Digestive haemorrhage in patients with anti-Anisakis antibodies.

OBJECTIVE: Anisakis simplex parasite causes anisakidosis, a disease that often mimics other gastrointestinal diseases such as peritonitis, appendicitis, Crohn's disease, ulcer, etc. Patients with digestive haemorrhage caused by ulcers, varices or Mallory syndrome were analysed for antibodies to the worm A. simplex. METHODS: Antibody detection was carried out by enzyme-linked immunosorbent assay (ELISA) and immunoblot using crude extracts of antigen and excretory/secretory products. Total immunoglobulin (Ig), IgG, IgM, IgA and IgE were studied. RESULTS: Eighty-seven patients were studied. The following prevalence rates were found with crude antigen: total Ig 30% (95% confidence interval 21-40), with values for IgG, IgM, IgA and IgE of 22 (CI 14-31), 17 (CI 10-26), 37 (CI 27-47) and 12% (CI 6-20), respectively. Twenty-four positive sera for total Ig response and crude products were selected for determination of specific antibodies with excretory/secretory antigens. We obtained 8, 13, 3 and 16 positive cases for total Ig, IgG, IgM and IgA respectively. The percentages of positivity within the varices and Mallory groups of patients were higher although differences were not significant (35 and 50% respectively). In a healthy population, the prevalence for total Ig is much lower (6%). Twenty-five positive sera for total Ig response were tested by means of immunoblot analysis against crude larval antigen. Concerning total Ig antibody response, 12 of the sera showed an immuno-recognition pattern concordant with the human anisakidosis reference serum (E17). Specific IgG bands were visualized in 30 sera; specific IgM and IgA in 6 and 12, respectively. Different clinical variables of these patients were studied: leucocytes, eosinophils, haemoglobin, prothrombin activity, thromboplastin time, fibrinogen, platelets and erythrocyte sedimentation rate. There were a few significant differences: for total Ig in prothrombin activity and platelets, and for IgM in eosinophils. CONCLUSIONS: The prevalence of detectable antibodies against A. simplex is higher in patients with digestive haemorrhage than in the healthy population.

Anti-anisakis antibodies in the clinical course of Crohn's disease.

BACKGROUND: Human anisakidosis is a parasitic disease whose intestinal form shares several characterictics with Crohn's disease (CD), like the ileocolonic location predominantly and the presence of granuloma. METHODS: 73 CD patients diagnosed according to Lennard-Jones' criteria were studied. The disease activity was measured by means of Harvey-Bradshaw Index. Different clinical variables were analyzed. Antigenic extract of Anisakis simplex larvae extracted from blue whiting was prepared. Determination of IgG, IgM, IgA and total specific immunoglobulin against A. simplex were carried out in sera by enzyme-linked immunosorbent assay. Immunorecognition patterns of these sera were realized by immunoblotting analysis. Results were compared with prevalence of antibodies anti-Anisakis in 251 healthy controls. RESULTS: 29% of CD patients showed detectable specific total Ig (G+M+A) against A. simplex (95% CI, 19-41). For IgG, IgM and IgA the percentages were 44, 18 and 53% (95% CI, 32-56, 10-26, and 41-65) respectively. In positive specific total Ig cases CD was localized mainly in ileum. In IgG-, IgM- and IgA-positive cases the location was predominantly ileocolonic. CD activity index was higher for groups with positive IgA (2.86 vs. 5.55; p < 0.05). CONCLUSION: Patients with CD have high prevalence of specific immunoglobulin against A. simplex. Specific IgA are associated to higher activity index of CD.

Analysis of stage-specific and shared antigens derived from Rhipicephalus sanguineus by electrophoresis and western blotting.

We carried out an SDS-PAGE analysis of antigens of Rhipicephalus sanguineus using extracts of eggs (EE), larvae (LE), nymphs (NE), male salivary glands (MSGE), male midguts (MME), female salivary glands (FSGE) and female midguts (FME). Under non-reducing conditions a common band of about 205 kDa was observed. EE, LE and NE extracts showed groups of bands between 150 and 75 kDa. A protein pattern was observed in FSGE extract with a group of bands between 75 and 50 kDa and four bands between 15 and 6.5 kDa. In this case an apparently exclusive band of molecular weight about 25 kDa was observed. Under reducing conditions similarities between LE and NE extracts increased, separating from the EE pattern. On the other hand, we have determined the presence of stage-specific and common antigens on EE, LE, NE, MSGE, MME, FSGE and FME extracts of R.sanguineus by means of immunoblots using polyclonal sera of rabbits infested with larvae, nymphs or adults of this tick. EE extract was only recognized by the anti-larva sera. Higher reactivity was observed when the extracts were tested with anti-adult sera. In these experiments a very prominent band of molecular weight about 45 kDa was detected. This band was not observed under reducing conditions. Higher reactivity with anti-adult sera was observed against FSGE extract.

Immunological response in rabbits infested with Rhipicephalus sanguineus.

In order to clarify the importance of humoral antibody in host resistance to ticks, in the present work we studied the immunological response of rabbits infested with larvae, nymphs or adults of Rhipicephalus sanguineus, using extracts of eggs (EE), larvae (LE), nymphs (NE), male salivary glands (MSGE), male midguts (MME), female salivary glands (FSGE) and female midguts (FME). When serum from rabbits infested with larvae or nymphs was tested using enzyme-linked immunosorbent assay, no reactions were observed with any of the extracts including the homologous LE or NE. In sera from rabbits infested with adult ticks, the reactions were observed in both homologous (MSGE, MME, FSGE and FME) and heterologous (EE, LE and NE) system. However, differences were seen regarding the type of antigen used. When the experiment was carried out using extracts from adults higher responses were found. With FSGE and FME antigens, antibody levels were systematically higher than those observed when MSGE and MME were used.