Expression of circulating leucocytes before, during and after myiasis by Dermatobia hominis in experimentally infected rats

Expression of circulating white blood cells was investigated in rats (Rattus norvegicus) experimentally infected with larvae of Dermatobia hominis, the human bot fly. Leucocytes were counted prior to infection (control group) as well as at 6, 10, 15, 20 and 28 days post-infection (dpi) and at 7, 15, 30 and 60 days post-larval emergence (dple). Total leucocyte numbers did not differ markedly among the groups. Significant differences were registered when values from control and animals harboring each larval stage of D. hominis were compared; with crescent rank: L1-, L2-, control and L3-infected groups. Leucocyte numbers were significantly higher in the control, 15, 20 or 28 dpi groups than in the 6 dpi animals. Higher counts were observed in control, L2- or L3-infected rats than L1-infected animals. Neutrophils, eosinophils and both large and small lymphocytes were also counted and analyzed. Basophils and monocytes were insufficient in number to permit statistical studies. These results stimulate the continuity of the studies about the host-parasite relationship in the dermatobiosis.

Partial characterization of proteolytic activity in Giardia duodenalis purified proteins

This report describes a preliminary characterization of proteolytic activity of proteins isolated from lysate of Giardia trophozoites of an axenic Brazilian strain. Fractions obtained by high-performance liquid chromatography (FPLC) were tested in SDS-polyacrylamide gel for the protein profiles, and the proteases activity was analyzed using gelatin impregnated SDS-PAGE. The proteases characterization was based on inhibition assays employing synthetic inhibitors for cysteine (E-64, IAA), serine (PMSF, TPCK, TLCK, and elastatinal), metalo (EDTA) and aspartic (pepstatin) proteases. Among thirty eluted fractions, polypeptide bands were observed in eight of them, however, proteolytic activity was detected in four ones (F23, F24, F25 and F26). Protein profiles of these fractions showed a banding pattern composed by few bands distributed in the migration region of 45 to < 18 kDa. The zymograms revealed proteolytic activity in all the four fractions assayed, mainly distributed in the migration region of 62 to 35 kDa. Among the profiles, the main pronounced zones of proteolysis were distinguished at 62, 55, 53, 50, 46 and 40 kDa. In inhibition assays, the protease activities were significantly inhibited by cysteine (E-64) and serine proteases (TPCK, TLCK and elastatinal) inhibitors. Gels incubated with other cysteine and serine protease inhibitors, IAA and PMSF, respectively, showed a decrease in the intensity of hydrolysis zones. Indeed, in the assays with the inhibitors EDTA for metalloproteases and pepstatin for aspartic proteases, none inhibition was detected against the substrate. These observations are relevants, especially if we consider that to define the real role of the proteases in host-parasite interaction, the purification of these enzymes for detailed studies may be warranted.

In vitro ANTIGIARDIAL ACTIVITY OF THE CYSTEINE PROTEASE INHIBITOR E-64

The quest for new antiparasitic alternatives has led researchers to base their studies on insights into biology, host-parasite interactions and pathogenesis. In this context, proteases and their inhibitors are focused, respectively, as druggable targets and new therapy alternatives. Herein, we proposed to evaluate the in vitro effect of the cysteine protease inhibitor E-64 on Giardia trophozoites growth, adherence and viability. Trophozoites (105) were exposed to E-64 at different final concentrations, for 24, 48 and 72 h at 37 °C. In the growth and adherence assays, the number of trophozoites was estimated microscopically in a haemocytometer, whereas cell viability was evaluated by a dye-reduction assay using MTT. The E-64 inhibitor showed effect on growth, adherence and viability of trophozoites, however, its better performance was detected in the 100 µM-treated cultures. Although metronidazole was more effective, the E-64 was shown to be able to inhibit growth, adherence and viability rates by ≥ 50%. These results reveal that E-64 can interfere in some crucial processes to the parasite survival and they open perspectives for future investigations in order to confirm the real antigiardial potential of the protease inhibitors.

