Hematological alterations caused by high intensity of L3 larvae of Contracaecum sp Railliet & Henry, 1912 (Nematoda, Anisakidae) in the stomach of Hoplias malabaricus in lakes in Pirassununga, São Paulo.

The objective of the present study was to discover the hematological indices of Hoplias malabaricus infected by larvae of Contracaecum sp. A total of 105 fish were collected from two lakes located in the municipal district of Pirassununga, São Paulo, Brazil, of which 95.2% were infested by L3 larvae of Contracaecum sp., with an average intensity of 348.7 ± 231.55 helminths/fish. A total of 76 fish were analyzed to establish hematological parameters. Following analysis, it was found that there was a significant difference between hematological parameters Hct, erythrocytes (Er), thrombocytopenia (p = 0.01), and Hb (p = 0.02) in lakes A and B, while there was no significant difference for leukocyte (p = 0.68), mean corpuscular volume, mean corpuscular hemoglobin (p = 0.06), and mean corpuscular hemoglobin concentration (p = 0.47). The intensity of infection correlated negatively with Er (r(s) = -0.42; p < 0.01) and positively for (r(s) = 0.48; p < 0.01). The high intensity of infection by Contracaecum sp. in H. malabaricus did not affect the health of the fish despite causing alterations in hematological variables.

A novel approach based on antigen, antibody and immune complex detection in bronchoalveolar lavage fluid samples from rats experimentally infected with Strongyloides venezuelensis.

This study was performed in order to develop a novel approach based on antigen, antibody and immune complex detection by enzyme-linked immunosorbent assay (ELISA) in bronchoalveolar lavage fluid (BALF) samples. For that purpose Wistar rats immunosuppressed or not were experimentally infected with Strongyloides venezuelensis. The microtiter plates were coated with alkaline parasite extract for antibody detection and with IgG anti-S. venezuelensis for antigen and immune complex detection. The immune serum was able to detect 1.56 µg/mL of L3 antigens in BALF samples. ELISA sensitivity was 96.6%, 71.6% and 91.6% for antigen, antibody and immune complex, respectively, and the specificity was 100% for all methods. Antigen detection in BALF samples showed to be a good approach for evaluating the kinetics of infection in non immunosuppressed or immunosuppressed rats. IgG was detected in non immunosuppressed rats from day 8 p.i. and in immunosuppressed rats from day 2 p.i. Moreover, immune complex was detected during the entire kinetic for both groups. In conclusion, association of antigen, antibody and immune complex detection in BALF samples seems to be an alternative approach for early strongyloidiasis diagnosis particularly in immunosuppressed individuals.

Dexamethasone reduces bronchial wall remodeling during pulmonary migration of Strongyloides venezuelensis larvae in rats.

Strongyloidiasis is an intestinal parasitosis with an obligatory pulmonary cycle. A Th2-type immune response is induced and amplifies the cellular response through the secretion of inflammatory mediators. Although this response has been described as being similar to asthma, airway remodeling during pulmonary migration of larvae has not yet been established. The aim of this study was to identify the occurrence of airway remodeling during Strongyloides venezuelensis (S. v.) infection and to determine the ability of dexamethasone treatment to interfere with the mechanisms involved in this process. Rats were inoculated with 9,000 S. v. larvae, treated with dexamethasone (2 mg/kg) and killed at 1, 3, 5, 7, 14 and 21 days. Morphological and morphometric analyzes with routine stains and immunohistochemistry were conducted, and some inflammatory mediators were evaluated using ELISA. Goblet cell hyperplasia and increased bronchiolar thickness, characterized by edema, neovascularization, inflammatory infiltrate, collagen deposition and enlargement of the smooth muscle cell layer were observed. VEGF, IL1-ß and IL-4 levels were elevated throughout the course of the infection. The morphological findings and the immunomodulatory response to the infection were drastically reduced in dexamethasone-treated rats. The pulmonary migration of S. venezuelensis larvae produced a transitory, but significant amount of airway remodeling with a slight residual bronchiolar fibrosis. The exact mechanisms involved in this process require further study.

Antigen, antibody and immune complex detection in serum samples from rats experimentally infected with Strongyloides venezuelensis.

