Evaluating host-parasite co-adaptation relationships involving Angiostrongylus costaricensis.

Angiostrongylus costaricensis is a parasite that infects rodents, including the wild cotton rat Sigmodon hispidus and pygmy rice rats Oligoryzomys spp., among others. However, urban Rattus norvegicus and Mus musculus have not been identified as important hosts of A. costaricensis. In this study, Swiss mice (SW), Wistar R. norvegicus (RN), wild Oligoryzomys nigripes (ON) and a local strain of M. musculus (RGS) were experimentally infected with A. costaricensis. Survival, elimination of L1 (total sum per group, A0), and the number of adult worms recovered divided by the dose of each L3 inoculum (yield ratio, YR) were examined for each group after a 40-day post-infection period. The survival rates, A0 and YR values were: 27%, 207,589 and 0.42 for the SW group; 81%, 8691 and 0.01 for the RN group; and 63.6%, 26,560 and 0.16 for the RGS group, respectively, in each case. The survival rate for the ON group was 100% and the A0 value was 847,050. A YR was not calculated for the ON group since the ON group was maintained up to 565 days post-infection (pi) to examine long-term mortality. At 500 days pi (16 months), 50% of the ON group had died, while one animal (10%) survived 595 days pi (20 months). Taken together, these data indicate that A. costaricensis has undergone a greater degree of adaptation to the wild rodent, O. nigripes, than to R. norvegicus or a local M. musculus strain. In addition, titre curve (A0) modelling of adaptation status proved to be useful in evaluating A. costaricensis-rodent interactions.

Invasive slug Meghimatium pictum (Stoliczka, 1873) infected by Angiostrongylus costaricensis Morera & Céspedes, 1971, and the possible risk of human infection associated with grape consumption.

Many molluscs may be infected with angiostrongylid larvae. Following the histopathological diagnosis of abdominal angiostrongyliasis in a grape farmer from southern Brazil, molluscs in the area were investigated. During a nocturnal search, 245 specimens of slugs were collected and identified as the invasive Chinese slug Meghimatium pictum. Angiostrongylus costaricensis worms were recovered from mice that were experimentally infected with larvae obtained from 11 (4.5%) of the molluscs. This study presents the first report of M. pictum being identified as an intermediate host for A. costaricensis. Most of the slugs were collected from grape plants, which suggests that transmission may be associated with grape consumption.

Effect of different helminth extracts on the development of asthma in mice: The influence of early-life exposure and the role of IL-10 response.

It is not currently clear whether different parasites have distinct effects on the airway inflammatory response in asthma and whether exposure in early life to helminths have a stronger impact in a potential inhibitory effect on asthma. The aim of this study is to evaluate the effect of exposure to different helminth extracts on the development of allergic pulmonary response in mice, including early-life exposure. Different helminth extracts (Angiostrongylus costaricensis, Angiostrongylus cantonensis and Ascaris lumbricoides) were studied in female adult BALB/c and C57BL/6 IL-10-deficient mice in a protocol of murine asthma, injected intraperitoneally in different periods of exposure (early, pre-sensitization and post-sensitization). Cell counts in bronchoalveolar lavage (BAL), eosinophil peroxidase (EPO) from lung tissue, cytokine levels from BAL/spleen cell cultures, and lung histology were analyzed. Airway cellular influx induced by OVA was significantly inhibited by extracts of A. cantonensis and A. lumbricoides. Extracts of A. lumbricoides and A. costaricensis led to a significant reduction of IL-5 in BAL (p < 0.001). Only the exposure to A. lumbricoides led to an increased production of IL-10 in the lungs (p < 0.001). In IL-10-deficient mice exposed to A. costaricensis pre-sensitization, eosinophil counts and IL-5 levels in BAL and EPO in lung tissue were significantly reduced. In the early exposure to A. cantonensis, lung inflammation was clearly inhibited. In conclusion, different helminth extracts inhibit allergic lung inflammation in mice. IL-10 may not play a central role in some helminth-host interactions. Early exposure to helminth extracts could be a potential strategy to explore primary prevention in asthma.

Helminth extracts inhibit eosinophilic inflammation in a murine model of allergic rhinitis

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Abdominal angiostrongyliasis in rodent experimental infection: evidence for systemic circulation of first stage larvae.

Eggs of Angiostrongylus costaricensis embrionate and hatch in the course of their migration in the intestinal wall, and first stage larvae (L1) are released in feces. Aiming to investigate the possible systemic circulation of L1, we inoculated mice and, four weeks later, examined their peritoneal cavities and several organs for the presence of L1. A total of 65 larvae were found in extra-intestinal organs (kidney, heart, spleen, liver, lungs). No larvae were found in blood or in the peritoneal cavity. 320 and 578 L1 were found in intestinal wall and intestinal contents, respectively. The experiment was repeated and it confirmed that the metastrongylid larvae found in several organs were larval stages of A. costaricensis. Ten Oligoryzomis sp. rodents, a natural host for A. costaricensis, were also infected and in three animals L1 could be recovered from several organs as well as from bronchoalveolar lavage (BAL) in one of them. These data indicate that systemic circulation and bronchial elimination of L1 may represent an alternative route for release of L1 of A. costaricensis into the environment for transmission to the intermediate host.

