Ocular larva migrans caused by Toxocara cati in Mongolian gerbils and a comparison of ophthalmologic findings with those produced by T. canis.

To elucidate the pathogenic potential of Toxocara cari, we observed the ophthalmologic changes of the fundi in Mongolian gerbils, Meriones unguiculatus, after oral inoculation of 17 embryonated eggs/g body weight. Ophthalmic conditions in 8 T. cati-infected gerbils were monitored using an ophthalmoscope from day 0 to day 156 and were compared with those of 57 T. canis-infected gerbils. The results showed that T. cati larvae migrated into the eye of the gerbil and then elicited ophthalmic changes, including retinal (25%) and vitreous (50%) hemorrhaging, vasculitis (37.5%), and exudative lesions (25%). Lesions were less prevalent, however, in T. cati-infected than in T. canis-infected gerbils. Unlike in T. canis-infected gerbils, the hemorrhagic lesions did not reappear in T. cati-infected gerbils after they were absorbed. These findings suggested that T. cati larvae are a potentially hazardous pathogen for ocular toxocariasis and that Mongolian gerbils infected with T. cati may be a useful model for the study of human ocular toxocariasis caused by T. cati. This is the first study to report that T. cati larvae can induce ophthalmic lesions in the retina of gerbils.

Differential susceptibility to Brugia pahangi infection in Mongolian gerbils (Meriones unguiculatus) of different coat colour.

The influence of intraspecific host variables on the response to parasitic infections is an important aspect of host-parasite relationships, yet little is known about this aspect of filariasis for lack of a model. This study presents coat colour mutants of the Mongolian gerbil (Meriones unguiculatus) as potential new models for research into the effects of host genetic variation on response to filarial infection. Peak level of microfilaraemia, eosinophil response, body weight and degree of splenomegaly in gerbils infected with Brugia pahangi varied with agouti, albino, and black coat colour. These results suggested that coat colour-related genes might influence host immune response to developmental stages of the parasite and eosinophil-mediated reaction might cause host damage.

New animal model for human ocular toxocariasis: ophthalmoscopic observation.

BACKGROUND/AIMS: Although human ocular toxocariasis causes severe vision defect, little is known about its aetiology, diagnosis, and treatment. To develop a new animal model for human ocular toxocariasis, ophthalmological findings of fundi in Mongolian gerbils, Meriones unguiculatus, and BALB/c mice were investigated following infection with Toxocara canis. METHODS: Using an ophthalmoscope, which was specifically developed to observe the fundi of small animals, ocular changes of fundi of 20 gerbils and 11 mice were monitored after oral infection with embryonated eggs of T canis. RESULTS: Vitreous, choroidal, and retinal haemorrhages were consistently observed in Mongolian gerbils, but rarely in mice. Severe exudative lesions and vasculitis were often present in gerbils but not in mice. Migrating larvae were also frequently observed in gerbils. CONCLUSION: Mongolian gerbils are more appropriate animal model for human ocular toxocariasis than previously used experimental animal such as mice, guinea pigs, rabbits, and monkeys because of its high susceptibility of ocular infection.

Down-regulation of murine susceptibility to cerebral malaria by inoculation with third-stage larvae of the filarial nematode Brugia pahangi.

In areas where malaria is endemic, helminthic infections, caused by intestinal or filarial parasites, commonly coexist with malaria in the same individual. This study investigates the course of Plasmodium berghei malaria infection in CBA/J mice inoculated with irradiated attenuated 3rd-stage larvae (L3) of Brugia pahangi. Peripheral eosinophil counts, serum IgE levels and cytokine production revealed that the filarial antigen induced T-helper type 2 (Th2) cell predominance in these mice, which protected them against the development of cerebral malaria. These mice significantly prolonged their survival, compared with the control mice after P. berghei infection. All of the mice not inoculated with irradiated L3 died within 12 days with acute neurological manifestations unrelated to the level of parasitaemia after infection of P. berghei. Conversely, most of the inoculated mice lived more than 3 weeks following infection with P. berghei, dying in the fourth week of severe anaemia and overwhelming parasitaemia. This suggests that Th2-dominant responses lead to the down-regulation of susceptibility to murine cerebral malaria.

