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Journal of Clinical Microbiology, November 2001, p. 3889-3894, Vol. 39, No. 11
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.11.3889-3894.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Origin of Cryptococcus neoformans var. neoformans Diploid Strains

Massimo Cogliati,1 Maria C. Esposto,1 David L. Clarke,2 Brian L. Wickes,2 and Maria A. Viviani1,*

Laboratorio di Micologia Medica, Istituto di Igiene e Medicina Preventiva, Università degli Studi, IRCCS Ospedale Maggiore, 20122 Milan, Italy,1 and Department of Microbiology, University of Texas Health Science Center at San Antonio, San Antonio, Texas 78229-39002

Received 8 May 2001/Returned for modification 18 June 2001/Accepted 12 August 2001

The basidiomycetous yeast Cryptococcus neoformans is an important human fungal pathogen. Two varieties, C. neoformans var. neoformans and C. neoformans var. gattii, have been identified. Both are heterothallic with two mating types, MATa and MATalpha . Some rare isolates are self-fertile and are considered occasional diploid or aneuploid strains. In the present study, 133 isolates, mostly from Italian patients, were investigated to detect the presence of diploid strains in the Igiene Università Milano culture collection. All of the diploid isolates were further investigated by different methods to elucidate their origins. Forty-nine diploid strains were identified by flow cytometry. PCR fingerprinting using the (GACA)4 primer showed that the diploid state was associated with two specific genotypes identified as VN3 and VN4. Determination of mating type on V8 juice medium confirmed that the majority of the strains were sterile. PCR and dot blotting using the two pheromone genes (MFa and MFalpha ) as probes identified 36 of the 49 diploid isolates as MATa/alpha . The results of pheromone gene sequencing showed that two allelic MFalpha genes exist and are distinct for serotypes A and D. In contrast, the MFa gene sequence was conserved in both serotype alleles. Amplification of serotype-specific STE20 alleles demonstrated that the diploid strains contained one mating locus inherited from a serotype A parent and one inherited from a serotype D parent. The present results suggest that diploid isolates may be common among the C. neoformans population and that in Italy and other European countries serotype A and D populations are not genetically isolated but are able to recombine by sexual reproduction.

* Corresponding author. Mailing address: Laboratorio di Micologia Medica, Istituto di Igiene e Medicina Preventiva, Università degli Studi di Milano, Via Sforza 35, 20122 Milan, Italy. Phone: 39 02 55188373. Fax: 39 02 55191561. E-mail: marianna.viviani{at}unimi.it.

Journal of Clinical Microbiology, November 2001, p. 3889-3894, Vol. 39, No. 11
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.11.3889-3894.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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