PARACOCCIDIOIDOMYCOSIS TREATMENT

SUMMARYConsidered to be an emerging endemic mycosis in Latin America, paracoccidioidomycosis is characterized by a chronic course and involvement of multiple organs in immunocompromised hosts. Infection sequelae are mainly related to pulmonary and adrenal insufficiency. The host-parasite interaction results in different expressions of the immune response depending on parasite pathogenicity, fungal load and genetic characteristics of the host. A few controlled and case series reports have shown that azoles and fast-acting sulfa derivatives are useful treatment alternatives in milder forms of the disease. For moderate/severe cases, more prolonged treatments or even parenteral routes are required especially when there is involvement of the digestive tract mucosa, resulting in poor drug absorption. Although comparative studies have reported that shorter treatment regimens with itraconazole are able to induce cure in chronically-infected patients, there are still treatment challenges such as the need for more controlled studies involving acute cases, the search for new drugs and combinations, and the search for compounds capable of modulating the immune response in severe cases as well as the paradoxical reactions.

Human immune responses during schistosomiasis mansoni

Studies of immune responses as they occur in patients with schistosomiasis appear to progress relative to corrent technological advances, and to advance despite the understandable inability to pursue in vivo manipulations in this host/parasite system. Emphasis is most often placed on making immunological comparisons between such patient groups as reinfected/non-reinfected, intestinals/hepatosplenic, high/low intensities of infection, infected/uninfected within endemic areas, and those born of infected/uninfected mothers. Based on these types of comparisons, reasonable conjectures can be made regarding the immunological occurrences during this chronic exposure condition. Some consideration is now being given to the immune mechanisms of some of the observations made, and while some of these must then be carried back to experimental models for further manipulation-based analysis, new technological developments continue to assist in the field/bench ability to ask questions that might assist our understanding to a point where this knowledge can be applied to shaping developmental approaches to vaccine development and the goal of alleviating morbidity.

Angiostrongylus costaricensis and the intermediate hosts: observations on elimination of L3 in the mucus and inoculation of L1 through the tegument of mollucs

Human accidental infection with Angiostrongylus costaricensis may result in abdominal disease of varied severity. Slugs from the Veronicellidae family are the main intermediate hosts for this parasitic nematode of rodents. Phyllocaulis variegatus, Phyllocaulis soleiformis and Phyllocaulis boraceiensis were experimentally infected to describe the kinetics of L3 elimination in the mucus secretions of those veronicelid species. A maximum of 2 L3/g/day was found in the mucus, while the number of L3 isolated from the fibromuscular tissues varied from 14 to 448. Productive infection was established by inoculations in the hyponotum or in the body cavity, through the tegument. Intra-cavity injection is a less complex procedure and permits a better control of inocula. A preliminary trial to titrate the infective dosis for P. variegatus indicated that inocula should range between 1000 and 5000 L1. The data also confirmed the importance of P. variegatus as an intermediate host of A. costaricensis.

Hepatic pathology in Capillaria hepatica infected mice

Septal fibrosis of the liver regularly develops in rats infected with Capillaria hepatica. To find out whether such fibrosis also occurs in mice, 20 animals were submitted to infection with either 100 or 300 embryonated eggs and histologically examined after several periods of time, from 30 to 110 days afterwards. Results showed that mice developed acute, severe, diffuse and focal hepatic lesions that were soon modulated to focal areas of fibrosis containing eggs and worm remnants, despite the fact that a few worms remained alive, at least up to 110 days after inoculation. Areas of perisinusoidal fibrosis appeared in the proximity and around focal parasitic lesions, but clear-cut septal fibrosis was not observed. Why septal fibrosis forms in rats, but not in mice during C. hepatica infection, only further studies can clarify. Mice seem to show better host/parasite relationship than rats in regard to C. hepatica infection.

Evaluation of the phagocytic activity of peripheral blood monocytes of patients with Jorge Lobo's disease

Studies on host-parasite interaction in Jorge Lobo's disease are scarce, with no report in the literature on the phagocytosis of Lacazia loboi by phagocytic mononuclear cells. Thus, the objective of the present study was to assess the phagocytic activity of blood monocytes in the presence of L. loboi in patients with the disease and in healthy subjects (controls) over 3 and 24 hours of incubation. Statistical analyses of the results showed no significant difference in percent phagocytosis of the fungus between patient and control monocytes. With respect to incubation time, however, there was a significant difference, in that percent phagocytosis was higher at 3 hours than at 24 hours (p <0.01). These results suggest that monocytes from patients with the mycosis are able to phagocyte the fungus, as also observed in control individuals.

