one proven, 17 probable, and 60 possible) and 83 episodes in patients with no IA according to the EORTC/MSG criteria. Patients specimens were tested by Mycassay Aspergillus PCR (first commercial real‐time PCR test) and in house real‐time PCR to investigate the presence of Aspergillus DNA, and by ELISA for detect the galactomannan (GM) antigen. We systematically investigated the medical background that can be effective on the test results. The hospitalisation period was longer in proven/probable episodes when compared with no IA (P = 0.001) and possible episodes. With regard to duration of neutropenia, the differences between both proven/probable with no IA (P = 0.023) and possible with no IA (P = 0.002) were highly significant. Similarly, the rates of T cell suppressant therapy in group proven/probable and possible episodes were significantly higher than in no IA (P = 0.005). There are significant differences in the performance of GM and PCR‐based tests among studies, and standardisation is required. Therefore, it can be useful to determine the effective factors on these tests. The use of larger volume of sera improved the performance of real‐time PCR for detection of Aspergillus DNA in high‐risk adult patients in the present study. Some host factors such as duration of neutropenia and administration of T cell suppressants related to the development of IA.

The prevalence of tinea pedis and tinea manuum in adults in rural areas in Turkey.

The aim of this study was to determine the frequency of tinea pedis and manuum (dermatophyte infections of the hands and feet) in adults in rural areas of Turkey, the risk factors and self-administered treatment options. A total of 2,574 people living in a rural area were enrolled in the study. Participants were asked demographic data, hygienic habits in a questionnaire. KOH preparations and culture were performed from suspicious lesions. Medical and alternative therapy methods and former dermatophytosis diagnosis history were taken from the respondents with suspicious lesions. Microbiological samples were taken from 285 (11.1%) participants. Culture was positive in 109 (4.2%) of those. The most common agent was Trichophyton rubrum. The predisposing factors were found as age older than 40, male gender and obesity. Forty-nine (44.9%) of patients had taken a medical therapy, 56 (51.4%) had performed non-medical methods (cologne, Lawsonia inermis-Henna and softener creams). Patients education about the treatment compliance is important.

Potential of polymerase chain reaction and galactomannan for the diagnosis of invasive aspergillosis in patients with febrile neutropenia.

The incidence of invasive aspergillosis (IA) has increased over the last years, especially in immuncompromised patients with high mortality rates. Because of difficulties about the diagnosis; serological methods [galactomannan (GM) antigen test] and polymerase chain reaction (PCR) developed in recent years. MycAssay Aspergillus PCR performance in the diagnosis of IA was evaluated and compared with the GM and in‐house PCR. This study was conducted with 358 serum samples obtained from 99 patient with febrile neutropenic episodes who were followed in haematology and bone marrow transplantation units. Patients were classified by the European Organization for the Research and Treatment of Cancer/Mycoses Study Group criteria, 18 of them is proven and probable IA. GM antigen test and two different real‐time PCR; one of them is fist commercial PCR for IA; Mycassay Aspergillus and the other one is in‐house real‐time PCR performed. Sensitivity values were Mycassay Aspergillus PCR, in‐house PCR, and GM 65.38%, 11.53% and 23.07%, respectively. The high sensitivity obtained from Mycassay Aspergillus PCR and sensitivity is increased by using a combination of diagnostic methods. GM antigen test and real‐time PCR could be beneficial for early diagnosis and treatment of IA. For routine usage of PCR as diagnostic assay more studies needed in future.

Postantifungal effect of the combination of caspofungin with voriconazole and amphotericin B against clinical Candida krusei isolates

We evaluated the postantifungal effects (PAFEs) of caspofungin (CAS), voriconazole (VOR), amphotericin B (AmB), and the combinations of CAS + VOR and CAS + AmB against 30 clinical Candida krusei isolates at 0.25, 1 and 4 times the MIC of each individually and in the indicated combinations. Antifungals were removed after 1 hour and colony counts were performed at 0, 2, 6, 24, and 48 h. VOR did not display any measurable PAFE regardless of antifungal concentrations, while AmB and CAS exhibited dose-dependent PAFE. The most effective agent producing a prolonged PAFE in this study was CAS. Although the combination of CAS with VOR generated longer PAFEs at 0.25 and 1 times their respective MICs in comparison with CAS alone, this combination was indifferent rather than synergistic. However, the combination of CAS with AmB at 4 times their MICs exhibited the best performance, reducing the colony counts during the 48 h after removal of drugs and resulted in synergic interaction in respect to 20 (67%) isolates. Consequently, CAS has a prolonged PAFE in vitro against C. krusei isolates, and the combination of AmB + CAS may increase significantly the efficacy of CAS. Our data may be useful in optimizing dosing regimens for these agents and their combinations, although further studies are needed to explore the clinical usefulness of our results.