Animal Production

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Browsing Animal Production by Author "Ahmed Yasine, Yasine"

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    Trypanosoma equiperdum: Venereal transmission and Pathogenesis
    (2019-08) Ahmed Yasine, Yasine; Prof. Hagos Ashenafi, Prof. Dr. Bruno Goddeeris; Prof. Dr. Luc Duchateau
    Dourine, a venereal transmitted trypanosomosis caused by Trypanosoma equiperdum, has different clinical signs related to the reproductive and nervous system. It is one of the life-threatening venereal diseases in equidae. So far, there is no clear evidence on how and when stallions become infectious, nor which tissues are affected by the parasite in diseased animals. Post-infection, after a transient, temporary phase of parasitaemia, the parasite disperses to different tissues in an unknown distribution pattern. A review of the available literature was performed and knowledge of previous reports concerning dourine and T. equiperdum was gathered (Chapter 1). T. equiperdum is very closely related to T .evansi (the causative agent of surra) in morphology, biochemical and molecular characteristics. Differentiation is possible by use of PCR targeting the kinetoplast DNA gene more specifically,the maxicircle and minicircle genes. Pathologic tissue changes associated with the disease are poorly described so far. The necessity to impose management strategies for the control and eradication of dourine from endemic areas is in demand of more evidence-based data about more accurate feasible diagnostic measures and a better understanding of possible pathways of disease transmission. The aim of this thesis was to get more insights into the pathogenesis and venereal transmission of T. equiperdum (chapter 2). Histopathological lesions in four naturally T. equiperdum-infected horses in the chronical stage of dourine from Arsi-Bale highland of Ethiopia, exhibiting obvious clinical signs of dourine, were examined. Tissues collected at necropsy revealed on PCR test characteristics of T. equiperdum. A full post-mortem examination on the horses did not reveal typical gross lesions in the organs that were assumed to be responsible for the symptomatology. On histopathology, genital organs (especially testicles, uterus and vagina) were affected with mononuclear cell infiltration and erosions leading to degeneration of seminiferous tubules and perivascular lymphoplasmacytic cuffing in the uterus. In the nervous system, mononuclear cell infiltration was located in peripheral nerves and ganglia and in the white matter of the spinal cord, leading to axonal degeneration and fragmentation. Starting from tissue samples from the predilection sites of the parasite, real-time and conventional PCRs, . Clinical and haematological parameters were recorded and finally the effect of Cymelarsan® treatment given at the chronic stage of the disease, was assessed. At the same time we looked whether this treatment could alleviate the clinical signs and health status of the animal and induces differences in the pathology and tissue distribution of the parasite or not (chapter 4). Mares were artificially inseminated with T. equiperdum Dodola 943 spiked semen whereas stallions were infected by blood transfusion of the same strain. The course of the disease was monitored by parasitological (Woo), serological (CATT/T. evansi) and molecular (PCR) tests and clinical signs and haematological parameters were recorded. At 120 days post infection, they had a full necropsy for histopathology and PCR. A similar pattern of parasitaemia, disease progression and tissue distribution were seen in all horses with different routes of infection. Ejaculated semen in the preclinical stage and epididymal semen in the chronic stage of the disease was positive for T. equiperdum on PCR and caused infection in mice. Cymelarsan® treatment in the chronic stage did not result in a clinico-haematological or histopathological improvement. At necropsy, lesions were observed in the nervous and reproductive system. Histopathological lesions were most severe in the peripheral nerves and associated ganglia, the testicles and genital mucosae with multifocal infiltration of lymphocytes, plasma cells and histocytes. The parasites disseminated to several tissues including the nervous system, testicles and semen. Since treatment options for dourine are limited and results are disappointing, other measures should be taken into account in an attempt to eliminate dourine in endemic areas. Dourine is dealt with international legislative measures imposed by the World Organization for Animal Health (OIE) aimed at isolation, castration or slaughtering of positive horses. In developing countries like Ethiopia, it is economically and morally not feasible to impose strict slaughter policies to control dourine. When dealing with a disorder that is transmitted by venereal routes, it seems obvious that in an attempt to eliminate such a condition or even to diminish the morbidity of the disease, life cover should be replaced by artificial insemination. Purification of semen by single layer centrifugation (SLC) was attempted to remove trypanosomes from semen (Chapter 5). SLC has been proven to be successful in reducing venereal transmitted diseases when dealing with other pathogens. For this hypothesis semen was spiked using cryopreserved T. equiperdum stabilates (Dodola 943) in varying concentrations, from 102 to >5x106 trypanosomes/ml. Subsequently, SLC was performed following standard procedures. The presence of the parasite in the purified semen was checked by wet smear examination, ITS1 PCR and in vivo inoculation in mice. Before SLC, all spiked semen samples, except the negative controls, were positive on PCR analysis. After SLC, all the pellets were found to be negative for T. equiperdum on microscopic examinations. PCR analysis also could not detect any parasite-DNA in the SLC-pellet of semen spiked with the lower concentration (102 to 104 trypanosomes/ml). However, in the SLC pellets spiked with higher concentrations (104 - 5x104 trypanosomes/ml), only 1 out of the 4 replicates was negative for parasite DNA. All groups spiked with >5x104 trypanosomes were found to be positive on PCR. All mice in the positive controls exhibited parasitemia (5/5). Mice inoculated with SLC-purified semen that was spiked with lower than 5x104 trypanosomes remained free of parasitemia, similar to the negative controls. However, inoculation with SLC-pellets from samples with a higher number of trypanosomes (> 5x104 - 5x106 and > 5x106 /ml), induced parasitaemia in 2 out of 5 and 3 out of 5 mice, respectively. This study indicates that single layer centrifugation can be used to lear T. equiperdum infected semen but that the success is dependent on the concentration of the trypanosomes in the semen. Finally, all the obtained results were summarized and compared with the current knowledge and opinions and discussed in chapter 6. In conclusion, our results indicate that transmission of T. equiperdum is possible through semen and semen from symptomless stallions, in the preclinical-, clinical- and in the post-treatment periods. The results of the treatment trial to manage dourine especially at the chronic stage discourages the use of Cymelarsan®. The observed histopathological lesions in the reproductive organs, distal spinal cord and peripheral nerves might give an explanation to the diagnostic clinical incoordination of the hind legs in T. equiperdum-infected horses and its presence in the reproductive tract can exemplify the venereal transmission. Finally, our findings confirmed some of the assumptions about the pathogenesis and transmission of T. equiperdum and attributed to the growing knowledge about dourine in equines and trypanosomosis in general. When aiming to develop an all-encompassing eradication program, these insights may lead to a more efficient approach of dourine and thus add value to the agricultural and socio-economic situation in endemic areas