In order to establish an antigen, antibody and immune complex detection by enzyme-linked immunosorbent assay (ELISA) in serum samples, normal or immunocompromised Wistar rats experimentally infected with Strongyloides venezuelensis were used. The microtitre plates were coated with IgG anti-S. venezuelensis for antigen and immune complex detection and with alkaline parasite extract for antibody detection. Analysis revealed at least 12.5 µg/mL of S. venezuelensis specific antigens in serum samples. Assay for antigen detection was not a good approach for evaluating infection in normal or immunocompromised rats. In normal rats IgG specific for S. venezuelensis was preferentially detected during the first 13 days post-infection (p.i.) and immune complex detection was significantly reduced in 21 p.i. day. On the other hand, in immunocompromised rats, IgG and immune complex were detected during the entire kinetic (5, 8, 13 and 21 p.i). These results suggest that immune complex screening seems to be an alternative for early strongyloidiasis diagnosis in immunocompromised individuals.

Marmosa paraguayana (Marsupialia: Didelphidae) as a new host for Gracilioxyuris agilisis (Nematoda: Oxyuridae) in Brazil.

Didelphids (Didelphimorphia: Didelphidae) are a large and well-studied group of Neotropical marsupials. Although knowledge of the parasitic fauna of didelphids is still scarce, recent work has suggested that Neotropical marsupials are often hosts of pinworms. Here, we isolated oxyurid nematodes from fecal samples of Marmosa paraguayana (Marsupialia: Didelphidae) and provide a general description and measurements for male and female specimens. We concluded these specimens can be assigned to Gracilioxyuris agilisis (Ascaridida: Oxyuridae), an oxyurid recently described as a parasite of the didelphid Gracilinanus agilis (Didelphimorphia: Didelphidae). The finding of G. agilisis in a different, albeit closely related, host species strengthens the previous notion of a close association between pinworms and didelphids and contributes to the knowledge of the helminthic fauna of didelphid marsupials.

IgG avidity in differential serodiagnosis of human strongyloidiasis active infection.

IgG avidity assays have been developed for several parasitic diseases although there are no researches focused in strongyloidiasis diagnosis. Definitive diagnosis of strongyloidiasis is based on the presence of Strongyloides larvae in stool, but majority of cases involve low and irregular larval output. While limitations of serological assays for strongyloidiasis are well known, characteristics of persons who are misdiagnosed based on negative coproparasitological tests have been little explored. The aim of the present study was to evaluate the use of IgG avidity to detect patients with active strongyloidiasis and to characterize sources of disagreement between serology and coproparasitology. A total of 80 serum samples was analyzed, 40 from patients with Strongyloides larvae in stool (G1) and 40 from individuals with negative coproparasitology, but positive serology (G2). Serum samples were analyzed in an indirect IgG avidity ELISA using urea 6M in serial double dilutions from 1:80 to 1:2560. Avidity index (AI) was calculated to each serum dilution and analyzed as screening AI (serum dilution of 1:160) or mean AI of different serum dilutions that had a positive result. Statistical analyzes were performed by Mann-Whitney's (U) and Fisher's exact tests. At screening dilution, median of AI was 68% in G1 and 88% in G2 (P<0.0001), whereas median of mean AI in G1 was 72% and in G2 94% (P<0.0001), but there was no significant differences between both AI in each patient group. A cut off value established at AI of 75% demonstrated a significant difference between groups, with G1 sera showing AI<75% and G2 sera with AI>75% (P<0.0001). In conclusion, IgG avidity assays may distinguish active infection with Strongyloides stercoralis from suspect or serologically false positive cases.

Dexamethasone effects in the Strongyloides venezuelensis infection in a murine model.

