Uma Banerjee

Molecular Epidemiology of Clinical Cryptococcus neoformans Strains from India

ABSTRACT Little is known about the molecular epidemiology of the human pathogenic fungus Cryptococcus neoformans in India, a country now in the midst of an epidemic of AIDS-related cryptococcosis. We studied 57 clinical isolates from several regions in India, of which 51 were C. neoformans var. grubii, 1 was C. neoformans var. neoformans, and 5 were C. neoformans var. gattii. This strain set included 18 additional sequential isolates from 14 patients. Strains were characterized phenotypically by measuring the polysaccharide capsule and by determining the MICs of standard antifungals. Molecular typing was performed by a PCR-based method using the minisatellite-specific core sequence (M13), by electrophoretic karyotyping, by restriction fragment length polymorphisms with the C. neoformans transposon 1 (TCN-1), and by URA5 DNA sequence analysis. Overall, Indian isolates were less heterogeneous than isolates from other regions and included a subset that clustered into one group based on URA5 DNA sequence analysis. In summary, our results demonstrate (i) differences in genetic diversity of C. neoformans isolates from India compared to isolates from other regions in the world; (ii) that DNA typing with the TCN-1 probe can adequately distinguish C. neoformans var. grubii strains; (iii) that TCN-1 sequences are absent in many C. neoformans var. gattii strains, supporting previous studies indicating that these strains have a limited geographical dispersal; and (iv) that human cryptococcal infection can be associated with microevolution of the infecting strain and by simultaneous coinfection with two distinct C. neoformans strains.

Epidemiology of candidemia in a tertiary care centre of north India: 5-year study.

Background:To determine the distribution of species of Candida and the risk factors associated with candidemia in Indian population for which we conducted a retrospective study for 5 years in a tertiary care centre of North India.Materials and Methods:Blood samples from 7,297 patients aged from 3 days to 85 years, suspected with candidemia, were collected and tested for Candida. The susceptibility patterns toward fluconazole for the year 2005 isolates were tested by micro-dilution assay as described in the CLSI (M27A-2 method).Results:Most of the episodes have been caused by species other than C. albicans. Non-albicans candidemia was 79%–80% in both female and male populations. The most frequent species isolated from 275 patients in 5 years (January 2001–December 2005) was C. tropicalis (35.3%), followed by C. albicans (21.5%), C. parapsilosis (20%), C. glabrata (17.5%), C. krusei (3.3%), C. haemulonii (1.5%), and C. guilliermondii (1%). C. parapsilosis was the predominant in the fifth year of the study (2004–2005). Dose-dependant susceptibility to fluconazole was observed in 5% (n = 3) of the strains. Antifungal resistance was found in 11.7% (n = 7), which includes only C. glabrata.Conclusion:These results were comparable to those derived from other regions of India. C. tropicalis has been reported as the predominant species involved in the cases of candidemia. But in 2005 it has moved toward C. parapsilosis. No true antifungal resistance is reported. Further epidemiological surveillance is needed.

Susceptibility Pattern and Molecular Type of Species-Specific Candida in Oropharyngeal Lesions of Indian Human Immunodeficiency Virus-Positive Patients

ABSTRACT A study of oropharyngeal candidiasis (OPC) in Indian human immunodeficiency virus (HIV)/AIDS patients was conducted over a period of 15 months. This study revealed that 75% of the HIV/AIDS patients had OPC. MIC testing revealed that 5% of the Candida isolates were fluconazole resistant. A correlation between CD4+-T-cell counts and development of OPC in HIV/AIDS patients was also observed. Molecular typing of C. albicans isolates showed that all were genetically unrelated.

Dual Infections with Pigmented and Albino Strains of Cryptococcus neoformans in Patients with or without Human Immunodeficiency Virus Infection in India

ABSTRACT Cryptococcus neoformans is an encapsulated yeast-like fungus of worldwide distribution. Melanin production is an important virulence factor of C. neoformans. We report the identification of distinct cryptococcal isolates with either pigmented or white colony phenotypes on l-dihydroxyphenylalanine agar plates in three patients who presented with meningitis to the All India Institute of Medical Sciences in India. Two of the patients were also infected with human immunodeficiency virus. Biochemical studies, India ink analysis, immunofluorescence with antibodies specific to capsular antigen, and serotyping confirmed that the melanotic and albino strains were C. neoformans serotypes A and D, respectively. Genotyping with M13 and [GACA]4 primers revealed that all the C. neoformans isolates were genetically different. The CNLAC1 gene associated with melanin production was identified in all the strains by PCR. Standard MIC testing revealed that the strains had similar susceptibilities to amphotericin B, but time-kill assays with the antifungal showed reduced susceptibility in melanin-producing strains. Infection studies with A/Jcr mice showed that the melanin-lacking yeast were less virulent than melanin-producing isolates. These findings indicate that these patients had dual infections with pigmented and albino strains of C. neoformans that were phenotypically and biologically different. Continued surveillance of primary isolates from patients with cryptococcosis by analyzing phenotypic differences and by molecular methods may reveal that mixed infections occur more commonly than is currently realized.

Primary Cutaneous Mucormycosis in an Immunocompetent Host : Report of a Case

Abstract.Cutaneous mucormycosis is an uncommon disease and it usually affects immunocompromised, diabetic, and trauma patients with contaminated wounds or patients with underlying malignancies. It is very rare to find this disease in immunocompetent, nondiabetic patients. We herein report a case of primary cutaneous mucormycosis in an immunocompetent and nondiabetic patient. Our patient was a 50-year-old veterinary doctor. He was diagnosed to have cutaneous mucormycosis of the anterior abdominal wall, and was treated with multiple debridements of the wound and intravenous amphotericin B therapy. He received a total of 1 000 mg of amphotericin B. A high index of clinical suspicion and early institution of therapy in the form of surgical debridements and antifungal drugs are required to achieve a successful outcome.

