The present article describes our studies to know the usefulness of in situ hybridization (ISH) to identify various kinds of mold observed in tissue sections and ⁄ or cytological preparations from the lesions of patients with invasive fungal infection. To establish the precise procedure for ISH in formalin – fixed and paraffin-embedded sections, various pretreatments were attempted. The condition finally chosen is written here providing a favorable outcome regarding to both intensity and specificity of signals on outline of molds observed in the tissue sections when specimens were treated with both heat and proteinase K and, solutions were adjusted to higher pH value. Therefore, usefulness of promising probes, two each DNA and peptide nucleic acid (PNA) were verified with a favorable pretreatment condition, using lungs of mice experimentally infected and ⁄ or those obtained from autopsies with invasive mold infection. As the result, DNA probes targeting alkaline proteinase (ALP) gene and retrotransposon Afut – 1 gene of Aspergillus fumigatus showed specific signal intensity for the Aspergillus species and A. fumigatus, respectively. PNA probes for Candida albicans and the Fusarium species also showed satisfactory specificity. We wish to emphasize that ISH can be a valuable tool to identify medically important molds in formalin – fixed and paraffin – embedded tissue sections or cytological preparations.
Animal mycosis, particularly deep mycosis, is one of the most challenging conditions encountered by veterinarians. Pathogens causing mycotic infections in animals include fungi such as Cryptococcus neoformans, Candida spp., and Aspergillus spp. The antifungal drugs used for the treatment of deep mycoses in animals as well as humans are polyenes and azoles. However, the sensitivity of clinical isolates obtained from animals toward these drugs has rarely been assayed. In this study, the antifungal activities of itraconazole and voriconazole against clinical isolates of C. neoformans, Candida spp., and A. fumigatus isolated from animals with mycoses were examined using the broth microdilution method performed according to the guidelines provided by the Clinical and Laboratory Standards Institute. The minimum inhibitory concentrations (MICs) of itraconazole toward the C. neoformans, Candida spp., and A. fumigatus isolates were 0.125 – 1, 0.125 – 2, and 0.25 – 2μg⁄ml, respectively, and those of voriconazole were 0.0625 – 0.5, ≤0.0313 – 0.0625, and 0.0625 – 1μg⁄ml, respectively. The results of the MIC analyses implied that the fungal isolates obtained from infected animals exhibit an equivalent degree of susceptibility to itraconazole and voriconazole, as is observed in the case of isolates obtained from humans. The appropriate antifungal therapeutic strategy for the treatment of mycoses in animals must be selected taking into consideration the host immune status and organ function as well as the in vitro sensitivity of the pathogens to antifungal drugs.
Seven cases of cryptococcus meningitis in a tertiary care hospital from 2004 – 2007 were reviewed. 85.7% of the patients had headache as their predominant clinical feature. The spectrum of CT ⁄ MR findings ranged from no abnormality, basal ganglion lesion, to intracerebral and intraventricular granulomas. Findings of cerebrospinal fluid(CSF)cytology and biochemistry analysis were inconclusive. Patients were diagnosed by India ink(100%), CSF cryptococcal antigen detection(100%), and CSF culture in 6(85.7%). With the exception of two patients, co-morbidities associated were HIV, diabetes mellitus, and idiopathic CD4 + lymphocytopenia. Six patients were successfully treated with amphotericin B and discharged. A high index of clinical suspicion and laboratory diagnosis achieved early can reduce the overall morbidity and mortality among patients with cryptococcosis.