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Journal of Clinical Microbiology, April 2004, p. 1505-1510, Vol. 42, No. 4
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.4.1505-1510.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Genotyping and Coalescent Phylogenetic Analysis of Pneumocystis jiroveci from South Africa

Frans J. L. Robberts,1* Lynne D. Liebowitz,1 and Lynda J. Chalkley2

Department of Medical Microbiology, School for Basic and Applied Health Sciences, University of Stellenbosch, Tygerberg,1 Medical Research Council South Africa, Parow, South Africa2

Received 14 October 2003/ Returned for modification 1 December 2003/ Accepted 16 December 2003

Sequence analysis of Pneumocystis jiroveci internal transcribed spacer (ITS) regions has become an important epidemiological tool. The objectives of the present study were to investigate sequence variations in the ITS1-5.8S ribosomal DNA (rDNA)-ITS2 regions; determine the P. jiroveci genotypes present in Cape Town, South Africa; and resolve the lineage evolution of the types by use of the coalescent theory. ITS regions were amplified from samples collected from 19 patients. PCR products were cloned, and four to five clones were sequenced from each specimen. Statistical parsimony was applied for coalescence-based network genotype analysis. The most prevalent type was Eg (14 of 19 patients, 33 of 83 clones), followed by Gg (4 of 19 patients, 7 of 83 clones), Eu (3 of 19 patients, 5 of 83 clones), and Gh (2 of 19 patients, 2 of 83 clones). Four new combinations (Eo, Je, Ge, and No), 11 new ITS1 sequences, and 13 new ITS2 sequences were identified. A new ITS2 type was detected in three patients and was designated type u. Coinfection appeared to be common, with 15 of 19 patients harboring more than one type and with up to six types per specimen. The resultant parsimony network identified Eg as the most probable ancestral haplotype and supported the occurrence of recombinational events within the population studied. Although the 5.8S rDNA region revealed only 13 clones containing one to two nucleotide polymorphisms, it may assist in defining types. Coalescent theory proposed that Eg is an ancestral type from which microevolutionary subtypes radiate.

* Corresponding author. Mailing address: Department of Medical Microbiology, School for Basic and Applied Health Sciences, University of Stellenbosch, P.O. Box 19063, Tygerberg 7505, South Africa. Phone: 27 021 938 5228. Fax: 27 021 938 4005. E-mail: lourensr{at}sun.ac.za.

Journal of Clinical Microbiology, April 2004, p. 1505-1510, Vol. 42, No. 4
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.4.1505-1510.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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