Evaluation of the allergenicity of goat milk, cow milk, and their lactosera in a guinea pig model.

The object of this study was to determine the allergenicity of goat milk (GM) and cow milk (CM) and that of their respective lactosera (GML and CML), by in vivo and in vitro assays. Two systemic tests for anaphylaxis were carried out in guinea pigs, the animals being sensitized orally with the 2 types of milk and lactosera. Sera were taken from the orbital sinus of the experimental animals at 0 and 22 d of the experiment to perform the serological study and the passive cutaneous anaphylaxis test. For the latter, the guinea pigs were sensitized passively with antibodies against the 4 antigen solutions. Enzyme-linked immunosorbent assay and Western blot were used to determine the specific antibodies of the isotypes immunoglobulin G1 and immunoglobulin G(Fc) developed against the same 4 antigen solutions. From these anaphylaxis and antibody-production tests, it was concluded that GM is hypoallergenic when compared with CM. The lactosera produced more closely grouped results, with values always below those of the corresponding milk. None of the proteins in the 4 immunizing solutions were identified as being their main allergen. These results show the hypoallergenicity of GM versus CM, and also that both casein and lactoserum proteins may be responsible for allergy in each case. To analyze the possibility of producing an innocuous food for those allergic to milk proteins, it would be of interest to identify the epitope(s) responsible for such allergenicity.

Prevalence of Cryptosporidium infection in goats maintained under semi-extensive feeding conditions in the southeast of Spain.

The aim of the present study was to estimate the prevalence of Cryptosporidium parvum infection in goats maintained under semi-extensive feeding conditions in the southeast of Spain. Four farms took part in the study. Samples were obtained during the autumn and spring kidding seasons. The goats were divided into three groups: group I (0-15 days), group II (> 15 days-2 months) and group III (> 2 months-9 years). Faecal specimens were concentrated and smears of the sediment (25 microl) were stained by the modified Ziehl-Neelsen acid fast technique. Parasite oocysts were identified in all the farms; of the 582 goats, 111 were found to have the parasite in their faeces, i.e. the prevalence was 19.1%. The frequency distributions of the Cryptosporidium oocyst counts according to the age categories were 10.4%, 13.4% and 25.2% for groups I, II and III respectively. With respect to the kidding season, nodifferences were found. The present study revealed the high prevalence of Cryptosporidium infection in goats in the studied zone.

Thermogenesis associated to the intake of a diet non-supplemented or supplemented with n-3 polyunsaturated fatty acid-rich fat, determined in rats receiving the same quantity of metabolizable energy.

The beneficial effects of n-3 polyunsaturated fatty acids (PUFA) are well known, but their consumption in western countries is chronically insufficient, and so it is recommended that diets should be supplemented with a fat rich in these fatty acids. However, the effect of such diets on the energy expenditure remains a controversial question. Precise data concerning the effect of using under the same metabolizable energy intake, a diet non-supplemented or supplemented with a fat rich in n-3 PUFA are not available. This type of information was obtained using rats at weaning fed a diet supplemented or non-supplemented with 10% of fish oil. Between the 30th and 60th day after starting the experiment, the energy and protein balance was established by means of the comparative slaughter method. The blood levels of different metabolites were also determined. Although total thermogenesis did not vary between the two groups, consumption of the fish oil diet led to a lower level of thermogenesis associated with the oxidation of protein, and a higher one of that associated with the oxidation of fat. We conclude that the thermic effect of feeding is a combination of independent processes. Due to their specific metabolism, n-3 PUFA may be considered essential compounds to maintain the energy balance.

Polyunsaturated fatty acids and parasitism: effect of a diet supplemented with fish oil on the course of rat trichinellosis.

Dietary fish oil has a beneficial effect on heart and some bacterial diseases and apart from other effects, some studies have revealed their ability to modulate the course of inflammatory and autoimmune diseases. The study here reported was designed to evaluate the possible influence of a fish oil supplement on the course of a Trichinella infection. Nutritional, parasitological and immunological parameters were analyzed. Two groups of 20 Wistar rats, one fed a standard diet and the other one a standard diet supplemented with fish oil, were infected with 1000 L1 larvae. Other two uninfected groups served as control. Results were as follows: fish oil diet intake and infection have, respectively, a positive and a negative effect on growth and food utilization. The negative effect is detected later in animals fed the fish oil diet. A reduction of 30.9 and 36.6% in the number of adult worms and L1 larvae, respectively, was observed in the fish oil group as compared to the standard diet group. Production of IFNgamma (Th1 response) and IL4 (Th2) response was measured in stimulated splenic cells. The fish oil diet increased both IFNgamma and IL4 levels. At 6 days after infection both IFNgamma and IL4 responses were detected, but at 36 days after infection only IL4 was detected in the standard group. The level of somatic and cuticular antibodies was not affected by the diet.