Infection of BALB/c mice with Angiostrongylus costaricensis decreases pulmonary inflammatory response to ovalbumin.

SUMMARY The prevalence of asthma in developing countries is lower than in developed countries. Viral, bacterial and parasitic infections may be associated with this discrepancy. The relationship between parasitic infection and asthma prevalence is not clear. Previous controversial data have demonstrated that parasitic infection may either predispose or protect against the development of asthma. The aim of this study is to determine whether infection with Angiostrongylus costaricensis (A. costaricensis) decreases inflammatory lung response to ovalbumin (OVA) in mice. Seven BALB/c mice were infected with A. costaricensis by orogastric gavage (10 larvae/mouse) on day (D) 0. The mice were immunized against OVA by intraperitoneal injection on D 5 and D 12 and received an intranasal OVA challenge (40 micro L) on D 15 and D 17. On D 19 bronchoalveolar lavage (BAL) was performed. Six BALB/c mice (control group) were immunized with OVA using the same protocol, but were not infected with A. costaricensis. Interleukin (IL)-1beta and IL-6 levels were measured in the BAL fluid by using commercial ELISA assays. Total cell counts and differential cell counts were performed in the BAL fluid samples. The group infected with A. costaricensis had lower total cell count in the BAL fluid when compared with the control group (0.11 x 10(6)cells/mL and 0.3 x 10(6)cells/mL, respectively; P = 0.013). BAL fluid IL-1beta levels in the infected group were significantly lower than in the control group (P = 0.008). IL-6 levels in BAL fluid were not different between the groups studied. We conclude that Angiostrongylus costaricensis infection in mice decreases pulmonary inflammatory response to OVA.

Long-term observations on mouse strains experimentally infected with Angiostrongylus costaricensis.

BALB/c and C57BL/6 mice were experimentally infected with Angiostrongylus costaricensis and the parasitic parameters and antibody response during the acute and chronic phases of infection were analyzed. Following administration of six third-stage larvae (L3), there was no significant difference in the mean worm recovery or mean larval output. Coinciding with the maturation of worms in infected animals and with the egg output in mesenteric arteries, a strong increase in the humoral immune response was observed in both mouse strains. This response was characterized by a hypergammaglobulinemia, with a predominance of IgA and IgG1 during the acute phase of infection, and IgG1 and total IgE during the patent and post-patent periods. Significantly higher levels of IgM, IgG and IgG1 were found in BALB/c mice compared with C57BL/6 mice. On the other hand, a significantly higher concentration of IgA was detected at 6 and 7 weeks post-infection in C57BL/6 mice compared with BALB/c mice. Specific IgE could not be detected in any of the mouse strains. Our results suggest that immunoglobulins, mainly IgG1, contribute to the outcome of a primary A. costaricensis infection with respect to the period of patency and to mortality during the chronic phase.

Angiostrongylus costaricensis infection in C57BL/6 mice: MHC-II deficiency results in increased larval elimination but unaltered mortality.

During experimental Angiostrongylus costaricensis infections in several inbred mouse strains, genetic factors as well as different cytokine secretion patterns have recently been shown to play a role in the outcome of infection in terms of morbidity and mortality, e.g. BALB/c mice show a high and C57BL/6 mice a low mortality during the acute phase of infection. In this study, C57BL/6 MHC-II knockout mice infected with A. costaricensis did not show increased mortality during the acute phase of infection when compared with wild-type mice. Furthermore, MHC-II knockout mice showed a strongly diminished parasite-specific humoral and cellular immune response, which can be explained by the nearly complete lack of CD4+ T cells in the periphery. This defect in MHC-II genes, the lack of CD4+ T cells, and the resulting cellular and humoral unresponsiveness resulted in a three times higher output of first-stage larvae in feces compared with wild-type animals. The results indicate that during experimental A. costaricensis infection a parasite-specific immune response, directed via MHC-II molecules and CD4+ T cells, is not essential for the survival of C57BL/6 mice during the acute phase of infection, whereas the elimination of first-stage larvae seems to be regulated by a MHC-II- and CD4+ T-cell-dependent mechanism.

Interleukin-5 transgenic mice show augmented resistance to Angiostrongylus costaricensis infection.