Suppression of mitogen-induced IL-2R alpha expression in PBL by serum from microfilaraemic rats chronically infected with Brugia pahangi.

To study immunological responses in filarial infections, interleukin-2 receptor alpha (IL-2R alpha) expression in the peripheral blood lymphocytes (PBL) of Lewis rats infected with Brugia pahangi was observed by flow cytometry. During infection, no increase in IL-2R alpha was observed in either microfilaraemic or amicrofilaraemic rats. The PBL of rats were stimulated with phytohaemagglutinin (PHA) and the frequency of IL-2R alpha-positivity was examined. After microfilariae appeared, the increase in the rate of IL-2R alpha-positivity in microfilaraemic rats was less than in amicrofilaraemic rats. The anergy of IL-2R alpha expression in microfilaraemic rats was restored when their lymphocytes were cultured with normal rat serum. IL-2R alpha expression in normal uninfected rats decreased when cultured with serum from microfilaraemic rats. These results suggest that microfilariae-related factors in rat serum suppress PHA-induced IL-2R alpha expression in PBL and thus inhibit cell proliferation and host immunity.

Brugia pahangi: production of a monoclonal antibody reactive with the surface of infective larvae.

Monoclonal antibodies against infective third-stage larvae (L3) of Brugia pahangi were generated from mice immunized with L3 antigens. The monoclonal antibodies were L3 stage-specific or stage-nonspecific. A BpG1 monoclonal antibody (IgG1 subclass) showing L3 stage-specificity was examined in detail. BpG1 recognized the surface of B. pahangi L3 and also reacted with the surface of Brugia malayi L3 but not with the surface of filarial worms of other genera, such as Acanthocheilonema viteae and Litomosoides carinii. BpG1 promoted cellular adhesion to the surface of B. pahangi L3. BpG1 bound on living L3 was shed but the shedding rate was relatively slow. The surface antigen recognized by BpG1 had a molecular weight of 58 kDa. It was stable to heat and periodate treatments but sensitive to trypsin digestion and was released from living L3 by SDS but not by Triton X-100 or CTAB. Preincubation of L3 with BpG1 significantly reduced the recovery rate of worms compared with the preincubation with a monoclonal antibody (IgG1 subclass) against the inner tissues of B. pahangi L3 or control supernatant of P3U1 myeloma cells. This result suggests that the antigen containing the BpG1 epitope may be one of the targets of a protective immune response against Brugia infection.

Induction of Fc epsilon RII/CD23 on human T cells by excretory and secretory antigen of Dirofilaria immitis.

We investigated the capacity of excretory and secretory antigen (ES) derived from living filarial worms in the induction of CD23 expression on human peripheral blood T cells by using flow cytometry. ES (10 micrograms/ml) significantly induced the expression of CD23 on human T cells. Moreover, increased CD23 expression was completely abolished by preincubation with specific antibody to ES. The results suggest that ES might play a certain role in IgE antibody production by induction of CD23 expression on T cells.

Strongyloidiasis influences the elevation of adult T-cell leukemia-associated antigen antibody titer.

We have carried out a seroepidemiological study on the infectious situation of adult T-cell leukemia and strongyloidiasis in Okinawa and in the south-western islands of Japan. The mean titer of adult T-cell leukemia virus-associated antigen (ATLA) antibody of human T-lymphotropic virus type-1 (HTLV-1) carriers with strongyloidiasis was significantly higher than that of HTLV-1 carriers without strongyloidiasis. This suggests the possibility that strongyloidiasis is also associated with an elevation of the ATLA antibody titer.

Non-specific immune suppression by CD8+ T cells in Brugia pahangi-infected rats.

