Cellular responses and cytokine production in post-treatment hookworm patients from an endemic area in Brazil.

Human hookworm infections are distributed widely in tropical areas and have a significant impact on host morbidity and human health. In the present study, we investigated the cellular responsiveness and cytokine production in peripheral blood mononuclear cells (PBMC) from Necator americanus-infected schoolchildren who had recently received chemotherapy, and compared them with non-infected endemic controls. Hookworm patients and treated, egg-negative individuals showed a lower cellular reactivity against phytohaemagglutinin (PHA) and hookworm antigen when compared with egg-negative endemic controls. The baseline production of proinflammatory tumour necrosis factor-alpha (TNF-alpha) in PBMC from infected patients and treated, egg-negative individuals was elevated. On the other hand, PHA- or hookworm antigen-induced interleukin (IL)-12 and interferon (IFN)-gamma secretion was higher in endemic controls than in hookworm patients, who either continued egg-positive or were egg-negative after treatment. Also, PBMC from endemic controls secreted more IL-5 and IL-13 than the other patient groups. Opposite to that, the spontaneous as well as the antigen-driven IL-10 secretion was lower in endemic controls when compared with the other groups. In summary, patently hookworm-infected as well as egg-negative treated patients disclosed an elevated spontaneous cellular secretion of proinflammatory TNF-alpha, a prominent secretion of regulatory Th2-type IL-10 and an impaired production of IL-12, IFN-gamma, IL-5 and IL-13.

In vitro study on thiabendazole action on viability of Ascaris lumbricoides (Lineu, 1758) eggs.

The in vitro activity of thiabendazole on Ascaris lumbricoides eggs, which were recovered from uteri of worm excreted after chemotherapeutic treatment, was studied. Four concentrations of the drug were used: 1 - 2.5 - 5 - and 10 ppm during 24, 48 and 72 hours of exposure. Subsequently, the eggs were centrifuged, washed three times and H(2)SO(4)0.1N was added. The eggs were maintained in an incubator for 20 days at 28 degrees C. Finally, the percentage of embryonated eggs was determined under a lightmicroscope at a 100X magnification. After 48 and 72 hours of thiabendazole exposure, at a concentration of 10ppm, the drug showed complete inhibition of egg embryonation.

In vitro study on thiabendazole action on viability of Ascaris lumbricoides (Lineu, 1758) eggs

The in vitro activity of thiabendazole on Ascaris lumbricoides eggs, which were recovered from uteri of worm excreted after chemotherapeutic treatment, was studied. Four concentrations of the drug were used: 1 - 2.5 - 5 - and 10 ppm during 24, 48 and 72 hours of exposure. Subsequently, the eggs were centrifuged, washed three times and H(2)SO(4)0.1N was added. The eggs were maintained in an incubator for 20 days at 28 degrees C. Finally, the percentage of embryonated eggs was determined under a lightmicroscope at a 100X magnification. After 48 and 72 hours of thiabendazole exposure, at a concentration of 10ppm, the drug showed complete inhibition of egg embryonation.

Foi estudada, in vitro, a ação do tiabendazol, contra ovos de Ascaris lumbricoides , retirados de úteros de vermes eliminados após tratamento. Foram utilizadas quatro concentrações da droga: 1 ¾ 2,5 - 5 e 10 ppm em três diferentes períodos de tempo: 24, 48 e 72 horas. Decorridos estes tempos a suspensão com os ovos foi centrifugada, lavada por três vezes com água destilada, adicionada de solução de H2SO4, 0,1N e mantida em estufa a 28°C por 20 dias. Em seguida, os ovos foram levados ao microscópio óptico, no aumento de 100X, para observação do percentual de embrionamento. Na concentração de 10ppm, nos tempos de 48 e 72 horas de exposição à droga, verificou-se completa inibição do embrionamento dos ovos.

Susceptibility and resistance to Schistosoma mansoni reinfection: parallel cellular and isotypic immunologic assessment.

Cellular and humoral immune responses to Schistosoma mansoni antigen preparations were evaluated in individuals presumed to be susceptible or resistant to reinfection after chemotherapeutic cure. A consistent proliferative increase in the response to soluble egg antigen (SEA) was observed post-treatment in both the susceptible and resistant groups. However, this change was not related to resistance. Isotype studies showed that IgM antibody levels to soluble worm antigen preparation (SWAP) and cercariae antigens were significantly higher in the resistant group than in the susceptible group. Post-treatment, an increase in IgE anti-SWAP and anti-schistosomular tegument (STEG) responses and a decrease in IgG4 anti-SEA and anti-STEG responses were observed in the resistant group. These finding are similar to those we have reported previously for a putative resistant group termed endemic normals, and are compatible with immunologic studies in different endemic areas. Together, these findings indicate that even on the population level, high IgE specificities coupled with low IgG4 specificities correlate well with documented resistance to reinfection.

