Development of Slide Enzyme Linked Immunosorbent Assay (SELISA) for Detection of Trypanosoma evansi Infection in Bovines
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The Slide enzyme linked immunosorbent assay (SELISA) was standardized for detection of antibodies specific to Trypanosoma evansi and subsequently used for the screening of naturally infected bovine sera. A novel SELISA, a modification of the standard ELISA technique was used for the detection of antibodies against Trypanosoma evansi in bovines using positive and negative control sera. The test is based on immunostaining of the fixed whole Trypanosoma evansi organisms on microscopic glass slide, incubation with sera, antibovine IgG-HRPO conjugate and substrate Diaminobenzidine tetrahydrochloride (DAB). Finally the reaction was read under oil immersion of microscope. A total of 702 sera samples from bovines in Rayalaseema region of Andhra Pradesh were examined by SELISA and 192 were found positive for Trypanosoma evansi antibodies.Keywords:
Trypanosoma evansi
As parasitaemia is low and fluctuating during the chronic stage of infection, accurate detection of Trypanosoma evansi in blood is difficult. The primary aims of this investigation were to assess for the first time the seroprevalence of T. evansi in all agro-climatic zones of Punjab, by indirect enzyme-linked immunosorbent assay (iELISA) and card agglutination test (CATT/T. evansi), and to evaluate the risk factors associated with latent trypanosomosis. A total of 319 equine serum samples collected from 12 districts of Punjab (India) belonging to different agro-climatic zones revealed 39 (12.23%) and 9 (2.82%) samples to be positive by CATT/T. evansi and iELISA, respectively. The highest prevalence was recorded from the Ludhiana district (42.86% and 7.14% by CATT/T. evansi and iELISA, respectively) in the central plain zone (for which the overall prevalence was 15% and 4.17%, respectively). There was fair agreement between the tests for the detection of T. evansi (kappa = 0.345). Species was the most influential risk factor for infection, with odds ratios (ORs) of 2.81 and 5.63 for donkeys/ mules, in comparison with horses, by CATT/T. evansi and iELISA, respectively. The female equine population (OR = 3.13, 95% confidence interval [CI] = 1.32-7.67 [CATT]) was found to be at a higher risk of seropositivity for T. evansi, particularly on 'unorganised' (inappropriately managed) farms (OR = 3.18, 95% CI = 1.53- 6.65 [CATT]) and among animals used for commercial purposes (OR = 2.51, 95% CI = 1.20-5.21 [CATT]). In conclusion, to declare disease-free status, use of the iELISA followed by retesting of suspect samples by CATT/T. evansi is suggested.La détection minutieuse de Trypanosoma evansi dans le sang est difficile en raison du nombre faible et fluctuant de parasites pendant la phase chronique de l’infection. L’étude présentée par les auteurs vise, d’une part, à réaliser une première évaluation de la prévalence sérologique de T. evansi dans chacune des zones agro-climatiques du Pendjab en utilisant une épreuve immuno-enzymatique (ELISA) indirecte et le test d’agglutination sur carte pour la trypanosomose (CATT/T. evansi) et, d’autre part, à évaluer les facteurs de risque associés à une présence inapparente de la trypanosomose. Au total, sur les 319 sérums d’équidés prélevés dans 12 districts du Pendjab (Inde) appartenant à des zones agro-climatiques différentes, 39 échantillons (12,23 %) ont donné des résultats positifs avec le CATT/T. evansi et 9 échantillons (2,82 %) ont donné des résultats positifs à l’ELISA indirecte. La prévalence la plus élevée a été enregistrée dans le district de Ludhiana (42,86 % de résultats positifs avec le CATT/T. evansi et 7,14 % de résultats positifs avec l’ELISA indirecte) dans la zone des plaines centrales (où la prévalence globale s’élevait, suivant les méthodes de test, à 15 % et 4,17 %, respectivement). La détection de T. evansi par les deux tests a été concordante (kappa = 0,345). Le facteur de risque ayant le plus d’influence sur la probabilité d’infection était l’espèce, ce risque étant plus élevé chez les ânes et les mulets que chez les chevaux (rapport de cotes [odds ratio, OR] de 2,81 [CATT/T. evansi] et de 5,63 [ELISA indirecte]). Les femelles présentaient également un risque plus élevé de posséder des anticorps anti-T. evansi que les mâles (OR = 3,13 ; intervalle de confiance [IC] à 95 % : 1,32–7,67 [CATT]), en particulier dans les élevages « informels » (sans gestion sanitaire) (OR = 3,18 ; IC à 95 % : 1,53–6,65 [CATT]) ainsi que parmi les animaux utilisés à des fins commerciales (OR = 2,51 ; IC à 95 % : 1,20–5,21 [CATT]). En conclusion, pour la démonstration de l’absence d’anticorps, les auteurs recommandent d’utiliser l’ELISA indirecte puis de soumettre les échantillons douteux à un test de confirmation au moyen du CATT/T. evansi.La detección precisa de Trypanosoma evansi en la sangre resulta difícil porque en la fase crónica de la infección la parasitemia es baja y fluctuante. Los autores describen una investigación encaminada principalmente a determinar por primera vez la seroprevalencia de T. evansi en todas las zonas agroclimáticas del Punjab