SSCP-based identification of members within the Pseudoterranova decipiens complex (Nematoda: Ascaridoidea: Anisakidae) using genetic markers in the internal transcribed spacers of ribosomal DNA

Abstract

The anisakid nematodes morphologically corresponding with Pseudoterranova decipiens sensu lato (s.l.) (Krabbe, 1878) from different seal or sea lion hosts and geographical origins, previously identified as Pseudoterranova krabbei, P. decipiens (s.s.), P. bulbosa, P. azarasi and P. cattani by multilocus enzyme electrophoresis, were characterized using a DNA approach. Also a population of P. decipiens (s.l.) from Chaenocephalus aceratus, the blackfin icefish, from Antarctica and another from Osmerus eperlanus, the European smelt, from Germany were included in the study. The first (ITS-1) and second (ITS-2) internal transcribed spacers (ITS) of ribosomal DNA (rDNA) were amplified by PCR from individual nematodes and analysed by single-strand conformation polymorphism (SSCP), followed by selective sequencing. While no variation in single-stranded ITS-1 and ITS-2 profiles was detected among samples representing each of the species or populations (with the exception of slight microheterogeneity), SSCP analysis of the ITS-2 amplicons allowed the unequivocal differentiation of all of the 5 sibling species of P. decipiens (s.l.) examined, which was supported by sequence differences in ITS rDNA. Samples representing the P. decipiens (s.l.) population from O. eperlanus had the same SSCP profile as those of P. decipiens (s.s.), which was supported by a lack of nucleotide difference in the ITS between them, suggesting that the former represented P. decipiens (s.s.). Based on SSCP results and ITS sequence data, P. decipiens (s.l.) from C. aceratus was genetically most distinct with respect to all other members of Pseudoterranova examined, which indicated that it may represent P. decipiens E (based on geographical origin) or a distinct species. These findings and the molecular approach taken should have important implications for studying the life-cycles, transmission patterns, epidemiology and population genetics of these anisakid nematodes, and the diagnosis of their infections.

Anisakids of seals found on the southern coast of Baltic Sea

“…It is an expected result. Data collected from fish during previous studies in Baltic Sea confirm this observation , Mattiucci and Nascettii 2007, Zhu et al 2002. There were also some positive identifications of A. simplex -it has been defined as A. simplex s.s.…”

“…Subsequently some specific rDNA fragments (ITS-1, ITS-2) were amplified using a PCR technique. NC2 and NC5 primers were used (Zhu et al 2002). The next step was a digestion of PCR products using restriction enzymes: RsaI, HhaI, HinfI and TaqI.…”

Molecular evidence that Heligmosomoides polygyrus from laboratory mice and wood mice are separate species

“…This assumes a greater interspecific genetic distance between taxa than the intraspecific distances, a condition that appears to hold in the present case, in all pairwise comparisons of genetic distance except those including the Guernsey isolate of H. p. polygyrus. The cryptic Haemonchus placei, for example, was differentiated by 15 % sequence variation from H. contortus at the mitochondrial ND4 locus (Blouin et al 1997), while Zhu et al (2002) recorded differences in ITS sequence of approximately 2 % amongst the cryptic nematode species of the Pseudoterranova decipiens complex. In both cases, as in the H. polygyrus complex, species status is based on little more than small morphological differences, differences in host identity and sequence variation.…”

Helminths of Antarctic fishes: Life cycle biology, specificity and geographical distribution

“…Palm (1999) stated that one sibling species, Pseudoterranova decipiens E, occurred in Antarctic waters. The genetic distance of this species from other species within the Pseudoterranova decipiens complex was described by Zhu et al (2002). A frequent parasite of bony fishes, Hysterothylacium aduncum (Rudolphi, 1802) was recorded from two notothenioid species from environs of the South Shetland Islands and off South Georgia (Gaevskaya et al 1990, Rokicki unpubl.).…”