Subcutaneous zygomycosis in an immunocompetent patient caused by Basidiobolus ranarum.

Zygomycosis is a fungal infection in humans caused by orders Mucorales and Entomophthorales. The incidence of Mucorales causing mucormycosis is on a rise and is well documented, whereas Entomophthorales is rare. Among Entomophthorales, infections caused by Conidiobolus are more common than Basidiobolus Here we present a case of subcutaneous basidiobolomycosis in a female patient. The patient had hyperpigmentation in the thigh region for 6 months and serous discharge for 4 months. All initial findings suggested the inflammatory stage of morphea. Differential diagnoses of granuloma annulare, malignant melanoma and morphea were considered radiologically. A good suspicion of fungal aetiology by the dermatologist led to an appropriate diagnosis of subcutaneous basidiobolomycosis based on fungal culture and histopathological examination. Based on macroscopic and microscopic findings, the causative organism was confirmed to be Basidiobolus ranarum The patient was started on oral potassium iodide and itraconazole and showed a good prognosis.

In Vitro Production of Virulence Factors and Antifungal Susceptibility Pattern of Aspergillus Isolates from Clinical Samples in a Tertiary Care Center.

Objectives This study was aimed to investigate the association between virulence factors and antifungal susceptibility pattern among Aspergillus species. Materials and Methods This study was carried out in the Department of Microbiology, from May 2018 to June 2019. A total of 52 Aspergillus isolates obtained from various clinical samples were speciated based on microscopic identification by lacto phenol cotton blue (LPCB) mount and slide culture technique. The production virulence factors such as biofilm, lipase, phospholipase, amylase, and hemolysin were detected using standard phenotypic methods with Aspergillus niger ATCC (American Type Culture Collection) 6275 as the control strain. Antifungal susceptibility patterns of all Aspergillus isolates to amphotericin B, itraconazole, voriconazole, and posaconazole were evaluated in line with the Clinical Laboratory Standards Institute (CLSI) M38-A2 guidelines. Results The percentage of resistance was the highest in itraconazole (48.08%), followed by amphotericin B (28.85%) and voriconazole (9.62%). All amphotericin B-resistant isolates produced biofilm, itraconazole-resistant isolates exhibited phospholipase activity, and voriconazole-resistant isolates produced biofilm and demonstrated phospholipase and hemolytic activities. Regardless of the virulence factors produced, all isolates were susceptible to posaconazole. Conclusion Understanding the relationship between virulence factors and antifungal resistance aids in the development of new therapeutic approaches involving virulence mechanisms as potential targets for effective antifungal drug development.

Strain Typing of Trichosporon asahii Clinical Isolates by Random Amplification of Polymorphic DNA (RAPD) Analysis.

Objective The aim of this study was to identify and isolate Trichosporon asahii ( T. asahii ) from clinical samples and to assess the genetic relatedness of the most frequently isolated strains of T. asahii using random amplification of polymorphic DNA (RAPD) primers GAC-1 and M13. Methods All the clinical samples that grew Trichosporon species, identified and confirmed by polymerase chain reaction (PCR) using Trichosporon genus-specific primers, were considered for the study. Confirmation of the species T. asahii was carried out by T. asahii- specific PCR. Fingerprinting of the most frequently isolated T. asahii isolates was carried out by RAPD using random primers GAC-1 and M13. Results Among the 72 clinical isolates of Trichosporon sp. confirmed by Trichosporon -specific PCR, 65 were found to be T. asahii as identified by T. asahii- specific PCR. Fingerprinting of the 65 isolates confirmed as T. asahii using GAC-1 RAPD primer yielded 11 different patterns, whereas that of M13 primer produced only 5 patterns. The pattern I was found to be the most predominant type (29.2%) followed by pattern III (16.9%) by GAC-1 primer. Conclusions This study being the first of its kind in India on strain typing of T. asahii isolates by adopting RAPD analysis throws light on genetic diversity among the T. asahii isolates from clinical samples. Fingerprinting by RAPD primer GAC-1 identified more heterogeneity among the T. asahii isolates than M13.

Evaluation of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide method for assessing biofilm formation in vitro by Trichosporon spp.

BACKGROUND: Invasive infections due to Trichosporon spp. have increased recently and are frequently associated with indwelling medical devices. Such infections which are associated with biofilm formation do not respond to the routinely used antifungal agents and are often persistent, associated with high mortality rate. Various methods have been described by researchers to evaluate and quantify the biofilm formation. AIM: This study was conducted to compare two methods of biofilm production by Trichosporon sp, i.e., test tube method with crystal violet (CV) staining and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. MATERIALS AND METHODS: Seventy-two clinical isolates of Trichosporon spp. collected from various sources were considered for the study. The identity of all the isolates was genotypically confirmed by Trichosporon-specific polymerase chain reaction (PCR). The isolates were further speciated phenotypically using biochemical profile and growth characteristics which identified the isolates as Trichosporon asahii (64/72), Trichosporon asteroides (5/72), Trichosporon cutaneum (2/72), and Trichosporon mucoides (1/72). Biofilm production was then evaluated and compared by test tube-CV method and MTT assay. RESULTS: All the Trichosporon isolates produced biofilm by MTT assay, whereas only 42 (53.6%) of the isolates were detected to be biofilm producers by CV method. Furthermore, MTT assay could differentiate better between weak and moderate biofilm producers as compared to CV method. CONCLUSION: Hence, MTT assay is a reliable method for quantification of biofilm produced by Trichosporon spp. using 96-well microtiter plate.