To determine the possible role of eosinophils in Angiostrongylus costaricensis infection, both interleukin-5 (IL-5) transgenic (Tg) and non-transgenic (non-Tg) C3H/HeN mice were infected with A. costaricensis third-stage larvae. IL-5 Tg mice demonstrated greater resistance than non-Tg mice to A. costaricensis, as shown by lower adult worm recovery, smaller adults, fewer eggs in the intestinal wall and fewer larvae passed in the feces. Both mice showed similar antigen-specific IgA and IgGI antibody responses, although IgA was more prominent than IgG1. Egg deposition and inflammatory responses in the intestinal walls were milder in IL-5 Tg mice than in non-Tg mice. The eggs with developed larvae, deposited in the intestinal walls of IL-5 Tg mice, were surrounded by numerous degranulating eosinophils and sometimes with Splendore-Hoeppli deposits. The data suggest that eosinophils are involved in the resistance of the mouse during primary infection with A. costaricensis.

Cellular immune responses and cytokine production in BALB/c and C57BL/6 mice during the acute phase of Angiostrongylus costaricensis infection.

In our experimental study we were able to show that the contrasting outcome of Angiostrongylus costaricensis infection in C57BL/6 and BALB/c mice, in respect of morbidity and mortality, can be explained by divergent cellular immune responses and a different cytokine pattern in each strain. In BALB/c mice (i.e. those with high mortality), the initial high proliferation of ConA or LPS stimulated spleen cells dropped to very low levels after 2 weeks post-infection (p.i.), whereas in C57BL/6 mice (i.e. those with low mortality), only a minor reduction in lymphoproliferative responses after mitogenic stimulation was observed. The specific proliferation of spleen cells after stimulation with A. costaricensis adult worm antigen remained low in BALB/c mice throughout the experiment, but showed an augmented proliferation in C57BL/6 mice, especially from 2 weeks p.i. onwards. The mitogen-induced production of Th2-type cytokines (IL-4, IL-5, IL-6, IL-10) in spleen cell cultures remained low in BALB/c mice until 4 weeks p.i., but production of Th1-type cytokines (IL-2, IFN-gamma) was highly elevated at 14 and 28 days p.i. In C57BL/6 mice, an upregulated and balanced production of both Th1- and Th2-type cytokines was measured during the course of infection. In summary, a polarization of the immune response towards cellular hyporesponsiveness and a predominantly Th1 cytokine profile was observed in A. costaricensis infected BALB/c mice, which may contribute to pathogenesis and increased morbidity.

Detection of the acute phase of abdominal angiostrongyliasis with a parasite-specific IgG enzyme linked immunosorbent assay.

Angiostrongylus costaricensis may cause intestinal lesions of varied severity when it accidentally infects man in Central and South America. First-stage larvae have never been detected in stools. Therefore, a parasite-specific IgG ELISA was evaluated for the determination of the acute phase of infection. The specificity and the sensitivity of the immunoassay was shown to be 76.2% and 91.1%, respectively. Eight serum samples taken from patients with histopathological diagnosis, at different time points (3 to 15 months) after surgical treatment, showed a sharp and early decline in antibody reactivity. The titration of anti-A. costaricensis antibodies has proved to be a useful method for the diagnosis of acute abdominal angiostrongyliasis.

Report on the occurrence of Angiostrongylus costaricensis in southern Brazil, in a new intermediate host from the genus Sarasinula (Veronicellidae, Gastropoda).

Veronicellid slugs are the main intermediate hosts for Angiostrongylus costaricencis. In a rural locality in Nova Itaberaba (SC, southern Brazil) Sarasinula linguaeformis was identified as a crop pest. The parasitological examination revealed A. costaricencis infection in 43 out ot 50 slugs. The prevalence of 86% and the individual parasitic burdens are the highest sofar reported in Brazil and S. linguaeformis is the first species from the genus Sarasinula to be identified as intermediate host for A. costaricencis in southern Brazil.

Inactivation of infective larvae of Angiostrongylus costaricensis with short time incubations in 1.5% bleach solution, vinegar or saturated cooking salt solution.

Angiostrongylus costaricensis is a nematode parasite of wild rodents in the Americas. Man may become infected accidentally and, sometimes, suffers a very severe abdominal disease. Ingestion of raw vegetables has been proven to be a risk factor for the acquisition of A. costaricensis and, therefore, prophylaxis should include food disinfection. The larvicidal effect of wine vinegar, saturated cooking salt (SS) and a 1.5% bleach solution (BW) were compared with a 1-h-incubation period, at room temperature. Larval viability was tested through inoculation in Swiss mice. Only one out of 560 larvae treated with BW (97.3% of the animals were uninfected) was recovered as an adult worm, while 90/336 and 29/512 larvae treated, respectively, with SS and WV were recovered as adult worms. This larvicidal effect of BW was seen also in incubation times as short as 15 min. In conclusion, the 1.5% bleach solution may be helpful for prophylaxis of human abdominal angiostrongyliasis through disinfection of raw vegetables and unpeeled fruits.

