Immune response to chronic Toxocara canis infection in a mice model.

AIMS: The zoonotic nematode Toxocara canis causes larva migrans syndrome that induces an immune response characterized by the production of antibodies and eosinophilia. A Th2 polarization has been associated with the infection, but there are still details of the cellular and humoral immune response that need to be described. Thus, the aim of this study was to describe the systemic host immune response to T canis chronic infection in a mouse model. METHODS AND RESULTS: BALB/c mice were inoculated once with 500 T canis embryonated eggs, per os. After 49 days, the amounts of larval found in brain and muscle tissues were statistically two and four times higher, respectively, than the amounts found in lung, liver, kidney or heart tissues. Splenic proportions of F4/80+ cells, as well as B, cytotoxic T and CD4+ Foxp3+ lymphocytes, were statistically higher (P ≤ .05, P ≤ .01, P ≤ .001 and P ≤ .001, respectively) as compared with control mice. In lymph nodes, some of these proportions changed, with the exception of F4/80+ cells. IgG1 levels in infected mice sera were increased. IL-4, IL-10 and VEGF levels were statistically higher in spleen (P ≤ .05, all) and sera (P ≤ .01, P ≤ .05 and P ≤ .05, respectively) in the infected mice. Also, in infected animals, IL-5 serum levels were increased (P ≤ .01). CONCLUSION: These results suggest that T canis chronic infection in BALB/c mice results in a type 2 response with an incipient regulatory response.

Visceral larva migrans syndrome: analysis of serum cytokine levels in children with hepatic lesions confirmed in radiological findings.

Human toxocariasis is a helminth zoonosis resulting from accidental infection of humans by the roundworms Toxocara canis (T. canis) and cati (T. cati). The infection occurs in five forms: systemic (VLM), ocular, neurological, covert and asymptomatic. The aim of this study was to characterize the radiological and immunological findings in hepatic inflammation during the course of systemic infection by Toxocara sp. in children. Fifteen children, 2 to 17 years of age, with serological diagnosis of T. canis infection underwent abdominal ultrasonography and computer tomography (CT). Eosinophil counts, immunoglobulin E titres, interleukins IL-1α, IL-4, Il-6, IL-10 and interferon gamma were measured for all patients. Abdominal ultrasound revealed multiple hypoechoic areas in the livers of all patients. On the CT images, the hepatic lesions were seen as multiple, ill-defined, oval low-attenuating nodules that measured 6 to 9 millimetres in diameter. The nodules were usually best seen in the portal venous phase and were not seen on arterial-phase images. Significant intergroup differences were observed in the concentrations of IL-1α, IL-4, IL-6 and IL-10. The level of IFN-γ was not significantly elevated in patient sera relative to controls. The analysis shows that the production of anti-inflammatory cytokines is insufficient for granuloma formation in children presenting liver lesions in the course of VLM.

Factors affecting disease manifestation of toxocarosis in humans: genetics and environment.

Toxocara canis is regarded as the main cause of human toxocarosis but the relative contribution of T. cati is probably underestimated; serological and other diagnostic methods used in most studies of this zoonotic disease do not distinguish between the two parasites. The definitive hosts for T. canis are caniidae. Pups generally have higher infection rates than adult animals and are a major source of eggs in the environment. Humans usually acquire T. canis infection by accidental ingestion of embryonated eggs or encapsulated larvae from the environment or contaminated food, such infections may lead to visceral larva migrans (VLM), ocular larva migrans (OLM) or covert toxocarosis (CT). Although a mixed Th1- and Th2-mediated immunological response, particularly with high levels of IgE and eosinophilia is observed, the underlying mechanisms of molecular and immunopathogenesis for the development of the symptomatic syndromes of VLM, OLM, or of asymptomatic CT are largely unclear. Studies have indicated that immunological defences against various infectious diseases may be highly influenced by complex interactions of environmental and host genetic factors e.g. MHC class I and II, also known as human leucocyte antigen (HLA). Toxocara spp. infections are associated with a polarized CD4(+) Th2 response with high IgE levels and eosinophilia, mediated mainly by HLA class II molecules. Associations have been made between HLA class II and pathological severity and host genetic effects on exposure to infection. Recent research suggests Foxp3(+) CD4(+)CD25(+)-expressing T regulatory (Treg) cells play a role in regulation of the immunopathology of granulomas in experimental toxocaral granulomatous hepatitis and in enhanced expression of TGF-ß1, which is an important factor for the local survival and function of Treg observed during T. canis invasion in the mouse small intestine, liver, muscle, and brain. Since the potential susceptibility loci HLA class II molecules, are considered involved in the regulation of a Th2-dominant immunity which is highly controlled by Foxp3(+) CD4(+)CD25(+) Treg cells by stimulation through TGF-ß1, which thus provides a beneficial environment to T. canis larvae but severe injuries to local organs. However, TGF-ß1 variant Leu10Pro known to be involved in disease severity warrants further elucidation as this too may have a role in the severity of human toxocarosis. Exploration of TGF-ß1 polymorphism, Foxp3(+) CD4(+)CD25(+) Treg cells, and MHC polymorphisms may allow insight into the contribution made by environmental and genetic factors in influencing disease syndrome type and severity in humans with toxocarosis.

