title: Proteolytic activity in Hysterothylacium aduncum (Nematoda: Anisakidae), a fish gastrointestinal parasite of worldwide distribution.
abstract: Proteases have a significant role in the life cycle of parasites and the pathogen-host relationship, being regarded as important virulence factors. In the parasitic nematode Hysterothylacium aduncum proteolytic activity was measured during in vitro development from third larval stage (L3) to mature adult, using DQ red casein as a fluorogenic substrate. Proteolytic activity was detected in all the developmental stages studied and at all pH values within the range employed (2.0-7.5). The assay with specific inhibitors permitted the determination of metalloprotease activity, and, to a lesser extent, that of aspartate- and cysteine-protease. Serine-protease activity was the lowest of those studied. In L3 recently collected from the host fish (L3-0 h), the greatest activity was found at an optimum pH of 4.0 and was mainly inhibited by 1,10-phenathroline (metalloprotease inhibitor). This metalloprotease activity in L3-0 h (infective stage) may be related to the invasion of the host tissues by this larva. In the other developmental stages, the greatest protease activity was found at pH 5.5, although at pH 4.0 a lower activity peak was detected. On the other hand, our data show that the proteolytic activity of the nematode varies according to the presence of pepsin (an aspartic-protease) in the culture medium. Thus, at pH 4.0, activity was greater in the absence of pepsin, with increasing aspartic-protease activity. Together with the detection of aspartic-, cysteine- and metallo-protease (enzymes involved in digestion in invertebrates) in all the developmental stages of the parasite taking place in the digestive tract of the host fish, this allows us to suggest that the pepsin in the culture medium mimics the predigestion conditions in the habitat of the worm within the host and that the activity detected may have, amongst others, a digestive function.
authors.affiliation: Departamento de Parasitología, Facultad de Farmacia, Universidad de Granada, 18071 Granada, Spain.
authors.forename: David
authors.initials: D
authors.lastname: Malagón
authors.forename: Rocío
authors.initials: R
authors.lastname: Benítez
authors.forename: Francisco Javier
authors.initials: FJ
authors.lastname: Adroher
authors.forename: Manuel
authors.initials: M
authors.lastname: Díaz-López
mesh_terms: D000818:Animals; D017191:Ascaridida Infections / Q000469:parasitology / Q000662:veterinary*; D001199:Ascaridoidea / Q000201:enzymology* / Q000254:growth & development; D005393:Fish Diseases / Q000469:parasitology*; D006790:Host-Parasite Interactions; D006863:Hydrogen-Ion Concentration; D007814:Larva / Q000201:enzymology / Q000254:growth & development; D008018:Life Cycle Stages; D045726:Metalloproteases / Q000037:antagonists & inhibitors / Q000378:metabolism; D010434:Pepsin A / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D010473:Perciformes / Q000469:parasitology*; D010618:Phenanthrolines / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D059748:Proteolysis; D013030:Spain
chemical_list: D010618:Phenanthrolines; D011480:Protease Inhibitors; D045726:Metalloproteases; D010447:Peptide Hydrolases; D010434:Pepsin A; C025205:1,10-phenanthroline