Trichinella britovi human infection in Spain: antibody response to surface, excretory/secretory and somatic antigens.

A third outbreak of Trichinella britovi with 140 people involved, occurred in Granada Spain (December 1998). The source of infection was sausage made from uninspected wild boar meat. Fifty-two patients agreed to participated in this study. An elevated eosinophil level (> 5%) was detected in 59.6% of patients, and persisted in most of these cases for two months. A moderate IgG response was observed. At the onset of symptoms, Western blot (WB) test detected more positive cases than Enzyme linked immunosorbent assay (ELISA) and indirect immunofluorescence (IIF). Six months from infection, ELISA revealed fewer positive cases than the other two tests. It would appear that the response to somatic antigens starts earlier than those to cuticular and excretory/secretory (ES) antigens and that the response to ES antigens is the first to decrease.

Short report: Human infection with Trichinella britovi in Granada, Spain.

An outbreak of trichinellosis caused by ingestion of pork infected with Trichinella britovi occurred in the province of Granada in southern Spain in April-May 2000. Thirty-eight people were affected and 15 of them were hospitalized at the University Hospital of San Cecilio (Granada). The probable source of infection was sausage made from uninspected wild boar meat and inspected pork. Ninety-two percent of the patients had myalgias, 47.6% had diarrhea and/or vomited, 78.6% had periorbital edema, and 76.0% had fever. Twenty-two patients (15 hospitalized and 7 nonhospitalized) were serologically studied. Eosinophil levels were less than 5% of the total leukocyte count in 86.7% of the patients. Levels of creatinine phosphokinase (range = 200-2,213 U/L) and lactate dehydrogenase (range = 560-7,558 U/L) were elevated in 85.7% and 78.6% of the patients, respectively. Sixteen (72.7%) and 20 (90.9%) patients were positive for T. britovi by indirect immunofluorescence and Western blot, respectively.

Human trichinellosis in Sourthern Spain: serologic and epidemiologic study.

An outbreak of trichinellosis caused by wild boar meat occurred in the Iruela (Jaen) in southern Spain in February 1996. Thirty-five people were diagnosed on the basis of epidemiologic data, but only 24 patients agreed to participate in this study. Twenty-three (96%) had symptoms suggestive of trichinellosis. Immunofluorescent and Western blot test results for trichinellosis were positive in 18 persons, and 15 had circulating Trichinella spiralis antigens. These findings suggest that results of tests for circulating antigens in conjunction with clinical presentation are useful for the diagnosis of trichinellosis.

Humoral immune response and antigenemia in sheep experimentally infected with Schistosoma bovis. Cross-reactivity with Fasciola hepatica antigens.

Circulating antigen level, IgG antibody response to worm antigens and to excretory/secretory products (ES), and specificity to Fasciola hepatica antigens were determined in 6 Schistosoma bovis-infected sheep at weekly intervals for 15 wk. A noninfected control group was included. An enzyme-linked immunosorbent assay (ELISA) sandwich and a double-antibody ELISA test was used for antibody and antigen detection, respectively. The infection induced an early and relatively low IgG response to adult worm extract. This response was significantly elevated by 3 wk postinfection (PI), reached its maximum level at 9 wk PI, and was followed by a subsequent decrease. The response to ES antigens was slightly higher than that to adult worms, although the response started later, at 8 wk PI, and remained at its maximum level until 15 wk. A remarkable level of cross-reactivity was observed when adult F. hepatica extract was used. However, a low degree of cross-reactivity was found with ES antigen. The ELISA for circulating antigens was performed at weekly intervals for 8 wk. Antigens were detected as early as the first week of infection, although differences were statistically significant from week 5 onward. The highest values were observed at 7 week PI.

Fasciola hepatica: partial characterization of circulating antigens.