Re-evaluation of schistosomiasis mansoni in Minas Gerais, Brazil. III. "Noroeste de Minas" mesoregion.

This study was conducted to assess the presence of schistosomiasis mansoni in the "Noroeste de Minas" mesoregion, an area considered non-endemic. A malacologic survey and parasitologic stool examinations were undertaken in 13 municipalities of the mesoregion. A sample of 3,283 primary school students was submitted to fecal examination by the Kato-Katz method. A total of 3,627 planorbids was collected and examined. The molluscs were identified as Biomphalaria straminea in seven municipalities (Unaí, Bonfinópolis de Minas, Paracatu, Jaão Pinheiro, Vazante, Lagamar and Lagoa Grande) and as Biomphalaria peregrina in one (Presidente Olegário). All planorbids were negative for Schistosoma mansoni. Four students were diagnosed with schistosomiasis in the municipalities of Buritis, Formoso, Paracatu and Unaí, but none of these cases was considered autochthonous. The data obtained indicate that the "Noroeste de Minas" mesoregion continues to be non-endemic for schistosomiasis mansoni, although the presence of intermediate hosts associated with parasitized individuals emphasizes the need for epidemiological surveillance of schistosomiasis in this mesoregion.

Comparison of antibody isotype responses to Schistosoma mansoni antigens by infected and putative resistant individuals living in an endemic area.

The isotypic patterns of antibodies against Schistosoma mansoni antigenic preparations from eggs (SEA), adult worms (SWAP) and cercariae (CERC) have been studied in sera from two groups of individuals living in an area endemic for S. mansoni. One of the groups was comprised of individuals diagnosed as having S. mansoni infections based on their patency, i.e. those passing eggs in their faeces (patent infections, PI). The other group has been consider 'putatively resistant' due to their residence in an endemic area, their documented exposure to positive transmission sites, and their repeated negativity upon stool examinations (endemic normals, EN). There are strong specific responses of IgG1, IgG4 and IgM, particularly to SEA and CERC, by both groups. The reactivities of all isotypes were lower to SWAP. The responses of IgG4, IgM and IgE anti-CERC in EN and PI are higher than those found in normal individuals from outside endemic areas. In general, EN individuals express a relative higher level of anti-STEG IgE as compared to IgG4. On the other hand the pool of sera from PI showed the opposite pattern of higher IgG4 as compared to IgE. Several correlations are seen between isotypic responses to SEA, SWAP and CERC based on comparisons to the anti-SWAP IgE responses of the individuals in the two groups. These comparisons indicate the presence of distinct immunologic differences between individuals in the PI and the EN groups.

Schistosomiasis mansoni in the region of the Triângulo Mineiro, State of Minas Gerais, Brazil.

In order to reevaluate the possible presence of schistosomiasis mansoni in the Triângulo Mineiro, one of the areas of the State of Minas Gerais where this parasite is not commonly found, malacological survey and fecal examinations were undertaken in the region between October 1990 and June 1992. A sample of 7,032 1st grade school children from 29 counties had their feces examined using the Kato-Katz method. Amongst the children examined, two from Planura and one from each countie of Capinópolis, Conceição das Alagoas, Uberaba, Uberlândia, Prata and Gurinhatã were positive for Schistosoma mansoni. None of the children were identified as being autoctonous cases. In the malacological survey, 5,406 planorbid snails were examined. The specimens were identified morphologically and examined for S. mansoni by squashing between glass plates. The species were identified as Biomphalaria tenagophila in three counties, as B. straminea in ten and B. intermedia in 16. No snails were found in eight other counties studies. The snails were found to be negative for S. mansoni. The presence of intermediate hosts for S. mansoni, associated with parasitized individuals emphasizes the necessity of epidemiological surveillance for schistosomiasis in the region of Triângulo in the State of Minas Gerais.

Interferon-gamma production by peripheral blood mononuclear cells from residents of an area endemic for Schistosoma mansoni.