Toxocaríase: larva migrans visceral em crianças e adolescentes / Toxocariasis: visceral larva migrans in children

OBJETIVOS: Apresentar investigação detalhada de fatores de risco, sintomatologia, exames laboratoriais e de imagem que possam contribuir para o diagnóstico clínico-laboratorial da larva migrans visceral (LMV) em crianças e mostrar a importância do diagnóstico e do tratamento para evitar complicações oculares, hepáticas e em outros órgãos. FONTES DOS DADOS: Revisão de literatura utilizando os bancos de dados MEDLINE e LILACS (1952-2009), selecionando os artigos mais atuais e representativos do tema. SÍNTESE DOS DADOS: LMV é uma doença infecciosa de apresentação clínica inespecífica cuja transmissão está relacionada ao contato com cães, principalmente filhotes, podendo evoluir com complicações sistêmicas tardias em órgãos vitais como o olho e sistema nervoso central. Para diagnóstico laboratorial, pode ser utilizado IgG (ELISA) anti-Toxocara canis, cujos pontos de corte mais elevados sugerem doença recente e, os mais baixos, infecção leve ou em resolução. A resposta terapêutica pode ser avaliada por meio da contagem de eosinófilos no sangue. Esse artigo atualiza o pediatra em relação à LMV, doença de alta prevalência no mundo e no Brasil. CONCLUSÕES: O diagnóstico de LMV depende principalmente da epidemiologia da presença de cães no domicílio da criança, associada ao ELISA (IgG para T. canis), utilizando antígenos Toxocara de excreção e secreção. São necessários estudos prospectivos para avaliar a melhor droga na terapêutica. A prevenção é a estratégia mais importante devido à alta prevalência de T. canis na região urbana.

OBJECTIVES: To present a detailed investigation of risk factors, symptoms, and laboratory and imaging tests that may be useful to establish the clinical laboratory diagnosis of visceral larva migrans (VLM) in children, demonstrating the importance of diagnosis and treatment to prevent complications in the eyes, liver, and other organs. SOURCES: Literature review using the MEDLINE and LILACS (1952-2009) databases, selecting the most recent and representative articles on the topic. SUMMARY OF THE FINDINGS: VLM is an infectious disease with non-specific clinical presentation, whose transmission is related to contact with dogs, especially puppies, and which may progress to late systemic complications in vital organs such as the eyes and the central nervous system. IgG (ELISA) anti-T. canis can be used to establish the laboratory diagnosis. Higher cutoff points suggest recent illness and lower cutoff points demonstrate mild infection or infection in remission. Therapeutic response may be assessed by means of eosinophil blood cell count. The present article provides the pediatrician with updated information regarding VLM, a disease of high prevalence worldwide and in Brazil. CONCLUSIONS: The diagnosis of VLM depends mainly on the presence of dogs in the child's household, associated with ELISA (IgG anti-T. canis), using excretory-secretory antigens of Toxocara canis. Prospective studies are warranted to assess the best drug therapy. Prevention is the most important strategy because of the high prevalence of T. canis in urban areas.

