Genetic diversity, clonality and sexuality in Toxoplasma gondii

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Abstract

The majority of Toxoplasma gondii strains from a variety of human and animal sources have been grouped into three highly clonal but closely related lineages. The low occurrence of nucleotide differences among the three predominant lineages and their unusual dimorphic allelic composition suggest that they have arisen from a recent common ancestry. Less than 1% of the previously studied strains contain unique genotypes and high divergence of DNA sequence, and therefore are considered ‘exotic’ or ‘atypical’ strains. The seemingly low genetic diversity in T. gondii may have been underestimated because most parasite strains in previous studies were collected from human patients and domestic animals in North America and Europe. To investigate the genetic diversity of T. gondii, we analysed parasite strains isolated from remote geographical regions by multilocus microsatellite sequencing and phylogenetic analysis. The genetic diversity indices, the molecular analysis of microsatellite genotypes and the constructed phylogram considered together suggest that the global T. gondii population is highly diversified and not characteristic of a clonal organism. The most parsimonious hypothesis is that T. gondii presents a complex population structure with a mix of clonal and sexual propagation as a function of the environmental conditions. The comparison between domestic strains data on one hand and wild strains data on the other hand is in favour of more frequent sexual recombinations in wild environment even though Toxoplasma subpopulation in human and domestic animals is largely clonal.

Introduction

Toxoplasma gondii is a protozoan parasite that infects virtually all warm-blooded animal species worldwide. It has a two-host life cycle with felids as definitive hosts in which the sexual replication of the parasite occurs, while mammals and birds are intermediate hosts in which the asexual replication occurs. Even though there is no geographic boundary and host specificity, molecular genotyping studies have showed a fundamental clonal population structure with three clonal lineages, namely types I, II and III (Tibayrenc et al., 1991, Dardé et al., 1992, Sibley and Boothroyd, 1992, Howe and Sibley, 1995, Dardé, 1996, Ajzenberg et al., 2002a). Difference at DNA sequence level among the predominant clonal lineages is less than 2% (Grigg et al., 2001a). There are several explanations for the existence of clonal population structure in T. gondii. First, this parasite is able to transmit among intermediate hosts through carnivorism and scavenging, bypassing sexual recombination events in definitive host cats (Howe and Sibley, 1995, Su et al., 2003). Second, many macrogametes of the parasite remain unfertilised but are capable of forming oocysts in the small intestine of cats by parthenogenesis (Ferguson, 2002). Third, cats simultaneously infected with different strains of T. gondii are likely to be very rare events in nature, therefore there is a temporal barrier for recombination to occur. Though a largely clonal population structure predominates, genetic diversity does exist in T. gondii (Howe and Sibley, 1995, Dardé, 1996, Bossi et al., 1998, Dardé et al., 1998, Cole et al., 2000, Lehmann et al., 2000, Grigg et al., 2001b, Carme et al., 2002b, Ghosn et al., 2003, Miller et al., 2004). Moreover, recombination phenomena have been described as occasional events (Howe and Sibley, 1995). To better estimate the genetic diversity and sexual recombination of T. gondii, we analysed a substantial set of strains of wild and/or tropical origin, mainly from French Guiana, by multilocus microsatellite sequencing and phylogenetic analysis.

Section snippets

Toxoplasma and Hammondia strains

A total of 43 T. gondii strains were selected for this study (Table 1). Most strains were selected from a pool of 245 strains mainly collected by the BRC ToxoBS group and categorised based on length polymorphism of five microsatellite (MS) markers. Eighty-eight percent (216/245) of these strains belong to the three classical type I, II and III lineages, with a clear predominance of type II strains (188/245, 77%). For sequence analysis, 14 strains were arbitrarily selected to represent the three

Molecular analysis

All 43 T. gondii strains were successfully sequenced at the five MS loci and the sequences were submitted to GenBank (see Table 2 for accession numbers). Based on the polymorphisms of dinucleotide repeats in microsatellite regions and the single nucleotide polymorphisms (SNPs) in flanking regions, a number of total alleles were identified for each marker including four alleles at TUB2, 15 at W35, 10 at TgM-A, seven at B18 and 16 at B17 (Fig. 1 and Table 2).

The number of SNP alleles varies

Discussion

The clonal theory of T. gondii population structure, with three predominant lineages, is based on T. gondii samples collected mainly from the European or the North American domestic cycle either from clinical cases of human toxoplasmosis or from meat producing animals such as pigs, sheep and chicken (Dardé et al., 1992, Howe and Sibley, 1995, Ajzenberg et al., 2002a, Ajzenberg et al., 2002b). This collection may not reflect the true status of T. gondii in remote geographical areas or in

Acknowledgements

This work was supported by French Ministry of Research (support no. 02 g 0418 for the development of Biological Resource Centers). We wish to thank for strain providing the BRC ToxoBS group (French parasitologist network for Toxoplasma strain collection): J.M. Pinon, I. Villena (Reims, RMS), P. Thulliez (Paris Institut de Puériculture, IPP), P. Marty (Nice, LPN), M.H. Bessieres (Toulouse, TOU), J.P. Gangneux (Rennes, REN), L. Paris (Paris Pitié-Salpétrière, PSP), J. Berthonneau (Poitiers), A.

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    Nucleotide sequences reported in this paper are available in the GenBank™, EMBL and DDBJ databases under the accession numbers AY572648, AY572692, AY572736, AY572780, AY572562AY572604, AY572605AY572647, AY572649AY572691, AY572693AY572735, AY572737AY572779.

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