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halfbreed
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the brazil study
MATERIALS AND METHODS
All experimental work was carried out at the Bambui Research Centre of the Oswaldo Cruz Foundation, Brazil, where a research concerning the effects of reinfection on the pathology of T. cruzi infection in dogs was conducted. The dogs were young mongrels, maintained in the Centre kennels since birth. They were infected and re-infected with two different Brazilian strains of T. cruzi, being all the details pertaining to the experimental design and parasite behaviour and strain characteristics presented and discussed formerly (Machado et al. 2001). During a sudden infestation of R. sanguineus in the kennels, affecting all the animals, it became necessary to treat them in order to avoid anaemia and skin infections, as well as to prevent possible interference of tick infestation on the course of the experiments. For this, all the dogs were treated with subcutaneous injections of ivermectin (Detomax ®) at 20 mg a.i./kg. Ivermectin is the 22,23-dihydro derivative of avermectin B1, a macrocyclic lactone produced by the actinomycete Streptomyces avermitilis, which is active against a wide variety of nematode and arthropod parasites (Campbell et al. 1983, Azambuja et al. 1985, Campbell 1985, Rey 1991, Cimermann & Cimermann 1999, Barbosa & Campos 2001, PAHO 2001). The present data are a product of a casual situation, in which some dogs experimentally infected with T. cruzi became infested by R. sanguineus, so providing the opportunity to observe the possibility of a natural infection of ticks by that flagellate, as well as the eventual action of ivermectin against triatomines and trypanosomes. For this reason the experimental design of the work involves natural limitations, certainly being possible and desirable further investigations.
Fourteen young dogs with a high degree of R. sanguineus infestation were included in the present observations. Group I involved six dogs in the chronic phase of infection with both T. cruzi strains; Group II involved six dogs in the acute phase of infection; Group III involved two dogs uninfected with T. cruzi.
R. sanguineus was determined by the authors and confirmed by staff of the Zoology Department of the Federal University of Minas Gerais. To measure the infection rate of R. sanguineus by T. cruzi before the treatment with ivermectin, five fully engorged ticks were collected from each dog. Their digestive tract content was examined both by direct microscopy at 400 times (100 examined fields) and after cultivation in LIT/NNN medium (30 and 60 days of culture at 28 °C, considering two tubes for each examined tick). Pre- and post-treatment parasitaemia in the dogs was assessed by daily examination of fresh blood films (Brener 1961, Machado et al. 2001) for a period of 30 days post ivermectin treatment. Xenodiagnosis of each dog was also carried out at 13 days post-treatment using 20 laboratory-reared 4th instar nymphs of Triatoma infestans. The faecal material of these bugs was microscopically examined after 30, 60, and 90 days, after which negative bugs were dissected for examination of their entire digestive tract content.
To assess the effect of the ivermectin treatment on feeding triatomine bugs, a xenodiagnosis box containing five 4th instar nymphs of T. infestans was applied for 30 min to each of the dogs at intervals of 24, 72, 144, and 312 h after ivermectin treatment. These bugs were then maintained individually and offered a weekly feed on white mice, in order to assess their subsequent survivorship over the following 35 days. To compare the effects on different triatomine species, ten 4th instar nymphs of T. infestans and ten 4th instar nymphs of Rhodnius neglectus were allowed to feed from the same dog 24h after ivermectin treatment. These bugs were then maintained for 35 days, with a bloodfeed on chicken offered after 15days, in order to assess subsequent survivorship.
RESULTS
Action of ivermectin against tick infestation - The ivermectin treatment appeared to be highly effective against R. sanguineus, leading to complete elimination of all signs of tick infestation of the dogs within one week of treatment. Two months later, however, some tick reinfestation was noted presumably due to ticks hiding within the structure of the kennels. Accordingly, ivermectin treatment was repeated, but accompanied also by residual spraying of the kennels themselves with deltamethrin SC (25 mg a.i./m2 ). No ticks have been found since then, either on the dogs or within the kennel structure. There were no discernable side-effects of ivermectin treatment on the dogs, confirming previously reported studies (Barbosa & Campos 2001, Campbell 1985).
Parasitism of ticks by T. cruzi - No evidence of T. cruzi infection was found amongst ticks collected from chronically infected dogs (Group I). However, one of 37 ticks collected from dogs in the acute stage of T. cruzi infection (Group II) showed by microscopy a sparse infection of flagellates. LIT/NNN cultures from these ticks showed no flagellates at 30 days, but one culture from 27 of 37 ticks (73%) showed positive at 60 days (Table I).
Influence of ivermectin on T. cruzi infection in dogs - Considering the opportunistic character of this observation, in spite of the natural limitations in terms of technical approach, no apparent influence of ivermectin treatment was discernible on T. cruzi parasitaemia in the infected dogs. All treated dogs in the acute phase of infection (Group II) presented detectable parasitaemia by direct blood film examination during the first three weeks of infection, with similar levels of parasitaemia to those observed in untreated dogs during other periods of the long-term study (see Machado et al. 2001). In our experience, dogs in the chronic phase of T. cruzi infection never present detectable parasitaemia by direct examination, and this was unchanged by the ivermectin treatment. Similarly, xenodiagnosis of chronically infected dogs did not reveal T. cruzi infection, either in ivermectin treated animals or in previously studied untreated animals (Table II).
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