No abstract

No abstract

Genetic markers (ribosomal DNA and mitochondrial DNA) were used for molecular dissection of the Anisakis simplex sensu lato (s.l). complex populations. Host fish were caught off Moroccan coasts, where only Anisakis pegreffii is present, the sympatric area comprising Spanish coasts, and the Little Sole Bank fishing area from Nordeast Atlantic Ocean where the only present species is A. simplex sensu stricto(s.s.). Sequence variations in the amplification products were then assessed indirectly by digestion with restriction endonucleases or directly by sequencing for 623 L3 larvae. The sequences were used to infer the relationships between the two species under study using various methodological approaches. We reveal the high genetic diversity of Anisakis simplex s.s. and A. pegreffii in both mitochondrial and nuclear genes. We detected 10 and 2 fixed differences between A. simplex s.s and A. pegreffii in the Cox2 and ITS1, respectively. We found a proportion of putative hybrids below 20% with similar figures on the Atlantic and Mediterranean coasts. Moroccan hybrids were more similar to A. pegreffii reflecting backcrosses between these mixed genotypes and his ancestor A. pegreffii. We discuss the possible interpretation of these putative hybrids.

Monoclonal antibodies were produced and characterized to the metacyclic trypomastigotes stage of Trypanosoma cruzi. All the monoclonal antibodies (GR-12C/5; GR-P2B/5; GR-F2B/1; GR-J2C/6; GR-E6B/3; GR-A5B/5) also recognized antigens of epimastigotes of T. cruzi. These antigens are associated with the plasma membrane, flagellum, and an intracellular structure located by the nucleus. Immunoprecipitation using the monoclonal antibodies GR-J2C/6 and GR-E6B/3 followed by SDS-PAGE analysis has led to the following results: GR-J2C/6 precipitated two molecules with apparent molecular weights of 58 and 60 kD; the antigen recognized by GR-E6B/3 was a molecule with molecular weight of 68 kD. Only two of the monoclonal antibodies (GR-12C/5 and GR-P2B/5) recognized antigens of promastigotes of both Leishmania mexicana and L. infantum, and are associated with the plasma membrane.

A two-site enzyme-linked immunosorbent assay (ELISA) based on the avidin-biotin system was developed to study circulating antigens in the sera of rats orally or intraperitoneally infected with L3 larvae of Anisakis simplex s.l. Somatic and excretory-secretory antigens were detected from 24 h post-infection in all infected rats. A direct relationship between the number of larvae inoculated and the concentration of antigens in the sera was demonstrated; however, the amount of antigen decreased with the course of the infection. Comparative analysis revealed that intraperitoneal inoculation is most appropriate for detecting excretory-secretory antigens in serum. The minimum concentrations of A. simplex antigen detectable were approximately 2.5 microg/ml for somatic and excretory-secretory antigens. This antigen detection assay may have application in the diagnosis of anisakiasis.