RESUMO
Germline transformation of a parasitic nematode of mammals has proven to be an elusive goal. We report here the heritable germline transformation of Parastrongyloides trichosuri, a nematode parasite whose natural hosts are Australian possums of the genus Trichosurus. This parasite can undergo multiple free-living life cycles and these replicative cycles can be maintained indefinitely in the laboratory. Transformation was achieved by microinjection of DNA into the ovary syncytium of either free-living or parasitic adult females. By selecting for the transgenic progeny of successive free-living life cycles, it was possible to establish and maintain transgenic lines. All three transgenic lines tested were shown capable of establishing patent infections in possums and to transmit the functional transgene to their progeny. The transgene, driven by the Pt hsp-1 promoter, was constitutively expressed in intestinal cells at all stages of both parasitic and free-living life cycles, although gene silencing appears to occur in some transgenic progeny. This is the first report of heritable transgenesis in a parasitic nematode of a mammal and we discuss a variety of previously inaccessible experimental avenues that will now be possible with this powerful model system.
Assuntos
Técnicas de Transferência de Genes , Genes de Helmintos , Strongyloides/genética , Estrongiloidíase/parasitologia , Animais , Feminino , Expressão Gênica , Estágios do Ciclo de Vida , Masculino , Microinjeções/métodos , Strongyloides/crescimento & desenvolvimento , Transformação Genética , Transgenes , Trichosurus/parasitologiaRESUMO
BACKGROUND: The last 10 years have seen growing recognition of the significance of the genes encoding enzymes responsible for hepatic alcohol metabolism as protective factors in the development of alcoholism. METHODS: We have developed DNA sequencing assays for measuring genetic variation at the alcohol dehydrogenase 2 (ADH2), ADH3, and aldehyde dehydrogenase 2 (ALDH2) loci. These have been used to survey volunteer control subjects from three New Zealand ethnic groups (white, Asian, and Polynesian) and young male alcoholics recruited from white and New Zealand Maori patients in a local treatment program. RESULTS: The allele frequency values for whites and Asians obtained in our study closely match those obtained previously in other laboratories. Our data (the first for Polynesians) are 0.42 for ADH2*2, 0.78 for ADH3*1, and 0.00 for ALDH2*2. In the New Zealand Maori alcoholic patients, the ADH2*2 frequency is significantly lower (0.15; p < 0.01). The frequency of ADH3*1 is also lower in this group (0.60), but this value is not significant (0.05 < p < 0.06). CONCLUSIONS: In young male New Zealand Maori, the ADH2*2 allele is a protective factor against alcoholism even in the absence of ALDH2*2.