During human schistosomiasis host responses to antigens of various parasite life-cycle stages may contribute to whether the severe, hepatosplenic state develops or the patient remains relatively asymptomatic throughout infection, and may play a role in resistance. This study evaluated production of interferon gamma (IFN-gamma) in vitro by schistosome antigen-stimulated peripheral blood mononuclear cells (PBMCs) from asymptomatic patients, and by PBMCs from apparently uninfected, untreated persons living in areas endemic for Schistosoma mansoni ('endemic normals'). IFN-gamma production parallels PBMC proliferation in that schistosomal egg antigens stimulate patent patients' cells poorly, but strongly stimulate PBMCs from 'endemic normals'. This is proportionally true for antigens from adult worms and cercariae. Although asymptomatic patent patients' cells produced little or no IFN-gamma in response to the 3 schistosomal antigenic extracts, their PBMCs, and PBMCs from 'endemic normals', produced expected amounts of IFN-gamma when exposed to phytohaemagglutinin. This implies that persons with patent infections have schistosome antigen-specific defects in their ability to respond to IFN-gamma production that are not exhibited by putatively resistant 'endemic normals'.

[The susceptibility of Planorbidae from the metropolitan area of Belo Horizonte, MG (Brazil) to Angiostrongylus costaricensis (Nematoda, Angiostrongylidae)]. / Suscetibilidade de planorbídeos da região metropolitana de Belo Horizonte, MG (Brasil) ao Angiostrongylus costaricensis (Nematoda, Angiostrongylidae).

Biomphalaria glabrata (control), B. tenagophila and B. straminea from our laboratory colonies iniciated with molluscs collected in the municipality of Belo Horizonte, MG (Brasil), were experimentally infected with first-stage larvae of Angiostrongylus costaricensis. The number of molluscs of each species exposed was 139, 77 and 149. About 25 days later, surviving molluscs were individually examined by artificial digestion. Of 87 B. glabrata examined, 62 (71.3%) were positive and between one and 61 third-stage larvae were found; of 42 B. tenagophila, 21 (50.0%) contained between one five third-stage larvae; and of 89 B. straminea, 69 (77.5%) presented between one and 72 third-stage larvae. The three molluscan species are susceptible to A. costaricensis infection, but B. glabrata and B. straminea are most suitable for maintaining the nematode cycle in laboratory.

[Sarasinula marginata (Semper, 1885) (Mollusca, Soleolifera) from Belo Horizonte (MG, Brasil) as a potential intermediate host of Angiostrongylus costaricensis Morera, Cespedes, 1971]. / Sarasinula marginata (Semper, 1885) (Mollusca, Soleolifera) de Belo Horizonte (MG, Brasil) como hospedeira intermediária potencial do Angiostrongylus costaricencis Morera & Céspedes, 1971.

Specimens of Sarasinula marginata were collected in kitchen and house gardens of Belo Horizonte, Minas Gerais. The susceptibility of these molluscs for Angiostrongylus costaricensis was tested by infecting 15 laboratory--reared slugs (F1). The positivity demonstrated was of 80.0%.

Development of Ascaris lumbricoides eggs from females eliminated after chemotherapy in man

The development of Ascaris lumbricoides eggs obtained from females eliminated after treatment of infected individuals with a single oral dose of the antihelminthic drugs thiabendazole (50 mg/kg - 33 patients) or levamisole (250 mg - independent of body weigth - 20 patients) was studied. Every female eliminated up to 72 h after treatment were dissected, the uterus isolated and sectioned into small fragments. The eggs were transferred to plastics tubes and incubated at 28 degrees centigrades in 0.1 N H2 SO4 for 100 days. Every 20 days, starting from the 20 th up to the 100 th day, the extent of egg embryonation ratio was determined. The culture of A. lumbricoides eggs obtained from females from patients treated with thiabendazole did not contain embryonated eggs until the final period of observation. In contrast, the eggs obtained from females eliminated by patients treated with levamisole (control) presented an embryonation rate of 0.0 - 98.0% in the same period

Development of Ascaris lumbricoides eggs from females eliminated after chemotherapy in man.

The development of Ascaris lumbricoides eggs obtained from females eliminated after treatment of infected individuals with a single oral dose of the antihelminthic drugs thiabendazole (50 mg/kg--33 patients) or levamisole (250 mg--independent of body weight--20 patients) was studied. Every female eliminated up to 72 h after treatment were dissected, the uterus isolated and sectioned into small fragments. The eggs were transferred to plastics tubes and incubated at 28 degrees C in 0.1 N H2SO4 for 100 days. Every 20 days, starting from the 20 th up to the 100 th day, the extent of egg embryonation ratio was determined. The culture of A. lumbricoides eggs obtained from females from patients treated with thiabendazole did not contain embryonated eggs until the final period of observation. In contrast, the eggs obtained from females eliminated by patients treated with levamisole (control) presented an embryonation rate of 0.0-98.0% in the same period.