Capillaria hepatica-induced septal fibrosis in rats: a contribution to the study of liver fibrogenesis

INTRODUCTION: Septal fibrosis of the liver regularly develops in rats infected with the nematode Capillaria hepatica. Curative treatment of the infection prevents the development of septal fibrosis when intervention occurs up to postinfection day (PID) 15, but not later. The present investigation aimed to demonstrate which parasitic factors are present when the process of septal fibrosis can no longer be prevented by curative treatment. METHODS: Wistar rats were infected with 600 embryonated eggs of C. hepatica administered by gavage and treated with ivermectin and mebendazole in separate groups at PIDs 10, 12, 15, 17 or 20. Rats from each group and their nontreated controls, were killed and examined 40 days after the end of treatment. RESULTS: Findings by PID 15 were compatible with the stage of complete maturation of infection, when worms and eggs were fully developed and a complex host-parasite multifocal necroinflammatory reaction showed greater intensity, but with no signs of septal fibrosis, which appeared from PID 17 onward. CONCLUSIONS: Since the worms spontaneously died by PID 15, not only septal fibrosis production, but also its maintenance and further development appeared dependent on the presence of eggs, which were the only parasitic factor remaining thereafter.

Triatominae (Hemiptera, Reduviidae) in the Pantanal region: association with Trypanosoma cruzi, different habitats and vertebrate hosts

ABSTRACTINTRODUCTION: The transmission cycle of Trypanosoma cruzi in the Brazilian Pantanal region has been studied during the last decade. Although considerable knowledge is available regarding the mammalian hosts infected by T. cruzi in this wetland, no studies have investigated its vectors in this region. This study aimed to investigate the presence of sylvatic triatomine species in different habitats of the Brazilian Pantanal region and to correlate their presence with the occurrences of vertebrate hosts and T. cruzi infection.METHODS: The fieldwork involved passive search by using light traps and Noireau traps and active search by visual inspection. The light traps were placed at five selected points along forested areas for seven nights during each of the nine excursions. At each point where a light trap was set, eight Noireau traps were placed in palm trees and bromeliads.RESULTS: In all, 88 triatomine bugs were collected: two and one individuals from light traps and Noireau traps, respectively; three from peridomestic areas; 23 in coati nests; and 59 in thornbird nests. In this study, active search in microhabitats showed higher efficiency than passive search, since 95% of the triatomine bugs were caught in nests. Further, triatomine bugs were only found to be infected by T. cruzi in coati nests.CONCLUSIONS: Coati nests might act as a point of convergence and dispersion for triatomine bugs and mammal hosts infected by T. cruzi, thereby playing an important role in the sylvatic cycle of T. cruziin the Pantanal region.

Observations on two biological races of Schistosoma mansoni

A comparative study of the BH strain of Schistosoma mansoni from Belo Horizonte, Minas Gerais state, infective to Biomphalaria glabrata from the same locality, and the SJ strain from São José dos Campos, São Paulo state, infective to B. tenagophila from the latter locality, showed the following differences: 1. Length of adult worms and size of eggs significantly larger in the BH strain. 2. Higher infection rates in the B. glabrata-BH strain association than in the B. tenagophila-SJ strain association, following exposure of each snail to 1 or 10 miracidia. 3.Longer prepatent period (from penetration of miracidium to first shedding of cercariae) in the B. tenagophila-SJ strain association. 4. Infection of both Biomphalaria species when exposed to hybrid miracidia from crosses between the two strains, at lower levels than those resulting from exposure of each snail species to miracidia of the pure sympatric strain. (Both Biomphalaria populations are practically refractory to infection with the allopatric strain). These results are interpreted as pointing to a better host-parasite adjustment in the B. glabrata-BH strain association than in the B. tenagophila-SJ association. The interfertility between the two strains, which produced viable hybrids infective to both Biomphalaria species, supports the conclusion that the observed differences are merely intraspecific, and that the two strains may be considered distinct biological races of Schistosoma mansoni.