The aim of this study was to investigate the immunomodulatory effects of glucocorticoids on the immune response to Strongyloides venezuelensis in mice. Balb/c mice were infected with S. venezuelensis and treated with Dexamethasone (Dexa) or vehicle. Dexa treatment increased circulating blood neutrophil numbers and inhibited eosinophil and mononuclear cell accumulation in the blood, bronchoalveolar, and peritoneal fluid compared with control animals. Moreover, Dexa decreased tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-3 (IL-3), IL-4, IL-5, IL-10, and IL-12 production in the lungs and circulating immunoglobulin G1 (IgG1), IgG2a, and IgE antibody levels while increasing the overall parasite burden in the feces and intestine. Dexa treatment enhanced the fertility of female nematodes relative to untreated and infected mice. In summary, the alterations in the immune response induced by Dexa resulted in a blunted, aberrant immune response associated with increased parasite burden. This phenomenon is similar to that observed in S. stercoralis-infected humans who are taking immunosuppressive or antiinflammatory drugs, including corticosteroids.

Detergent fraction of heterologous antigen to detect IgA and IgG in strongyloidiasis using saliva and serum paired samples.

Strongyloides stercoralis causes chronic asymptomatic infections in immunocompetent human hosts and systemic invasion in immunocompromised patients, developing into a fatal hyperinfection syndrome. IgA and IgG detection in saliva and serum paired samples were tested using total saline extract from Strongyloides venezuelensis (SE(*)) and its detergent phase (D) extracted with Triton X-114. Saliva and serum paired samples were obtained from: 25 patients with confirmed strongyloidiasis; 25 patients with other parasitoses and 20 from apparently healthy individuals. Sensitivity, specificity, diagnostic efficiency, positive and negative predictive values and likelihood ratio were calculated at the optimum point of reaction. Using D phase sensitivity and specificity to detect IgA in saliva were 76.0% and 88.9% and in serum 80.0% and 86.7%, respectively. To detect IgG, D phase showed sensitivity and specificity of 88.0% and 88.9% in saliva and 88.0% and 84.4% in serum, respectively. D phase proved to be specific and efficient and could be utilized as an alternative antigen for IgA and IgG detection in saliva and serum samples for strongyloidiasis diagnosis.

A new faecal antigen detection system for Strongyloides venezuelensis diagnosis in immunosuppressed rats.

This study was performed with the objective of developing innovative procedures for the diagnosis of strongyloidiasis. Enzyme-linked immunosorbent assay (ELISA) was conducted to detect coproantigen in the faecal samples of normal and of immunosuppressed rats using an anti-L3 polyclonal antibody produced in rabbits. Analysis revealed the kinetics of egg shedding in the non-immunosuppressed and immunosuppressed rats infected with S. venezuelensis. Further analysis verified the ability of the immune serum to detect L3 antigens in faecal samples from infected animals. The number of eggs recovered in the faeces at 8 days p.i was significantly higher for both groups. Immunosuppressed animals eliminated increased quantities of eggs. The immune serum was able to detect 0.39 microg/ml of L3 antigens. The antigen recognition in the immunosuppressed group was anticipated on the 8th day p.i. In conclusion, these results may represent a first step in the development of a rapid coproantigen detection kit for strongyloidiasis.

Cyclooxygenase-derived mediators regulate the immunological control of Strongyloides venezuelensis infection.

The aim of this study was to define the immunoregulatory role of prostaglandins in a mouse model of Strongyloides venezuelensis infection. Strongyloides venezuelensis induced an increase of eosinophils and mononuclear cells in the blood, peritoneal cavity fluid, and bronchoalveolar lavage fluid. Treatment with the dual cyclooxygenase (COX-1/-2) inhibitors indomethacin and ibuprofen, and the COX-2-selective inhibitor celecoxib partially blocked these cellular responses and was associated with enhanced numbers of infective larvae in the lung and adult worms in the duodenum. However, the drugs did not interfere with worm fertility. Cyclooxygenase inhibitors also inhibited the production of the T-helper type 2 (Th2) mediators IL-5, IgG1, and IgE, while indomethacin alone also inhibited IL-4, IL-10, and IgG2a. Cyclooxygenase inhibitors tended to enhance the Th1 mediators IL-12 and IFN-gamma. This shift away from Th2 immunity in cyclooxygenase inhibitor-treated mice correlated with reduced prostaglandin E(2) (PGE(2)) production in infected duodenal tissue. As PGE(2) is a well-characterized driver of Th2 immunity, we speculate that reduced production of this lipid might be involved in the shift toward a Th1 phenotype, favoring parasitism by S. venezuelensis. These findings provide new evidence that cyclooxygenase-derived lipids play a role in regulating host defenses against Strongyloides, and support the exploration of eicosanoid signaling for identifying novel preventive and therapeutic modalities against these infections.