Non-specific suppression of the immune response was investigated in Brugia pahangi-infected Lewis rats. The proliferative response of peripheral blood lymphocytes or splenic non-adherent cells to mitogens was significantly reduced by B. pahangi infection. The degree of hyporesponsiveness of splenic non-adherent cells to mitogens was comparable between microfilaremic and non-microfilaremic animals. The suppressed proliferative response of splenic non-adherent cells was restored by blocking with anti-CD8 monoclonal antibody. After separation of T cells into CD4+ and CD8+ subpopulations, only CD8+ T cells from B. pahangi-infected rats suppressed the proliferative response of normal spleen cells to concanavalin A. CD8+ T cells from normal rats had no suppressive effect. On the other hand, the proliferative response of CD4+ T cells to concanavalin A was comparable between normal and infected rats. These results suggest that CD8+ T cells participate in the non-specific suppression of immune response in experimental filariasis.

Amino acid composition of the highly purified allergen from Dirofilaria immitis.

The amino acid analysis of the purified allergen from Dirofilaria immitis revealed that it had a total number of 98 amino acid residues. The allergen was remarkably rich on glutamic acid (15 residues), lysine (14 residues) and aspartic acid (12 residues). No half-cystine was obtained. The ratio of the acidic residues to the basic ones was 1.4. The weak positive charge of the allergen is contributed by the fewer number of basic residues relative to acidic residues.

Seroepidemiological studies on the effects of filarial parasites on infestation of adult T-cell leukemia virus in the Goto Islands, Japan.

Patients with filariasis were commonly observed until 1961-1970 in the Goto Islands, an area where adult T-cell leukemia (ATL) is endemic. The positive rate of antibodies to ATL virus-associated antigen among persons with a high antibody titer to filarial antigen was higher than that among persons with a low antibody titer in both sexes. Thus, filarial parasites might have some promoting effects on ATL virus infection and/or ATL virus proliferation among inhabitants in the endemic areas of filariasis and ATL.

Preparation of a highly purified allergen from Dirofilaria immitis. Reaginic antibody formation in mice.

A Dirofilaria immitis protein allergen was purified; it had a molecular weight of 15,000-20,000 and a carbohydrate content of 2%. The allergenic activity of adult Dirofilaria extracts was assayed by passive cutaneous anaphylaxis (PCA) in rats using mouse sera obtained by immunization with various fractions. The mouse-Dirofilaria system was used to study the degree of purification of allergen. The purified Dirofilaria allergen appeared as one band after sodium dodecyl sulphate polyacrylamide gel (SDS-gel) electrophoresis and one precipitin arc by immunoelectrophoresis (IEP). It was inclined to aggregate in buffered solution. The results suggested that the allergen--reagin axis was a simple single antigen-antibody interaction. Immunological responses to the purified allergen were compared among four inbred strains, two hybrid strains and two outbred strains of mice. They produced relatively high titres of reaginic antibody but did not produce detectable indirect haemagglutinating test (IHA) antibody. Among the strains tested, BALB/c was a high responder and also contained to produce the reaginic antibody for longer than the other strains.

Immunological and physicochemical properties of a highly purified allergen from Dirofilaria immitis.

Allergen in crude extract of Dirofilaria immitis was purified and separated from IgG-inducing antigens by a combination of DEAE-Sephadex A-50 chromatography, Sephadex G-200 gel filtration and starch gel zone electrophoresis. The purified preparation was proved to be one protein band by sodium dodecyl sulfate polyacrylamide gel (SDS-gel) electrophoresis and one precipitin arc by immunodiffusion. The molecular weight of the purified allergen was estimated to be approximately 20,000 by gel filtration and 15,000 by SDS-gel electrophoresis. The carbohydrate content of the preparation was apparently low, about 2%. The allergen was positively charged, and its determinant group was protein in nature. It was resistant to tryptic, pepsic and chymotryptic digestion, periodate oxidation and DNase and RNase digestion but very sensitive to pronase digestion. Allergen was inclined to aggregate each other in the buffered solution. It was also very resistant to vibration, heat (80 degrees C for 1 h) and acid (pH 2.5) and alkali (pH 11.0) treatments. Rats as well as mice immunized with allergen developed only a reaginic antibody and no hemagglutination antibody.