Molecular characterization and in vitro antifungal susceptibilities of 75 clinical Zygomycetes isolated in Delhi, India

S. Kathuria, C. H. Klaassen, P. Roy, M. J. Najafzadeh, G. S. de Hoog, J. F. Meis, H. S. Randhawa and A. Chowdhary


Zygomycosis is a highly aggressive disease generally characterised by rhino-orbital-cerebral and pulmonary manifestations, usually fatal in immunocompromised patients. Species identification is epidemiologically and clinically important because it impacts the choice of specific antifungal therapy. We report molecular characterization and in vitro antifungal susceptibility profiles of 75 clinical isolates of Zygomycetes determined by the CLSI broth microdilution method and Etest for posaconazole and amphotericin B. The isolates originated from 68 patients investigated in tertiary care hospitals of Delhi during 2004– 2011. Sixty (80%) of the isolates were from pulmonary, 7 (10%) cutaneous and 8 (11%) rhino-orbital-cerebral zygomycosis patients. Identification was done by the standard mycological procedures and DNA sequencing of ITS and D1/D2 regions of rDNA. Molecular characterization was done with amplified fragment length polymorphism (AFLP). The Zygomycetes investigated were: Rhizopus oryzae (n = 35), R. microsporus (n = 17), R. stolonifer (n = 2), R. azygosporus (n = 1), Syncephalastrum racemosum (n = 9), Apophysomyces elegans (n = 4), Lichtheimia corymbifera (n = 2), L. ramosa (n = 2), and Mucor circinelloides (n = 3). Phenotypic determination revealed misidentification in 7 (9%) isolates. AFLP classified isolates into 8 clades.
Amphotericin B showed the excellent activity (GM, 0.06 lg ml-1). Of the azoles, posaconazole exhibited the highest activity (GM, 0.4 lg ml- 1), followed by itraconazole (GM, 1.15 lg ml-1) and Isavuconazole (GM, 1.2 lg ml-1). Itraconazole and Isavuconazole exhibited activity in 46% and 36% of the isolates (MIC, £0.5 lg ml-1) respectively. Voriconazole (GM, 8 lg ml-1) was the least active azole. Notably, terbinafine was active against all the species except R. oryzae and R. stolonifer. Etest results of posaconazole revealed varied susceptibilities, MICs ranging <0.002 – 32 lg ml-1 with MIC50, 0.5 lg ml-1. The MICs by Etest and CLSI of posaconazole revealed no concordance whereas it was 85% for AMB. The commonest underlying condition in zygomycosis was diabetes mellitus (53%), followed by COPD (35%), malignancy (15%), HIV (4%) and trauma (4%). Treatment and outcome records were available for only 26 patients with fatalities in 11 cases (15%). Zygomycosis is an emerging mycoses in India and herein we present one of the largest series of antifungal susceptibility data of this important clinical entity. Also, the role of correct molecular identification is emphasized.

abstract No: 

    • ISHAM 18th (2012)