Counterregulation of Th2 immunity by interleukin 12 reduces host defenses against Strongyloides venezuelensis infection.

The aim of this study was to investigate the role of interleukin 12 (IL-12) during Strongyloides venezuelensis infection. IL-12(-/-) and wild-type C57BL/6 mice were subcutaneously infected with 1500 larvae of S. venezuelensis. On days 7, 14, and 21 post-infection, we determined eosinophil and mononuclear cell numbers in the blood and broncoalveolar lavage fluid (BALF), Th2 cytokine secretion in the lung parenchyma, and serum antibody levels. The numbers of eggs in the feces and worm parasites in the duodena were also quantified. The eosinophil and mononuclear cell counts and the concentrations of IL-3, IL-5, IL-10, IL-13, and IgG1 and IgE antibodies increased significantly in infected IL-12(-/-) and wild-type mice as compared with uninfected controls. However, the number of eosinophils and mononuclear cells in the blood and BALF and the Th2 cytokine levels in the lungs of infected IL-12(-/-) mice were greater than in infected wild-type C57BL/6 mice. In addition, serum IgE and IgG1 levels were also significantly enhanced in the infected mice lacking IL-12. Meanwhile, parasite burden and fecal egg counts were significantly decreased in infected IL-12(-/-) mice. Together, our results showed that the absence of IL-12 upregulates the Th2 immune response, which is important for control of S. venezuelensis infection.

The ubiquitin-proteasome system in Strongyloididae. Biochemical evidence for developmentally regulated proteolysis in Strongyloides venezuelensis.

Nematode parasites from the genus Strongyloides spp. are important pathogens of the intestinal mucosa of animals and humans. Their complex life cycles involve alternating developmental adaptations between larvae stages and the adult parthenogenetic female. Here, we report, primarily through homology-based searching, the existence of the major components of the ubiquitin-proteasome system in this genus, using the available EST data from S. ratti, S. stercoralis, and Parastrongyloides trichosuri. In this study, S. venezuelensis was used as our model organism for detection of proteasome activity and ubiquitinated substrates in cytosolic preparations from the L3 larvae and the adult female. Marked differences in proteasome capabilities were found when these two stages were compared. A preference for degradation of chymotryptic synthetic peptides was found in both stages with the adult exhibiting a higher rate of hydrolysis compared to the larvae. Due to the high evolutionary conservation of proteasome alpha subunits, an anti-human proteasome antibody was able to recognize proteasome subunits in these preparations by Western blotting, supporting the proposal that the activity of the ubiqutin-proteasome system is developmentally regulated in this nematode.

Major histocompatibility complex (MHC) class II but not MHC class I molecules are required for efficient control of Strongyloides venezuelensis infection in mice.

Strongyloides stercoralis is an intestinal nematode capable of chronic, persistent infection and hyperinfection of the host; this can lead to dissemination, mainly in immunosuppressive states, in which the infection can become severe and result in the death of the host. In this study, we investigated the immune response against Strongyloides venezuelensis infection in major histocompatibility complex (MHC) class I or class II deficient mice. We found that MHC II(-/-) animals were more susceptible to S. venezuelensis infection as a result of the presence of an elevated number of eggs in the faeces and a delay in the elimination of adult worms compared with wild-type (WT) and MHC I(-/-) mice. Histopathological analysis revealed that MHC II(-/-) mice had a mild inflammatory infiltration in the small intestine with a reduction in tissue eosinophilia. These mice also presented a significantly lower frequency of eosinophils and mononuclear cells in the blood, together with reduced T helper type 2 (Th2) cytokines in small intestine homogenates and sera compared with WT and MHC I(-/-) animals. Additionally, levels of parasite-specific immunoglobulin M (IgM), IgA, IgE, total IgG and IgG1 were also significantly reduced in the sera of MHC II(-/-) infected mice, while a non-significant increase in the level of IgG2a was found in comparison to WT or MHC I(-/-) infected mice. Together, these data demonstrate that expression of MHC class II but not class I molecules is required to induce a predominantly Th2 response and to achieve efficient control of S. venezuelensis infection in mice.