Serotype distribution of Cryptococcus neoformans in patients in a tertiary care center in India

The prevalence of specific serotypes of Cryptococcus neoformans in a given area bears on regional epidemiological patterns, the expected spectrum of clinical disease, and predicted response to therapy. In this retrospective study we analyzed the serotypes of 45 degrees C neoformans isolates from 36 North Indian patients with varied clinical presentations. The majority of the isolates were serotype A (87%), and surprisingly, a significant number were serotype B (five isolates, 11%), which caused infection in patients both positive and negative for HIV. One unusual isolate was not typable with factor sera. Study of serotype distribution in patients showed serotypes A and B to be present, respectively, in 92% and 8% of 36 patients. In one apparently immunocompetent patient two serotypes, A and B, were isolated simultaneously from two different sites, lung and scalp abscess. This is the first reported case in which an individual was infected with two serotypes at the same time. In one HIV-infected child serotype A was isolated from blood. Our results suggest that the distribution of serotypes in Indian clinical isolates is different than that found in other regions.

A two-step schedule for the treatment of actinomycotic mycetomas.

Actinomycotic mycetomas usually respond slowly to treatment with antibiotics. In an attempt to hasten clinical resolution, we used a 2-step regimen consisting of an intensive phase of therapy with penicillin, gentamycin and co-trimoxazole for 5-7 weeks, followed by maintenance therapy with amoxicillin and co-trimoxazole. Seven patients were treated, all of whom showed significant reduction in discharge and swelling after the intensive phase. Maintenance therapy was continued for 2-5 months after the lesions became completely inactive. Five patients completed maintenance therapy, which was given for 6-16 months (mean 10.7 months), and remained free of disease during a mean post-treatment follow-up period of 6.4 months. The other 2 patients also responded satisfactorily and continue to receive maintenance therapy. Side-effects necessitating a modification of the treatment schedule occurred in 2 patients but reversed on discontinuation of the drugs responsible. This treatment schedule produces a rapid clinical response during the initial, intensive phase and promotes compliance with the longer maintenance phase of treatment necessary to achieve a complete cure.

Current scenario of cryptococcosis and antifungal susceptibility pattern in India: a cause for reappraisal

This study analysed the spectrum, antifungal susceptibility pattern, clinical course and molecular epidemiology of cryptococcosis. Four hundred and thirty‐nine samples obtained from 378 meningitis patients were processed by standard procedures. Minimum inhibitory concentration (MIC) of fluconazole and amphotericin B for the isolates was tested by broth micro dilution and by E‐strip method. Molecular analysis by random amplified polymorphic DNA‐PCR of eight isolates was performed using M13 primer. Cryptococcosis was diagnosed in 35 patients [HIV‐1 seropositive (19) and apparently immunocompetent (16)]. Cryptococcus neoformans var. neoformans (serotype A and D) was the predominant isolate on phenotypic identification. Three C. neoformans var. gattii were isolated from HIV‐1 seropositive (2) and apparently immunocompetent (1) patients. MIC 90 for amphotericin B and fluconazole were 1 and 8 μg ml−1 respectively. On RAPD‐PCR, less diversity was seen among Indian isolates. AIDS remains the single most important risk factor for cryptococcosis. Rising MIC of the available induction and maintenance drugs is of grave concern. The DNA typing technique showed less diversity among Indian strains. Routine surveillance and application of molecular typing methods are crucial to know the baseline and existing pattern of cryptococcosis.

Cladosporiosis (Cerebral Phaeohyphomycosis) of brain — a case report

A case of cerebral cladosporiosis caused by Cladosporium trichoides (bantianum) now known as Xylohypha bantiana is described and illustrated. Predisposing debilitating diseases were not detectable. The Cladosporiosis diagnosis was based on visualisation of hyphal element in direct Grams stain, direct KOH preparate of pus from brain abscess and on repeated successful cultivation of Cladosporium trichoides from specimen and by histopathology. Following surgery and anti-fungal chemotherapy the patient was cured.

Differences in the cell wall architecture of melanin lacking and melanin producing Cryptococcus neoformans clinical isolates from India: an electron microscopic study

Cryptococcus neoformans e um importante fungo oportunista patogenico que causa infeccao no sistema nervoso central, e que pode levar o paciente a morte. Um dos principais fatores de virulencia do C. neoformans e a producao de melanina na parede celular. Utilizando microscopia eletronica de transmissao, nos estudamos as paredes celulares de tres pares de isolados obtidos de pacientes com dupla infeccao pelo fungo, onde um isolado melanizado e um albino foram isolados do liquor de cada paciente. A microscopia eletronica de transmissao revelou que as cepas albinas nao apresentavam a camada de melanina enquanto que uma camada de melanina estava associada com a parede celular de cepas melanoticas, constituindo aproximadamente 75% da area da parede celular. O tamanho da parede celular das celulas produtoras de melanina foi aproximadamente o dobro do tamanho da parede celular dos isolados albinos (p < 0,003). Neste estudo, a microscopia eletronica de transmissao revelou que as diferencas na estrutura dos isolados albinos sem melanina e dos isolados produtores de melanina estava associada a parede celular e a camada de melanina.