trypanosome

Trypanocides are essential in trypanosomosis management, but evidence regarding treatment efficacy in equids is scarce. The objective of this study was to establish the relative efficacy of three trypanocides (Diminasan® 3.5 mg/kg IM, Cymelarsan® 0.25 mg/kg IV and Samorin® 0.5 mg/kg IV) with respect to improvement of clinical parameters and parasitic burden and to evaluate adverse drug reactions. A prospective randomised clinical efficacy study was performed in ten villages in The Gambia. Owners were invited to present horses and donkeys for free examination (history, clinical examination and jugular blood sample for packed cell volume (PCV) and total serum protein). Horses and donkeys were enrolled if they fulfilled at least 2/5 inclusion criteria for trypanosomosis (anaemia (PCV<24%), poor body condition (≤1.5), limb or ventral oedema, abortion or pyrexia). Randomised trypanocide treatment was administered and the animals were observed for adverse reactions. Follow up evaluation was performed at one and two weeks to assess treatment effect. Blood samples for each animal collected at weeks 1, 2 and 3 underwent PCR analysis with validated specific primers1 for T. vivax west (TVW), T. congolense savannah (TCS) and T. brucei (TBR). 254/710 animals examined fulfilled study inclusion criteria with follow up data obtained for 243. Age, gender, species, median PCV (22%; range 8-26) and body condition score (median 1.5/5; range 0.5-2.5) were comparable between treatment groups (p>0.1). No immediate adverse reactions occurred following Cymelarsan® or Diminasan®. Immediate reactions occurred in 12/45 (27%) of donkeys treated with Samorin® ranging from neck scratching, lip smacking to tachycardia, cold extremities, sweating and hypothermia. Demeanour classifications improved following treatment with Samorin® or Diminasan® (p=0.002). PCV increased at 1 and 2 weeks post treatment for all treatment groups (p<0.001). On preliminary analysis of PCR results (n=65), animals representing four villages were positive before treatment (week 1) for TVW (55/65; 85%), TCS (44/65; 67%) and TBR (17/65; 26%) with mixed infection common (45/65; 69%). Trypanosome species profile varied between villages (p<0.05). Post treatment positives occurred in all groups for all Trypanosoma sp but with a marked decrease in prevalence (Fig 1). Positives were most common in the Cymelarsan® group, particularly for TCS (7/13; 54%). Two weeks post treatment Diminasan® (15/19; 79%) and Samorin® (19/23; 83%) had reduced parasitaemia below the threshold of detection in most cases. The data support the continuation of treatment with Diminasan® and Samorin® (with careful titration of dosing in donkeys). Further investigation to quantify parasitaemia in post treatment positives will aid differentiation between treatment failure, reduced parasitaemia, new infections and residual non-viable parasite DNA.

Acknowledgements

This work was funded by The Donkey Sanctuary.

In many countries diseases affecting working equid performance and productivity are detrimental both to equid welfare and to local economic development. Central nervous system (CNS) trypanosomosis, caused by Trypanosoma brucei spp 1, is a severe manifestation of trypanosomosis, which is usually fatal. The causative agent of this condition in the Gambia was investigated further in this study, to determine genotypic variation, mode of transmission and future management interventions. The presence of trypanosomes in local tsetse flies (Glossina spp.) was also investigated due to suspected vector involvement in disease transmission. Working equids exhibiting signs of CNS trypanosomosis were clinically evaluated. Blood was stored in EDTA and on FTA® cards prior to DNA extraction. In advanced neurological disease where prognosis was hopeless euthanasia was performed. CSF and CNS tissue samples were collected post-mortem. CSF was stored on FTA® cards and tissue samples were collected in formalin and RNAlater®. To confirm CNS T.brucei spp infection, immunohistochemistry and T.brucei-specific PCR 2 was performed on tissue samples. DNA was also extracted from blood collected from patients with evidence of generalised T.brucei infection with normal neurological function, and from the midguts of locally caught tsetse. Parasite population structure was investigated using a panel of microsatellite markers 3 together with a reference strain of T.brucei equiperdum (OVI) and a T.b.brucei positive control.Ten cases (5 horses, 5 donkeys) with naturally occurring CNS trypanosomosis were included. Horses presented with rapidly progressive spinal ataxia while donkeys showed slowly deteriorating cerebral dysfunction and cranial nerve abnormalities. CNS trypanosomosis was confirmed post-mortem using immunohistochemistry and PCR. Histopathological evaluation revealed diffuse lymphocytic-plasmacytic meningoencephalomyelitis. Microsatellite fragment analysis showed a heterogenous parasite population with a large range of alleles present, inconsistent with a clonal population. Parasite populations from donkey versus horse, and from blood versus CNS tissue were not found to be significantly different, suggesting that host factors are important in progression of neurological disease. Of 405 tsetse trapped locally and dissected, 11 contained microscopically visible midgut trypanosome infections. DNA extracted from the positive tsetse midguts was positive for equid DNA in 3/11 cases, confirming vector involvement. 5/11 flies were positive for T.brucei but with different microsatellite patterns to that found in infected CNS tissue. Further work is required to develop an optimal panel for use in both tsetse and equine-derived samples. Continued efforts are required to improve understanding of the transmission of this disease to enable the development of effective preventative measures.

Acknowledgements

This work is funded by The Donkey Sanctuary.