Estudio en niños con diagnóstico presuntivo de toxocariasis en Santa Fe, Argentina / Analysis of children with a presumptive diagnosis of toxocariasis in Santa Fe, Argentina

La toxocariasis humana es una parasitosis de amplia distribución en el mundo, cuyo agente etiológico más importante es el Toxocara canis, parásito del perro. El hombre adquiere esta zoonosis mediante la ingesta de huevos infectivos; en el intestino delgado desarticulan sus envolturas y las larvas se liberan atravesando la mucosa, ubicándose en diversos tejidos. Los niños son la población de mayor riesgo. El diagnostico clínico es dificultoso. El test de ELISA usando antígenos de excreción-secreción de la larva, es la técnica de elección. Se estudiaron por esta técnica inmunoserológica 100 niños con diagnóstico presuntivo de toxocariasis y se revisaron posteriormente sus historias clínicas de Hospital. La técnica de diagnóstico fue estandarizada y validada en el laboratorio. Su aplicación permitió identificar dos poblaciones de niños: infectados (59) y no infectados (41). La eosinofilia fue más frecuente en la población infectada (100% vs. 85.2%, p = 0.017); no así la leucocitosis (p = 0.950). La fuerza de asociación de ambos parámetros fue mayor en la población positiva (R = 0.918). La dificultad respiratoria se presentó más frecuentemente en pacientes con ELISA positiva, considerados como infectados (p = 0.05). La edad promedio de los positivos fue significativamente mayor que la de los negativos (p = 0.009). Se halló eosinofilia en el 100% de los infectados y en el 85.2% de los no infectados. La tenencia de canes en los domicilios no fue significativamente mayor entre los casos positivos que entre los negativos (p = 0.53). Sería necesario investigar esta enfermedad en niños consultantes con sospecha clínica, y promover las medidas de prevención, diagnóstico temprano y su correcto tratamiento.

Human toxocariasis is a parasitic disease found worldwide. The most important etiological agent is Toxocara canis, a dog parasite. Humans are infected by the ingestion of their eggs; the eggs hatch in the small intestine and the larvae migrate through the capillaries, taking up residence in different tissues. Clinical manifestations are associated with mechanical and/or reaction damage caused by these parasites larvae. Clinical diagnosis is difficult. The method applied in this work is the demonstration of antibodies against the helminth in the blood of children, target host population of this parasitic disease. An ELISA test was performed using T. canis larval excretory-secretory products as antigen. A total of 100 children presumptively diagnosed of toxocariasis that had been derived from different services of the Regional Children’s Hospital for complementary studies, were included in the analysis. The test detected two different populations: infected (59) and non-infected (41). The statistical analysis showed a non significant association between infection and sex (p = 0.279). Infected subjects tended to be older than the non infected (p = 0.009). Eosinophilia was detected in 100% of seropositive children and in 85.2% of the seronegative. There was no significant association between infection and leucocytosis ( = 0.950). The association of these two parameters was significantly higher among infected patients (R = 0.918). Respiratory symptoms and signs were more frequently detected in the positive population (p = 0.05). Dogs tenancy was as frequent among infected as in the non infected homes (p = 0.53). According to these results, prevention, early diagnosis and opportune treatment for toxocariasis should be considered as prioritary health activities in this region.

Prevalence of Toxocara-induced liver granulomas, detected by immunohistochemistry, in a series of autopsies at a Children's Reference Hospital in Vitoria, ES, Brazil.

The aim of this investigation was to study the frequency of visceral larva migrans (VLM) granulomas in autopsies at a Children's Reference Hospital in Vitoria, ES Brazil, where anti-Toxocara antibodies are frequently detected in the serum of children admitted at the hospital. Two liver fragments from 310 autopsies of children aged between 1 and 15 years were paraffin embedded, and sections were stained with hematoxylin and eosin and submitted to detection of Toxocara antigens using a rabbit anti-Toxocara serum. Among the 24 cases with granulomatous lesions, ten had eosinophil-rich granulomas positively stained with the anti-Toxocara serum. Some were typical epithelioid granulomas, with a positive reaction in multinucleated giant cells, epithelioid cells, or necrotic debris. The results showed that VLM granulomas are the most frequent granulomatous hepatitis in children in our county. This agrees with the high prevalence of anti-Toxocara antibodies in the serum of children admitted to the Children's Reference Hospital. The 3.2% frequency of liver VLM granulomas in autopsies is less than the 30-39% frequency of positive serology in these children, probably reflecting the low larval burden in infected children.

