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Molecular approaches to systematic problems in parasitic nematodes : ribosomol DNA variation within cystidicola spp. (Nematoda: Habronematoidea) and the superfamily dracunculoidea Miscampbell, Allyson E.


Morphological characters are traditionally used in nematode systematics, however, morphological convergence and marginal differences between close relatives can obscure species diversity and confound taxonomic studies. This thesis applies molecular approaches to systematic problems in two groups of parasitic nematodes where morphological data is ambiguous. Ribosomal DNA (rDNA) variable regions such as the first and second internal transcribed spacers (ITS-1 and ITS-2), and the D3 expansion loop of the large subunit have consistently distinguished nematode species and provided a limited basis for phylogenetic inference between close relatives. I assess rDNA variation within Cystidicola spp. (Nematoda; Habronematoidea) and the superfamily Dracunculoidea to examine species diversity in both groups, and phylogenetic relationships in the Dracunculoidea. Phenotypic variation in Cystidicola spp. suggests unresolved variation within the genus. Distinct life histories, host ranges, reproductive strategies, and adult and egg morphologies define the two recognized Cystidicola spp. Variable host specificity and egg morphology in Cystidicola farionis is difficult to interpret and could reflect genetic species-level variation. I sequenced four rDNA regions (ITS-1, ITS-2, 5.8S, D3) from Cystidicola spp. isolates from a total of seven host species and nine locations in Ontario (ONT), British Columbia (BC) and Finland (FIN). The ITS-1, 5.8S, and D3 regions displayed no inter or intraspecific variation. Two ITS-2 types were identified which differed at four nucleotide positions: the ITS-2 from C. farionis (BC) and C. stigmatura was identical and 365bp long; the ITS-2 from ONT and FIN C. farionis was identical and 368bp long. No relationship between egg morphology and genetic variation was apparent. ITS-2 differences between morphologically distinct C. farionis (ONT and FIN) and C. stigmatura were expected but comparison of this region among C. farionis isolates produced a surprising result. The ITS-2 distinguishes C. farionis (BC) from C. farionis (ONT and FIN) and suggests a closer relationship between C. farionis (BC) and C. stigmatura. Morphological resemblance among close relatives and a lack of phylogenetically informative characters in the superfamily Dracunculoidea reiterates this need for more precise taxonomic markers. I examined the D3 and ITS-2 regions from a total of nine dracunculoid species to distinguish cryptic species (e.g. Philonema spp.), place unidentified nematodes within the current classification system, and infer phylogenetic relationships within dracunculoid families (e.g. the Philometridae and Guyanemidae). I sequenced the D3 of two dracunculoid species, Philometroides huronensis and an unidentified nematode from Eopsetta exilis, adding these to an existing D3 data set of seven dracunculoids and sequenced the ITS-2 from all nine species. These regions varied in their ability to distinguish close relatives. The D3 region distinguishes Philonema agubernaculum and P. oncorhynchi but not Cystidicola spp. whereas the ITS-2 is identical in the former taxa and distinct in the latter. Both ITS-2 and D3 data supported previous suggestions that the family Philometridae may be artificial, and that members of the Guyanemidae are affiliated with some philometrids (e.g. Philonema spp.).

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