Interactions between Leishmania mexicana mexicana promastigotes and amastigotes and murine peritoneal macrophages in vitro

Unstimulated adherent mouse peritoneal cells were cultured in vitro and infected with equal numbers of a single strain of Leishmania m. mexicana amastigotes (AM), virulent promastigotes (VP), avirulent promastigotes (AVP) and fixed promastigotes (FP). Duplicate May-Grünwald-Giemsa stained coverslips were examined at time intervals up to 13 days. By 3 hr post infection, the number of macrophages containing parasites varied between 60.5% (VP) and 84% (AM) for macrophages exposed to living parasites, compared to 6.5% for macrophages exposed for FP. However, variable numbers of parasites showed degenerative changes by 3 hr, and the number of macrophages containing morphologically intact parasites varied significantly between cells infected with AM (84%) and those infected with VP (42%) or AVP(40%). The mean number on intacte parasites/macrophage also differed significantly between AM-infected cells and living or fixed promastigotes-infected cells. Quantitation of intact and degenerated parasites indicated parasite multiplication, as well as destruction, in VP-infected cells and parasite survival and multiplication in AM-infecte monolayers; in contrast no evidence of parasite multiplication was seen in AVP-infected cells. Changes in the mono layer itself (cell loss and macrophage vacuolization) were also evaluated. These results suggest that crucial events determining the outcome of infection occur in the host-parasite relationship during the fist 24 hours of infection. These events are apparently influenced not only by parasite or host strain but by environmentally induced variation within a given strain.

Prevalence and control of Babesiosis in the Americas

This review presents up-to-date information on the distribution and control measures of babesiosis in Latin America. Bovine babesiosis caused by Babesia bovis and B. bigemia will be emphasized. The disease is endemic is most countries and poses a serious economic burdenon livestock production in the region (U.S.$1365 million/year, FAO, 1989). Of the estimated 250 million cattle in Central and South America, approximately 175 million (70%) are in tick-infested regions. Humid, tropical and subtropical areas favor development of the main vector, the one-host tick Boophilus microplus. In many regions bovine babesiosis is enzootically stable as consequence of a balanced host-parasite relationship. However, Latin America offers a wide range of epidemiologica conditions that are influenced by variations from tropical to cool climates and by susceptible purebred cattle that are regularly imported to upgrade local stocks. The control measures employed in most countries for babesiosis esentially rely on chemotherapy, use of acaricides for B. microplus, and to a lesser degree, on immunization methods. In general, these measures are expensive, time consuming, and in many cases, provide limited success. Finally, the zoonotic potential ob babesiosis will be addressd, with special emphasis on the situation in the United States. Even though bovine babesiosis has long been eradicated from the U.S.A., human babesiosis in endemic in the northeastern region of the country.

Transmission immunity in malaria: reflections on the underlying immune mechanisms during natural infections and following artificial immunization

Malaria transmission-blocking immunity has been studied in natural malaria infections in man, during infections in animals and following artificial immunization of animals with sexual stage malaria parasites. Effective immunity, which prevents infectivity of a malarial infection to mosquitoes, has been observed under all of these circumstances. Two general types of effector mechanism have been identified. One is an antibody mediated mechanism which acts against the extracellular sexual stages of the parasite within the midgut of a blood feeding mosquito. The other is a cytokine mediated mechanism which inactivates the gametocytes of the parasites while in the circulation of the vertebrate host. Both effects have been observed during natural infections and following artificial immunization. The basis of induction of transmission-blocking immunity, including the nature of the memory for such immunity, however, may be very different in different host/parasite systems and during natural infection of following artificial immunization. Following artificial immunization a strong immune memory for transmission blocking immunity has been observed in animal systems. By contrast, following natural infections in man immune memory for transmission blocking immunity has been found to be weak and short lived if it occurs at all. It is suggested that the immunogens which induce natural transmission blocking immunity may be CD4+ independent.

Imunodiagnóstico

In the last years, the knowledge about immunodiagnosis in schistosomiasis has increased considerably. This was due to a better immunologic understanding of the host-parasite interaction and the evolution of the technical procedures by the introduction of new concepts and techniques. The search of more sensitive, specific, practical and less expensive diagnostic tests has led to the development of a great number of immunological tests that could complement the limitations of the parasitological diagnosis. The antigens of differnt life cycle stages of Schistosoma mansoni may be used as target for immunodiagnostic tests and the anti-schistosome antibodies in sera of infected patients can be detected. The research of circulating schistosome antigens, the production of specific antisera and the application of these antibodies in immunodiagnostic tests have also been discussed.