Immunological profiles of patients from endemic areas infected with Schistosoma mansoni.

Crude extracts of eggs (SEA) adult worms (SWAP) or cercariae (Cerc) have been used to stimulate Peripheral Blood Mononuclear cells (PBMC) and have provided rather distinct profiles of responses in different types of patients. In general it is clear that patients with early infections respond strongly to SEA while response to SWAP are develop more slowly. As infection progresses into the more chronic phases, a general pattern is seen which leads to lower anti-SEA proliferative responses in the face of higher responses to SWAP and variable anti-cerc responsiveness. Cured not re-exposed patients express very high levels of anti-SEA proliferation. It has recently been seen that those individuals who live in endemic areas and have continued water contact, but are repeatedly stool-negative (who are presumed to have self-cured or be putatively resistant; endemic normals) are strongly responsive to antigenic extracts, particularly to SEA. Furthermore, our results show that endemic normal individuals have significantly higher IFN gamma production upon PBMC stimulation with schistosome antigens than infected individuals. With the emergence of more studies it is becoming apparent that both the intensity and the prevalence of a given area may influence or shape the general responsiveness of the population under study.

Viability of Ascaris lumbricoides eggs eliminated after anti-helminthic therapy.

The viability of Ascaris lumbricoides eggs passed in the feces was evaluated after treatment of patients with one of the anti-helminthic drugs (thiabendazole, levamisole, cambendazole, pyrantel pamoate, mebendazole or praziquantel). For each drug, a group of 5 children was selected and their feces collected 24 h before treatment and 24, 48 and 72 h after drug administration, except for mebendazole, with the feces being collected throughout the period of treatment. After sedimentation, the total amount of eggs from each collection was transferred to tissue culture flasks containing 10 ml H2SO4 0,1N, with the addition of 3 drops of a miconazole solution, and incubated at 28 degrees C, individually, for 80 days. The flasks were maintained open and the culture were oxygenated daily by manual agitation. On the 80th day of culture, 20-days-old albino mice were inoculated with 3,200 embryonated eggs, per os. Larvae were recovered from their lungs and hearts, on the 8th day after infection, according to Baerman's method (Morais, 1948). Thiabendazole showed 100.0% ovicidal capacity as early as 48 h after treatment. Inhibition of embryonal development was observed when thiabendazole was used. This drug also had an effect on the eggs infectivity when inoculated into normal mice. No significant effect on embryonal development was observed for the other drugs tested.

[The study of Ascaris lumbricoides in the laboratory]. / Contribuição para o estudo do Ascaris lumbricoides em laboratório.

With the purpose of standardising techniques for the laboratory study of Ascaris lumbricoides, faeces were collected from children parasitised by A. lumbricoides, during a 24 hour period. The fecal samples were sieved and resuspended several times in water. The sediment containing the eggs was cultivated in H2SO4 0.1N in tissue culture flasks, at 28 degrees C. The culture of embryonated eggs was determined every ten days starting from the 20th day of culture achieving around 98.0% embryogeny on the 80th day of culture. The best day to recover larvae from mice was determined by infecting 9 groups of 5 mice per os with 200 embryonated eggs/mouse. Each mouse was sacrificed by cervical rupture starting on the 4 day up to the 12 day of infection. On the 8 day after the infection 0.45% of the larvae were recovered from the lungs. The inoculum determination was performed by using 5 groups of 10 infected mice with 200, 400, 800, 1,600 and 3,200 embryo eggs/mouse. The best age for recovery of infection was achieved by using 4 groups animals, with 5 mice/group, with age varying from 20 to 50 days and an inoculum of 3,200 eggs. The best recovery (1.9% was obtained from the group of 20 days of age.

[Schistosomiasis mansoni in the southwest of the State of Minas Gerais (Brazil)]. / Esquistossomose mansoni no sudoeste do Estado de Minas Gerais (Brasil).

A new focus of schistosomiasis mansoni at Passos, a town in the Southwest of the State of Minas Gerais (Brazil), region until now considered free of the disease is reported. Malacological surveys showed Biophalaria glabrata naturally infected with Schistosoma mansoni in a country club near Passos. All B. straminea captured at the pisciculture station of the Furnas hydroelectric dam were negative. Six out of seven individuals living in the country club were found to be infected with S. mansoni, including four children who had never been out of Passos. The epidemiological importance of these findings is discussed.