S. Geertsa, P. Kagerukaa, R. De Dekena, J.R.A. Brandta, J.M. Kazadia, B. Diarrab, M.C. Eislerc, Y. Lemmouchid, E. Schachtd, P.H. Holmesc
a Institute of Tropical Medicine, Nationalestraat, 155, B-2000 Antwerp, Belgium
b Laboratoire Central Vétérinaire, Bamako, Mali
c University of Glasgow, Veterinary School, Bearsden Road, Glasgow G61 1QH, UK
d University of Ghent, Laboratory of Polymers, Krijgslaan 261, B-9000 Ghent, Belgium
Introduction
Materials and methods
Results
Discussion
Acknowledgements
References
Given the promising results obtained in rabbits using sustained release devices (SRD) containing trypanocides (Geerts et al., 1993; Kageruka et al., 1996), it was decided to carry out some trials in cattle under experimental conditions.
Biodegradable sustained release devices (SRD) were prepared by extrusion of a mixture of poly (D, L-lactide) and isometamidium chloride (ISMM), (Samorin® or Trypamidium®, Rhône-Mérieux) or ethidium (Sigma). The SRD consisted of a cylindrical rod of 3 mm diameter, 3 cm long loaded with 25 % (w/w) of ISMM or ethidium. They were coated by dipping them in a chloroform solution (10 %) of the above mentioned polymer.
Dexamethasone (1 %, w/w) was added to the coating of the ethidium-SRD to reduce the tissue reaction around the SRD.
Two successive experiments were carried out each using six adult cows (520 - 799 kg). In the first experiment three cows were injected intramuscularly with ISMM at 0.5 mg/kg body weight. The other three cows received the same dose of the drug as subcutaneous implants. In the second experiment three animals were injected intramuscularly with 1 mg/kg ethidium, the three other cows received a SRD containing the same dose of ethidium.
The prophylactic effect of each drug formulation was evaluated by challenging the animals with an average of 8 Glossina morsitans infected with Trypanosoma congolense IL 1180 at monthly intervals from one month post treatment onwards. Dissection of the flies showed that metacyclic infections were present in 50 to 90% of the flies. Blood samples were taken weekly and examined by the buffy coat technique (Murray et al., 1977). The experiment was continued until all the animals had become positive for a maximum of 24 months after treatment. The infectivity of the trypanosome infections at each challenge was confirmed by exposing at least one control cow (each time a different one) to a challenge with a similar number of infected flies as the treated animals. Each of these challenged control animals subsequently became positive.
ISMM analysis of the serum samples was carried out using the ELISA method as described by Eisler et al., 1993.
ISMM sustained release devices
The average protection period was 20 months (range: 18-22) for the implanted group versus 5.7 months (range: 4-7) for the i.m. treated group. The protection period was thus 3.2 times longer for the ISMM-SRD treated group than for the animals which were treated with an i.m. injection of ISMM. At the implantation site of the SRD an inflammatory reaction was noticed shortly after the treatment. During the first weeks a quite large nodule (± 3 cm diameter) developed in all cattle, which disappeared gradually afterwards.
The kinetics of the concentration of ISMM in the sera of the implanted animals was completely different from that in the i.m. treated group. The average ISMM concentration remained quite constant (at least until 150 d.p.tr.) in the SRD treated group with peak levels of 0.8 ng/ml at about one month and still 0.4 ng/ml 5 months post treatment, whereas there was a rapid drop of the ISMM concentration in the i.m. injected group from an average of 5 ng/ml on day one until 0.1 ng/ml (detection limit of ELISA) at 85 days post treatment.
Ethidium sustained release devices
The average prophylactic period using the ethidium-SRD was 8.3 months (range: 5-10) versus 3 months (range 2-4) for the i.m. treated group. This means an extension of the protection period by a factor of 2.8 in comparison with an i.m. injection of ethidium. The inflammatory reaction at the implantation site was less pronounced than the one resulting from the ISMM-SRD (dexamethasone was added to the coating solution of the ethidium-SRD). The swelling disappeared slowly over a period of a few months.
Trypanolytic antibodies were not detected in any of the animals as long as they were protected by the treatment. The antibodies did appear only in those animals, in which a parasitaemia developed.
The advantages of the use of SRD are quite clear. Besides the significant extension of the protection period, it avoids the development of multiple fibrous lesions (and possibly toxic residues) at the injection sites deeply in the muscles (Boyt, 1971). Since the implants are degradable, they-will be resorbed if administered sufficiently long before slaughter.
The possible development of drug resistance after use of SRDs containing anthelmintic or other drugs is an important factor to be taken into consideration (Anderson, 1985; Peregrine, 1994). Comparison of the sensitivity of the breakthrough isolates obtained from implanted cattle with those from i.m. injected animals (based on CD80 and ED80 values using the mouse test according to Sones et al., 1988) did not indicate the development of resistance in trypanosomes isolated from the former group of animals, although a slight loss of sensitivity was observed in comparison with the parent trypanosome population (at least for ethidium). A similar loss of sensitivity of the breakthrough isolates as compared to the original stock of T. congolense (Kolo, Zaïre) was observed in the rabbit experiments using ethidium-SRD, but this was not the case for the isolate from the rabbits, which received the ISMM-SRD (Kageruka et al., 1996). Further research is necessary in order to examine this aspect in more detail.
Field trials are currently being conducted in Mali and Zambia in order to evaluate the efficacy of SRD in cattle maintained under conditions of heavy or moderate tsetse challenge.
This research project was financially supported by the EU-STD programme (contract No. TS3-CT93-240).
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