Intraclonal mating in Trypanosoma brucei is associated with out-crossing

Abstract

To examine whether mating can occur within as well as between clones of Trypanosoma brucei, we transformed three T. brucei subspecies stocks with heterologous genes conferring resistance to either hygromycin or Geneticin and carried out a series of inter-and intraclone matings in all possible double drug combinations. Double drug-resistant hybrids were recovered from three of the six out-crosses, but not from any of the three intraclone matings. However, further analysis of cloned progeny trypanosomes from one of the out-crosses using RFLP markers, molecular karyotyping and RAPD (random amplification of polymorphic DNA) produced unequivocal evidence that intraas well as interclone mating had occurred. The progeny of interclone mating were double drug-resistant and heterozygous at 9 of 13 loci examined. In contrast, the progeny of intraclone mating had no demonstrable input of genetic material from the hygromycin-resistant parent and were similar to the Geneticin-resistant parent for most markers, except for five loci which were heterozygous in the Geneticin-resistant parent but homozygous in these clones (aldolase, THTl glucose transporter, procyclin, tubulin and cDNA 23). In addition, PFGE showed considerable karyotypic rearrangements in these clones and loss of genetic material was evident from RAPD and VSG (variant surface glycoprotein) gene fingerprint analysis. We conclude that intraclone mating can occur in trypanosomes, but only during out-crossing, suggesting that meiosis and/or fusion are triggered by a diffusible factor.

Identification of the meiotic life cycle stage of Trypanosoma brucei in the tsetse fly

“…We infer that meiosis may be a normal part of the trypanosome developmental cycle in the fly, contrary to the traditional narrative that includes only mitotic divisions. In laboratory crosses, mating (and hence meiosis) is considered to occur rarely and only when two different trypanosome strains are present, although low frequencies of intraclonal mating have been detected in both single transmissions and cotransmissions of different strains (32)(33)(34). Even though we observed fluorescence in only a small proportion of trypanosomes at any one time, it remains possible that all SG trypanosomes undergo meiosis at a certain point during establishment of the SG infection.…”

Minisatellite marker analysis of Trypanosoma brucei : Reconciliation of clonal, panmictic, and epidemic population genetic structures

“…The relationship between these genotypes suggests that they could have originated by self-fertilization; for example, genotype M21, which was first detected in this area in 1959, is heterozygous for alleles at MS42 and 292 could have self-fertilized to produce genotype M4, which is homozygous at both loci, and genotype M5, which is homozygous at minisatellite locus 292 (the major genotype detected between 1988 and 1990). Self-fertilization has been described in T. brucei (15,16). It would appear that for the Busoga (Ugandan) focus at least, human infectivity is associated with one genotype and its predicted self-fertilization products, implying little or no recombination between the human infective and nonhuman infective stocks.…”

A novel GFP approach for the analysis of genetic exchange in trypanosomes allowing the in situ detection of mating events

“…For the experimental cross, flies were infected with a mixture of clones K11 (a GFP TetR transfectant derived from TH2) and T. b. brucei KP2N, a previously transfected derivative of GPAP\CI\82\KP2-I (CLONE 23) (Letch, 1984) carrying the NEO gene in the tubulin gene array (Gibson & Whittington, 1993). Drug-resistant clones of these two parental stocks, TH2 and KP2, were successfully mated in a previous cross (Gibson et al, 1997b …”

“…These factors, coupled with the relative inaccessibility of genetic exchange within the tsetse fly vector, have hampered direct observation of the process. After analysis of crosses using selectable drug resistance markers, hybrids were found only in trypanosome populations derived from the salivary glands, not midguts, suggesting that genetic exchange probably takes place in the salivary glands (Gibson & Bailey, 1994;Gibson et al, 1997b). Analysis of the inheritance of genetic markers in hybrid progeny suggests that a meiotic division occurs at some stage Sternberg et al, 1988Sternberg et al, , 1989Gibson, 1989;Turner et al, 1990;Gibson & Stevens, 1999), but triploid hybrids also occur with some frequency Wells et al, 1987;Gibson et al, 1992Gibson et al, , 1997bGibson & Bailey, 1994;Hope et al, 1999) and a haploid life cycle stage has not been found (Shapiro et al, 1984;Tait et al, 1989).…”

“…After analysis of crosses using selectable drug resistance markers, hybrids were found only in trypanosome populations derived from the salivary glands, not midguts, suggesting that genetic exchange probably takes place in the salivary glands (Gibson & Bailey, 1994;Gibson et al, 1997b). Analysis of the inheritance of genetic markers in hybrid progeny suggests that a meiotic division occurs at some stage Sternberg et al, 1988Sternberg et al, , 1989Gibson, 1989;Turner et al, 1990;Gibson & Stevens, 1999), but triploid hybrids also occur with some frequency Wells et al, 1987;Gibson et al, 1992Gibson et al, , 1997bGibson & Bailey, 1994;Hope et al, 1999) and a haploid life cycle stage has not been found (Shapiro et al, 1984;Tait et al, 1989). The observation that kinetoplast (mitochondrial) DNA is inherited from both parents in hybrid progeny supports the surprising suggestion that somehow the complex parental kinetoplast DNA networks swap DNA (Gibson & Garside, 1990;Turner et al, 1995;Gibson et al, 1997a).…”