Report on the occurrence of Angiostrongylus costaricensis in southern Brazil, in a new intermediate host from the genus Sarasinula (Veronicellidae, Gastropoda)

Veronicellid slugs are the main intermediate hosts for Angiostrongylus costaricencis. In a rural locality in Nova Itaberaba (SC, southern Brazil) Sarasinula linguaeformis was identified as a crop pest. The parasitological examination revealed A. costaricencis infection in 43 out ot 50 slugs. The prevalence of 86% and the individual parasitic burdens are the highest sofar reported in Brazil and S. linguaeformis is the first species from the genus Sarasinula to be identified as intermediate host for A. costaricencis in southern Brazil.

Lesmas veronicelídeas são os principais hospedeiros intermediários de Angiostrongylus costaricencis. Em uma localidade rural de Nova Itaberaba (SC, no sul do Brasil) Sarasinula linguaeformis apresenta-se como peste agrícola. O exame parasitológico das lesmas demonstrou infecção pelo A. costaricencis em 43 de 50 animais. A prevalência de 86% e as cargas parasitárias são as mais altas registradas até o momento no Brasil e S. linguaeformis é a primeira espécie do gênero Sarasinula a ser identificado como hospedeiro intermediário do A. costaricencis no sul do país.

The effect of temperature on mobility of Angiostrongylus costaricensis third stage larvae.

Third stage larvae (L3) from Angiostrongylus costaricensis were incubated in water at room temperature and at 5 degrees C and their mobility was assessed daily for 17 days. Viability was associated with the mobility and position of the L3, and it was confirmed by inoculation per os in albino mice. The number of actively moving L3 sharply decreased within 3 to 4 days, but there were some infective L3 at end of observation. A mathematical model estimated 80 days as the time required to reduce the probability of infective larvae to zero. This data does not support the proposition of refrigerating vegetables and raw food as an isolated procedure for prophylaxis of human abdominal angiostrongylosis infection.

Experimental Angiostrongylus costaricensis infection in mice: immunoglobulin isotype responses and parasite-specific antigen recognition after primary low-dose infection.

Immunoglobulin isotype responses and parasite-specific antigen recognition were investigated in experimental Angiostrongylus costaricensis infection in two different mouse strains. Even in a low-dose infection with third-stage larvae (L3), BALB/c mice showed high mortality until 28 days postinfection (p.i.) in association with a low patency rate in surviving animals. On the other hand, low mortality and a high rate of patent infection was observed in C57BL/10 mice. Parasite-specific IgM, total IgG, and IgG subclasses against crude adult-worm antigen (AcAg) rose in both groups of mice from day 14 onward, with IgG and IgG1 being significantly elevated in BALB/c mice at 21 and 28 days p.i., respectively. For total IgE, significantly elevated concentrations were detected at 14 days p.i. in BALB/c mice as compared with C57BL/10 mice. A. costaricensis-specific antigen recognition by total IgG, IgG1, or IgG2a was similar in both mouse strains, intensifying from 3 to 4 weeks p.i., with recognition of immunodominant AcAg ranging between 80 and 210 kDa. This study provides evidence that in BALB/c and C57BL/10 mice, immunoglobulins, with the possible exception of IgE and IgG1, do not decisively contribute to the outcome of a primary A. costaricensis infection with respect to immunopathogenesis or parasite permissiveness.

Isolation of Angiostrongylus costaricensis first-stage larvae from rodent feces on a Percoll gradient.

Several density gradients were tested for the isolation of parasitic nematode, Angiostrongylus costaricensis, first-stage larvae from rodent feces. With a 45/72% Percoll gradient, 83-99% (89.56+/-6.57%) of the larvae were recovered in a clean preparation.

Study on the elimination of Angiostrongylus costaricensis first stage larvae in the experimental infection of Swiss mice.

Abdominal angiostrongylosis is a nematode infection wild rodents. Human infection may result in severe abdominal disease and has been reported from several countries in the Americas. The domestic mouse, Mus musculus, has not been found with natural infection and, like other urban rodents, should not be considered a natural host for Angiostrongylus costaricensis. Quantification of parasitic forms released for transmission may better express the coevolutionary status in parasite-host relationship. With this objective, five groups of experimentally infected Swiss mice were followed for up to 155 days post-infection (PI) days and the quantification of first stage larvae (L1) output revealed: an irregular elimination of L1 and a huge variation in the patency period (1 to 114 days) and in the number of L1 eliminated daily by individual animals (1 to 6340 L1/g). Overall mortality was 72% (range: 28% to 100%) at seven weeks PI. In conclusion, abdominal angiostrongylosis in M. musculus presents high mortality and a very variable and irregular elimination of L1 in feces.