Toxocariosis en niños de una region subtropical / Toxocariasis in children from a subtropical region

La toxocariosis está presente en todo el mundo, pero se considera en mayor riesgo a los habitantes de zonas com deficiencias sanitarias y particularmente a los niños. El objetivo de este trabajo fue conocer aspectos inmunológicos y clínicos de la infección infantil en un área subtropical de Argentina, para lo cual se estudiaron 182 niños de ambos sexos de la ciudad de Resistencia (Noreste de Argentina), de 0 a 16 años, con eosinofilia mayor al 10%. Se realizaron exámenes clínicos, encuestas epidemiológicas, exámenes copropa-rasitológicos y dosajes de IgG e IgM anti Toxocara canis por EIE; los sueros positivos fueron confirmados por Western Blot. De los 182 niños estudiados, 122 resultaron seropositivos (67%), 28.8% no contaban con agua potable en su domicilio, 58.8% no tenían cloacas, 91.1% habían tenido contacto con perros y/o gatos, 30.0% tenían antecedentes de geofagia y 86.7% vivían sobre calles sin pavimento. La infección se presentó en forma asintomática en el 77.8% de los casos, como larva migrans ocular en el 6.7% y como larva migrans visceral en el 15.5 % de los casos. En 22 niños el seguimiento serológico post-tratamiento hasta los 18 meses mostró que la IgG se mantuvo estable en 10 casos, en 11 disminuyó pero manteniendo valores elevados y em uno aumentó. Hubo 19 casos con IgM positiva; 8 disminuyeron sus títulos, uno se mantuvo estable y 10 se negativizaron. Hubo un caso de reinfección. Estos resultados reafirman la importancia que las autoridades sanitárias deben asignar a esta infección, particularmente en las regiones carenciadas, en las que habitualmente no se reconoce a la toxocariosis como un problema relevante de salud pública.

A seroepidemiological survey for toxocariasis in apparently healthy residents in Gangwon-do, Korea.

We investigated the sero-prevalence of toxocariasis among healthy Korean adults in 1999. A total of 314 sera from normal inhabitants in Whachon-gun, Gangwondo, Korea was examined for specific antibody levels against excretory-secretory products of second stage larvae of Toxocara (TES). The presence of cross-reactions with other helminthiases such as cysticercosis, paragonimiasis, sparganosis or clonorchiasis was also checked by specific IgG ELISA. Sera showing positive reaction against TES were also tested by IgG immunoblot and by IgE ELISA. Out of 314 subjects, 16 was found to be positive by TES IgG ELISA and immunoblot, among whom 12 were also positive by TES IgE ELISA. Among the 16 seropositive samples, two sera showed positive reaction against Paragonimus and sparganum antigen, respectively. These results inferred that cross-reactions were negligible between toxocariasis and other helminthiases. Toxocariasis seroprevalence among Korean rural adults was detected to be approximately 5%.

Human toxocariasis and the visceral larva migrans syndrome: correlative immunopathology.

The surface of T. canis is now recognized as a dynamic structure which turns over quite rapidly and serves as a renewable source of large quantities of antigen(s). The major host responses to these antigens include a marked eosinophilia and hyperglobulinemia. Both of these responses are apparently ineffective at ridding the body of infective larvae. Both eosinphils and IgE antibodies are manifestations of the Th2 subset of T helper cells and the cytokines that they secrete. Further, there is reason to believe that the antigens released from T. canis larvae favor the induction of this cellular population. Finally, there is mounting evidence that the chronic production of parasite antigen and its continued stimulation of the host immune system with a concomitant production of eosinophils can lead to a permanent alteration of the normal organization of the cardiopulmonary system. In the absence of any well-documented drugs capable of killing infective larvae, it would seem that immunological intervention may offer the only way to minimize or neutralize this 'gift from man's best friend'. This chapter was not intended to be an exhaustive review of the literature pertaining to toxocariasis. Several other recent publications will hopefully fulfill the need for more detailed information on the biology of this organism and the clinical spectrum of the disease it produces [16, 138-140]. Finally, a MEDLARS search of the current medical literature should bring anyone up to speed in a very short time.

