Cryptococcus gatti serotype B isolated in Sikkim (North-East India)—A new geographical niche

Abstract

Cryptococcus neoformans and Cryptococcus gatti both cause infection in immunocompromised patients. We report a case of meningitis with C. gatti in an AIDS patient. This case to our knowledge is the first case of C. gatti being reported from Sikkim (North East India).

1. Introduction

Cryptococcus gatti formerly known as Cryptococcus neoformans var. gatti has now emerged as a health threat in the western world with deadly outbreaks in British Columbia, Canada [1–3] and the Northwestern United states [4,5]. It is endemic in Australia and New Zealand [6] and sporadic cases have been reported from the Indian subcontinent. C. neoformans is a species complex comprising of C. neoformans var. grubii (serotype A) C. neoformans var. neoformans (serotype D) and C. gatti (serotype B and C) [7]. C. gatti differs from the most commonly isolated C. neoformans in clinical aspects, ecological niche and genetic makeup. Unlike C. neoformans it most commonly affects immunocompetent patients exposed to an environmental source causing crytococcomas and is known to be more resistant to antifungal agents [1]

Sikkim, a state of India is landlocked geographical region with the coordinates 27°20′N 88°37′E/27.33°N 88.62°E/27.3. It is situated in the Eastern Himalayas spread below the world's third highest mountain Kangchendzonga. It has a total area of 7096 sq km with total population of 0.54 million as per 2001 census. Annual rainfall varies between 2000 mm and 4000 mm and the temperature ranges from sub-zero during winters to 28 °C during summer [8]

2. Case

A 27-year-old male from, East Sikkim, was admitted in the casualty ward (day 0) of Sir Thotub Namgyal Memorial Hospital, Gangtok with complaints of severe headache, loss of appetite and weight loss. On examination he was conscious but disoriented. His headache worsened with episodes of vomiting. Broad spectrum antibiotics, antiemetics and analgesics were administered. Patient was positive for HIV 1 (day+1) screened by the rapid immunochromatographic method (Standard Diagnostic) and confirmed by following the National Aids Control Organisation testing guidelines. Anti-retroviral treatment with zidovudine 300 mg, lamivudine 150 mg, nevirapine 200 mg twice daily was initiated. His CD4 count was-33 cells/μl. Routine laboratory investigations revealed—Hb 7.4 g/dl, ESR 70 mm/1st hour (Westergreen), WBC 4890 cells/mm³, Neutrophil 3920 cells/mm³, Lymphocyte 330 cells/mm³, and Monocytes 200 cells/mm³. His liver function test and kidney function test parameters were within normal limit. He was negative for Hepatitis B (HBsAg) and C, both the tests done by the immunochromatographic method (Tulip diagnostics, India). Widal test (Tulip diagnostics, India) for enteric fever and rapid plasma reagin test (RPR—Tulip diagnostics, India) done for Syphilis was negative.

Magnetic Resonance Imaging of brain done on day+2 showed ischaemic changes in the left caudal head and basal ganglia. His condition stabilized on day+3. A fundus examination revealed a normal fundus and lumbar puncture was performed on day+4 and samples were sent for biochemical and microbiological investigation. CSF Examination showed WBC—15 cells/mm³ (predominantly lymphocytes). Protein: 59 mg/dl and Sugar: 33 mg/dl. Zeil Neelsen's stain showed no acid fast bacilli.

India ink preparation done from the un-centrifuged CSF sample showed round yeast cells with narrow based occasional budding having a clear halo around it, suggestive of Cryptococcus spp. under magnification 400×. Amphotericin B 0.7 mg/kg and fluconazole 400 mg was started when the microscopy report was available. Flucytosine could not be administered as it was not readily available in the market.

White, moist colonies were observed on Sabourauds dextrose agar with gentamicin after 4 days of incubation at 37 °C. Urease test and Inositol fermentation test were positive and it was inoculated on canavanine-glycine bromothymol blue (CGB) agar to differentiate C. neoformans from C. gatti. These tests subsequently confirmed phenotypically the isolate as C. gatti. Serotyping done by the latex agglutination test (Meridian diagnostics) found the isolate to be serotype B. The fungal spp. was confirmed as C. gatti based on the colony morphology on CGB agar, biochemical reaction and serotyping.

Patient complained of giddiness on the morning of day +8, his condition gradually deteriorated and he was declared dead at 5:40 pm on day +8.

3. Discussion

C. gatti has been reported from Northern India in 1993 where out of 18 clinical isolates 3 were C. gatti (serotype B) [9], there have been sporadic reports since then [10,11]. C. gatti Serotype B is known to be the most prevalent serotype in clinical and environmental samples unlike serotype C which is less commonly isolated [7]. In concordance with the other Indian studies our isolate was also serotype B.

The areas where C. gatti has been previously reported fall under the tropical zone unlike east Sikkim which has a prevalent temperate climate. With recent outbreaks in temperate regions like Canada and Northwestern United States it has been suggested that the pathogen might have adapted to a new climatic niche and may have a wider geographical distribution [7].

The introduction of antiretroviral therapy has improved the quality and length of life of HIV patients and decreased the incidence of opportunistic infections [12]. The HIV status of the patient was diagnosed late and HAART was initiated when the symptoms of AIDS had manifested. Since his CD 4 count was towards the lower end—33 cells/μl the patients cell mediated immunity may not have defended the body against the environmental pathogen.

Environmental exposure is known to be the dominant risk factor for C. gatti infection. The Eucalyptus family of trees has been constantly associated with C. gatti [13]; it has also been isolated from 20 species of 14 families of trees and from soil around it in various states of northern India and south India [14]. Sikkim has a diverse variety of flora and fauna with 2650 sq km of forest land under tree cover [8]. Although not a native tree of the Sikkim Himalayas, saplings of the variety E. grandis, E. citriodora, E. globosa, E. tetrocornis were imported from Australia and planted here as a reforestation drive.. Since samples from trees where the patient lived were not taken we can only say that the environment may have been the source for the yeast.

Cryptococcosis due to C. gatti is known to be prevalent among immunocompetent individuals and not only in the immunocompromised. C. gatti infection is underreported and the exact epidemiology in this region will remain unclear if physicians do not consider C. gatti as a possible pathogen while treating patients and the laboratories do not carry out the phenotypic confirmation of the species.

4. Conflict of Interest

There are none.