Interference of dexamethasone in the pulmonary cycle of Strongyloides venezuelensis in rats.

The aim of this study was to investigate the interference of a daily treatment of dexamethasone in the pulmonary cycle of Strongyloides venezuelensis infection in rats. Three principal effects were found: 1) increased alveolar hemorrhagic inflammation provoked by the passage of larvae into alveolar spaces; 2) significant decrease of eosinophil and mast cell migration to the axial septum of the lungs; and 3) impaired formation of the reticular fiber network, interfering with granuloma organization. This study showed that the use of drugs with immunomodulatory actions, such as dexamethasone, in addition to interfering with the morbidity from the pulmonary cycle of S. venezuelensis infection, may contribute to showing the mechanisms involved in its pathogenesis.

Immunoparasitological diagnosis of Strongyloides stercoralis in garbage collectors in Uberlândia, MQ Brazil

The objective of this study was to determine the presence of Strongyloides stercoralis in urban garbage collectors through the use of immunological and parasitological methods. A total of 92 individuals were evaluated from August, 1997, to June, 1998. For the parasitological diagnosis Baermann and Lutz' methods were applied. The immunological diagnosis involved the indirect fluorescence antibody test (IFAT) and the enzyme-linked immunosorbent assay (ELISA) to detect specific IgG antibodies. Of the 92 workers examined, six (6.5 percent) were infected with larvae of S. stercoralis. The IFAT detected 19 (16.3 percent) and the ELISA 17 (18.5 percent) positive serum samples. The differences between the results of parasitological and immunological methods were statistically significant (p<0.05). These results demonstrate that there is a need to improve the health conditions of this category of city employees.

Seroepidemiological aspects of human Strongyloides stercoralis infections in Chile.

To determine the frequency of Strongyloides stercoralis antibodies by means of the enzyme linked immunosorbent assay (ELISA) in Chile, in 2001-2003, 675 blood samples of patients of two psychiatric hospitals and 172 of healthy individuals (doctors, nurses and paramedicals) of these institutions, and 1,200 serum samples of blood donors of Northern region (Arica and Antofagasta), Central region (Valparaiso and Santiago) and Southern region (La Union) were collected. ELISA showed positivity of 12.1% in psychiatric hospitalized patients, none (0%) in the health personnel and 0.25% in blood donors (p < 0.05). Only in blood donors of Arica (1%) and La Union (0.5%) the ELISA test was positive suggesting that strongyloidiasis is focalized in determinate zones of the country. In Chile, human infections by S. stercoralis are endemic with very low frequency in apparently healthy individuals and high prevalence in risk groups such as the mentally ill hospitalized patients.

Seroepidemiological aspects of human Strongyloides stercoralis infections in Chile

To determine the frequency of Strongyloides stercoralis antibodies by means of the enzyme linked immunosorbent assay (ELISA) in Chile, in 2001-2003, 675 blood samples of patients of two psychiatric hospitals and 172 of healthy individuals (doctors, nurses and paramedicals) of these institutions, and 1,200 serum samples of blood donors of Northern region (Arica and Antofagasta), Central region (Valparaiso and Santiago) and Southern region (La Union) were collected. ELISA showed positivity of 12.1 percent in psychiatric hospitalized patients, none (0 percent) in the health personnel and 0.25 percent in blood donors (p < 0.05). Only in blood donors of Arica (1 percent) and La Union (0.5 percent) the ELISA test was positive suggesting that strongyloidiasis is focalized in determinate zones of the country. In Chile, human infections by S. stercoralis are endemic with very low frequency in apparently healthy individuals and high prevalence in risk groups such as the mentally ill hospitalized patients.