Persistence of specific antibody response in different experimental infection of mice with toxocara canis larvae

Estudou-se a cinetica de anticorpos anti-Toxocara em camundongos BALB/c infectados experimentalmente segundo tres esquemas: Grupo I (GI): 25 camundongos infectados com dose unica de 200 ovos embrionados de T. canis; grupo II (GII): 25 camundongos infectados com 150 ovos embrionados de T. canis, divididos em tres doses de 50 ovos, administrados no 1§, 5§ e 8§ dias; grupo III (GIII): 25 camundongos infectados com 150 ovos embrionados de T. canis, administrados em tres doses de 50 ovos no 1§, 14§ e 28§ dias...

MRI in cerebral toxocaral disease.

Toxocara canis, the common roundworm in the dog, can cause "visceral larva migrans" syndrome in humans, which may include generalised illness, eosinophilia, and symptoms arising from larval invasion of different organs. Of these, the clinically most important are liver, lungs, eyes and CNS. Involvement of the different parts of the CNS in human toxocaral disease has been described, but not the CT or MRI appearances of the cerebral lesions. In one case with a single focal epileptic fit, CT was described as normal. In the Toxocara canis case described, the cerebral lesions on MRI, before and after therapy, are shown.

A collaborative study on larval excretory/secretory antigens of Toxocara canis for the immunodiagnosis of human toxocariasis with ELISA.

Two batches of excretory/secretory (E/S) antigens from second stage larvae of Toxocara canis maintained in vitro were prepared independently in two different laboratories (Zürich and Basel) and analysed in order to obtain information for future efforts to standardize the enzyme-linked immunosorbent assay (ELISA) used for the serodiagnosis of human toxocariasis. SDS-PAGE and "Western-blotting" revealed at least 10 different antigenic components common to the two antigen preparations. However, distinct qualitative and quantitative differences among the two E/S-antigens were observed, since one antigen had a more complex composition than the other. Despite these differences, an accordance of serodiagnosis was obtained in 80% of 25 sera from patients with suspected Toxocara infection tested independently in two different ELISA systems (Basel and Zürich) with the corresponding E/S-antigens. The specificity was 93% as determined (BS-antigen, BS-ELISA) by testing 46 out of 3396 sera from patients with parasitologically proven extra-intestinal helminthic infections. Cross-reactions occurred mainly with sera from patients infected with filariae (5 from 13 cases) exhibiting very high extinction values in their homologous ELISA-system. The reproducibility (intra- and inter-test variations) of two ELISA systems using the corresponding E/S-antigens varied from 5-15%. The results demonstrate that T. canis E/S-antigens may well be applicable for standardization of the ELISA used for the serodiagnosis of human toxocariasis.

Meandering ocular toxocariasis.

Ocular toxocariasis affects the eye in three recognizable patterns: a peripheral inflammatory mass in a quiet eye, posterior pole granuloma, or painless endophthalmitis. The present case photographically documents a new form of ocular involvement, that of periodic intraretinal meandering, followed by quiescence with encapsulation, reemergence, and renewed migration. The similar behavior of the Toxocara canis larva in the animal model and the serologic confirmation of Toxocara canis by ELISA testing establishes the diagnosis in the present case. Photocoagulation is the treatment of choice.

Ocular toxocariasis: a review of the literature.

Ocular toxocariasis is invasion of the eye by the larvae of the Toxocara canis worm. Since its initial description 31 years ago, many cases involving all segments of the eye have been reported. Most age groups have been affected, although it is generally a childhood disease. Its distribution is worldwide. Several tests have been devised for its detection, but only recently with a degree of success. Many treatments have been proposed, but more evaluation is necessary. This review correlates historical and present knowledge concerning Toxocara. The history, pathophysiology, immunopathology, distribution, diagnosis, and treatment are discussed.

[Eosinophilia for years in a patient with antibodies directed against larva migrans visceralis (author's transl)]. / Jahrelange Eosinophilie bei positivem Antikörpernachweis auf Toxocara-canis-Larven.

A report is given on a 60 years old male patient with a varying eosinophilia and leucocytosis during a period of 8 years. The patient was suffering from coronary heart disease, which finally was the cause of death. Antibodies against larva migrans universalis were identified with the Ouchterlony technique as well as with the microprecipitation test for living toxocara larva twice within a period of 3 years. The post mortem examination showed severe sequelae of the coronary heart disease with callosities and aneurysma, whereas parasitic granulomas were missed. The question is discussed if the eosinophilia may be a sequela of the coronary heart disease or a result of an infection with larva migrans visceralis, in spite of the lack of a histological evidence.