Antigens present in sera from Fasciola hepatica-infected goats were partially characterized by means of polyacrylamide gel electrophoresis of immunoprecipitates obtained by counterimmunoelectrophoresis. Agar containing the 2 immunoprecipitin lines, agar containing no precipitin lines, a crude extract of F. hepatica, and normal rabbit IgG were assayed. The electrophoretic pattern of 1 precipitin line showed, apart from the light and heavy chains of IgG and other faint bands, 2 prominent polypeptides of about 70 and 85 kDa. The other precipitin line showed a similar pattern, but with the 70-kDa band absent.

Immunological characteristics and localization of the Trichinella spiralis glutathione S-transferase.

Trichinella spiralis glutathione S-transferase (TsGST) was isolated from crude extracts of L1 larvae by glutathione-affinity chromatography. Two closely migrating polypeptides with molecular masses of 28.5 and 28 kDa were identified by electrophoresis. Three isoforms of pI 5.6, 5.8, and 6.0 were detected by isoelectric focusing. Purified TsGST showed a low transferase activity as measured with 1-chloro-2,4-dinitrobenzene; glutathione peroxidase activity was also demonstrated using cumene hydroperoxide. A rabbit antiserum against TsGST reacted by western blot with crude extracts of Trichinella britovi and Trichinella nativa but not with extracts of Trichinella pseudospiralis, Fasciola hepatica, Schistosoma bovis, Schistosoma mansoni, Dirofilaria immitis, Toxocara canis, or Anisakis sp. TsGST was detected by western blot in extracts of T. spiralis adults, but not in newborn larvae or L1 excretory-secretory products; yet, an antiserum against T. spiralis excretory-secretory products reacted with TsGST. By immunoelectron microscopy, TsGST was found in the granules of the alpha- and beta-stichocytes of L1 larvae, as well as in some granules of the stichocytes of 72-hr adults. Rabbits experimentally infected with T. spiralis developed substantial levels of anti-TsGST antibodies. Moreover, circulating TsGST was detected in serum by a sandwich-enzyme-linked immunosorbent assay, isolated from serum by glutathione-affinity chromatography, and characterized as TsGST by western blot.

Resistance to Schistosoma bovis in sheep induced by an experimental Fasciola hepatica infection.

Sheep infected with Fasciola hepatica for 10 wk acquired a substantial level of resistance to challenge with Schistosoma bovis. The worm burden was reduced by 87.2% (P < 0.01) compared with that of a control group. But when sheep primarily were infected with S. bovis and 6 wk later with F. hepatica, no significant reduction in the S. bovis burden was observed.

Seroepidemiological studies on five outbreaks of trichinellosis in southern Spain.

Five outbreaks of trichinellosis, involving 86 people, took place in Andalucía (Southern Spain) between 1984 and 1987. Four of them were caused by eating infected pork and one by eating wild boar meat. The human sera were analysed by the immunofluorescence assay test, which showed circulating anti-Trichinella antibody titres of up to 1:10240. A micro-ELISA test, carried out with sera from two of the outbreaks, revealed the presence of circulating antigens 68 and 78 days after infection. Finally, 11 of 17 samples of products prepared with pork and wild boar meat showed Trichinella larvae after acid-pepsin digestion. The close geographical localization of three of the outbreaks suggests that the southwestern corner of Spain is an endemic focus of trichinellosis.

Differentiation between Trichinella spiralis and T. pseudospiralis infective larvae by a monoclonal antibody.

Crude saline extracts of Trichinella spiralis and T. pseudospiralis infective larvae were studied by Western blot analysis using a monoclonal antibody, named ES/TA2 and produced against T. spiralis larvae. This monoclonal antibody recognized seven major antigenic components in T. spiralis larvae with apparent Mr: 45, 48, 50, 68, 70, 92 and 105 kDa and five in T. pseudospiralis larvae: 38, 50, 70, 72 and 92 kDa. SDS-PAGE of both extracts did not reveal appreciable differences in the range of molecular weights recognized by ES/TA2. These facts show the existence of immunological differences among proteins with apparently identical molecular weights.

Human Gongylonema infection in Spain.

A case of gongylonemiasis in a 31-year-old woman of Granada, Spain, is reported. The worm, clearly belonging to Gongylonema genus, could not be identified definitively as Gongylonema pulchrum, the species usually described in humans.