Entre os anos de 2001-2003 foram coletadas amostras de sangue de 675 pacientes de dois hospitais psiquiátricos da região central do Chile, 172 de indivíduos sadios (médicos, enfermeiros e paramédicos) destas instituições e 1200 de doadores de sangue de cidades das regiões norte (Arica e Antofagasta), central (Valparaiso e Santiago) e sul (La Union) para determinar a frequência de anticorpos anti Strongyloides stercoralis mediante a reação de enzyme linked immunosorbent assay (ELISA). Foram observadas soropositividade de 12.1 por cento em pacientes de hospitais psiquiátricos e de 0,25 por cento em doadores de sangue (p < 0.05). Todas as amostras dos indivíduos sadios foram não reagentes. Entre os doadores de sangue a soropositividade ocorreu somente nos indivíduos de Arica (1,0 por cento) e La Union (0,5 por cento) sugerindo que a estrongiloidíase poderia estar localizada em determinadas áreas geográficas do país. Conclui-se que no Chile as infecções por S. stercoralis seriam endêmicas, de baixa freqüência e afetando especialmente grupos de risco como os pacientes psiquiátricos.

Leukotrienes play a role in the control of parasite burden in murine strongyloidiasis.

It is clear that leukotrienes mediate inflammatory response; new aspects of leukotriene function have recently been described. In this study, we demonstrate that leukotrienes are key chemical mediators in the control of parasite burdens in mice infected with Strongyloides venezuelensis. High leukotriene levels were detected in the lungs and small intestines of Swiss mice. In infected Swiss mice treated with MK886, a leukotriene synthesis inhibitor, numbers of adult worms, and eggs/g/feces were greater than in infected-only animals. The MK886 treatment inhibited leukotriene B(4) production in the lungs and small intestines, albeit on different postinfection days. Similarly, parasite burdens and eggs/g/feces were greater in 5-lipoxygenase(-/-) mice than in wild-type animals. These observation were confirmed by histopathological study of the duodena. We subsequently observed significant lower numbers of eosinophils and mononuclear cells in the blood, peritoneal cavity fluid, and bronchoalveolar lavage fluid of Swiss mice treated with MK886. In the lung parenchyma of infected animals, MK886 significantly inhibited synthesis of IL-5 at the beginning of infection, whereas levels of IL-12 increased progressively throughout the postinfection period. However, levels of leukotriene C(4), PGE(2), TNF-alpha, IL-3, IL-4, IFN-gamma, and IL-10 were comparable between the treated and untreated groups. Nevertheless, IgE and IgG1 (but not IgG2a) synthesis was also significantly inhibited by MK886 administration. Therefore, in S. venezuelensis-infected mice, adult worm and egg burdens are leukotriene dependent. These findings indicate potential immunostimulatory strategies involving leukotriene administration, and may serve as an alert to physicians treating Strongyloides stercoralis-infected patients presenting asthma-like symptoms because use of 5-lipoxygenase inhibitors may worsen the infection.

Exposure to larva migrans syndromes in squares and public parks of cities in Chile.

Between November 2001 and December 2002, 600 dog fecal samples were collected in main squares and public parks of 13 cities in Chile, from the extreme north to the extreme south of the country. The samples were processed in the laboratory by centrifugal sedimentation and the Harada-Mori methods. T. canis eggs were found in 12 cities. Detection rates ranged from 1.9 to 12.5% with an average of 5.2%. Seven percent of the samples had eggs and 9.5% had rhabditoid and/or filariform larvae of Ancylostomatidae. Strongyloides stercoralis were not found. Squares and public parks in Chile pose a potential risk of exposure to visceral, ocular, and/or cutaneous larva migrans syndromes.

Exposure to larva migrans syndromes in squares and public parks of cities in Chile

Entre novembro de 2001 e dezembro de 2002, 600 amostras de fezes de cão foram coletadas nas principais praças e parques públicos de 13 cidades do Chile, localizadas nas regiões norte ao estremo sul da nação. No laboratório, as amostras foram processadas mediante os métodos de sedimentação por centrifugação e de Harada-Mori. Ovos de Toxocara canis foram encontrados em 12 cidades em freqüências que variaram entre 1,9 a 12,5 por cento por cidade, com média de 5,2 por cento. Sete por cento das amostras apresentaram ovos e 9,5 por cento larvas rabditóides ou filarióides de Ancylostomatidae. Strongyloides stercoralis não foi encontrado nas amostras estudadas. Praças e parques públicos do Chile apresentam riscos potenciais para aquisição de larva migrans visceral, ocular ou cutânea.