Clinical, serologic, and epidemiologic characteristics of ocular toxocariasis.

The clinical, serologic, and epidemiologic characteristics of 17 cases of ocular toxocariasis (OT) were studied and compared with those of a control group of 15 cases of other ocular diseases whose differential diagnosis included retinoblastoma. The prevalence and mean titers of Toxocara antibody detected by enzyme-linked immunosorbent assay were greater (P less than 0.005) for patients with OT than for the control group, but not all clinically diagnosed OT cases had detectable antibody. The prevalence of pica was significantly greater in cases than in controls (P less than 0.05). Almost all case and control patients had a history of exposure to pet dogs and cats, but recent exposure to puppies (less than 3 months old) was significantly associated with Toxocara infection in this study group.

Toxocara-specific antibody in the serum and aqueous humor of a patient with presumed ocular and visceral toxocariasis.

Rarely have concurrent ocular and systemic toxocariasis been reported in the literature. We describe a patient with serologically proven visceral toxocariasis who had a granulomatous lesion in the iris, small rod-like lesions in the retina, and in whom Toxocara-specific antibodies were also demonstrated in the aqueous humor. Two older siblings of this patient also had demonstrable serum antibody to Toxocara. The ocular manifestations resolved rapidly with corticosteroid and thiabendazole therapy and the initial leucocytosis, hepatomegaly, and elevated IgM level were normal at 3.5 months. These changes might be attributed to either the thiabendazole and prednisone therapy or to the natural history of this disease.

Evaluation of serodiagnostic tests for visceral larva migrans.

Four serologic techniques for the diagnosis of visceral larva migrans caused by Toxocara canis, namely indirect hemagglutination (IHA), bentonite flocculation (BF), enzyme-linked immunosorbent assay (ELISA), and double diffusion in agar (Ouchterlony), were evaluated using sera sent to the Center for Disease Control from patients with a presumptive diagnosis of visceral larva migrans (VLM). Patients having 5-6 of the clinical or laboratory criteria for VLM were designated as cases while those with 0-2 criteria served as controls. The sensitivity of the ELISA was 78.3% compared to 18.2%, 25.8% and 65.2% for the IHA, BF, and Ouchterlony, respectively; the sepcificity of all four tests was greater than 92%. The predivtive value of positive test was greater than 85% for all tests except the IHA, while the predictive value of a negative test was greater than 85% only for the ELISA. The results of a ELISA were reproducible in different laboratories. Based on these findings, the ELISA using a larval antigen appears to be the serodiagnostic method of choice for VLM.

Naturally occurring human antiglobulins with specificity for E.

Human sera have been examined for antibodies with specific reactivity for gammaE using the tanned cell hemagglutination test. Cells tanned with three different gammaE myeloma proteins provided a reproducible test system. Inhibition of agglutination reactions by gammaE proteins, but not by gammaG, gammaA, gammaM, or gammaD confirmed the specificity of these reactions. 8.5% of 304 serial serum samples obtained from miscellaneous hospitalized patients showed clear-cut anti-gamma-globulins with specificity for gammaE. In most of these instances no definite clinical history of concomitant allergic disorders could be obtained. 53% of 73 patients with well-established allergic disorders (hay fever, extrinsic asthma) showed serum anti-gamma-globulins with reactivity for gammaE. Some patients studied before and after desensitization to Bermuda grass allergen showed an increase in titer or a conversion from negative to positive reactions for anti-gammaE antibodies following several month courses of progressive desensitization. Gradient and gel filtration studies indicated that anti-gammaE globulins were 19S gammaM in all instances. No clear correlation was noted between quantitative serum gammaE levels and titer of anti-gammaE antibodies.19S serum fractions with anti-gammaE antibody activity did not release histamine from normal human peripheral blood leukocytes, whereas specific rabbit anti-gammaE antisera consistently induced leukocytic histamine release. Moreover, macroglobulin fractions with anti-gammaE activity did not block allergen-specific leukocyte histamine release induced by in vitro leukocyte challenge with allergens such as Bermuda grass and leukocytes from allergic donors. In some instances 19S human serum fractions with anti-gammaE activity appeared to potentiate histamine release when incubated concomitantly with specific allergen and leukocytes from allergic individuals.