por ensayo inmunoenzimático indirecto (ELISAi) y por aglutinación en placa, así como los factores de riesgo asociados a la tripanosomosis latente. De un total de 319 muestras de suero equino procedentes de 12 distritos del Punjab (India) situados en diferentes zonas agroclimáticas, la aglutinación en placa deparó resultado positivo en 39 de ellas (un 12,23%) y el ELISAi en 9 (2,82%). El máximo nivel de prevalencia se registró en el distrito de Ludhiana (42,86% y 7,14% por aglutinación en placa y ELISAi, respectivamente), sito en la zona de la planicie central (que en conjunto deparó una prevalencia del 15% y el 4,17%, respectivamente). Ambas pruebas resultaron bastante coincidentes por lo que respecta a la detección de T. evansi (coeficiente kappa = 0,345). El factor de riesgo más influyente resultó ser la especie: en comparación con los caballos, los asnos o mulas presentaban una razón de probabilidad (RP) de 2,81 y 5,63 para la aglutinación en placa y el ELISAi respectivamente. Se observó que la población de yeguas (RP = 3,13; intervalo de confianza [IC] al 95% = 1,32–7,67 [aglutinación en placa]) presentaba un riesgo más elevado de seropositividad para T. evansi, especialmente en explotaciones «desorganizadas» (mal gestionadas) (RP = 3,18; IC 95% = 1,53–6,65 [aglutinación en placa]) y entre los animales utilizados con fines comerciales (RP = 2,51; IC 95% = 1,20–5,21 [aglutinación en placa]). Los autores concluyen proponiendo que, a los efectos de declarar la ausencia de enfermedad, se utilice en primer lugar el ELISAi, seguido de la prueba de aglutinación en placa para las muestras sospechosas.
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We report an Indian farmer who had fluctuating trypanosome parasitemia associated with febrile episodes for five months. Morphologic examination of the parasites indicated the presence of large numbers of trypanosomes belonging to the species Trypanosoma evansi, which is normally a causative agent of animal trypanosomiasis known as surra. Basic clinical and biologic examinations are described, using several assays, including parasitologic, serologic, and molecular biologic tests, all of which confirmed the infecting species as T. evansi. Analysis of cerebrospinal fluid indicated no invasion of the central nervous system (CNS) by trypanosomes. Suramin, a drug used exclusively for treatment of early-stage human African trypanosomiasis with no CNS involvement, effected apparent cure in the patient. This is the first case reported of human infection due to Trypanosoma evansi, which was probably caused by transmission of blood from an infected animal.
Trypanosoma evansi
Trypanosoma vivax
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African animal trypanosomosis, transmitted cyclically by tsetse flies or mechanically by other biting flies, causes serious inflictions to livestock health. This study investigates the extent of non-tsetse transmitted animal trypanosomosis (NTTAT) by Trypanosoma (T.) evansi and T. vivax in domestic animals in the tsetse-free regions of Northern Ethiopia, Afar and Tigray.A cross sectional study was conducted on 754 dromedary camels, 493 cattle, 264 goats, 181 sheep, 84 donkeys, 25 horses and 10 mules. The microhaematocrit centrifugation technique was used as parasitological test. Plasma was collected for serodiagnosis with CATT/T.evansi and RoTat 1.2 immune trypanolysis (ITL) while buffy coat specimens were collected for molecular diagnosis with T. evansi type A specific RoTat 1.2 PCR, T. evansi type B specific EVAB PCR and T. vivax specific TvPRAC PCR.The parasitological prevalence was 4.7% in Tigray and 2.7% in Afar and significantly higher (z = 2.53, p = 0.011) in cattle (7.3%) than in the other hosts. Seroprevalence in CATT/T.evansi was 24.6% in Tigray and 13.9% in Afar and was significantly higher (z = 9.39, p < 0.001) in cattle (37.3%) than in the other hosts. On the other hand, seroprevalence assessed by ITL was only 1.9% suggesting cross reaction of CATT/T.evansi with T. vivax or other trypanosome infections. Molecular prevalence of T. evansi type A was 8.0% in Tigray and in Afar and varied from 28.0% in horses to 2.2% in sheep. It was also significantly higher (p < 0.001) in camel (11.7%) than in cattle (6.1%), donkey (6%), goat (3.8%), and sheep (2.2%). Four camels were positive for T. evansi type B. Molecular prevalence of T. vivax was 3.0% and was similar in Tigray and Afar. It didn't differ significantly among the host species except that it was not detected in horses and mules.NTTAT caused by T. vivax and T. evansi, is an important threat to animal health in Tigray and Afar. For the first time, we confirm the presence of T. evansi type B in Ethiopian camels. Unexplained results obtained with the current diagnostic tests in bovines warrant particular efforts to isolate and characterise trypanosome strains that circulate in Northern Ethiopia.
Trypanosoma evansi
Trypanosoma vivax
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Buffy coat
Veterinary parasitology
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Trypanosoma evansi
White (mutation)
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During an outbreak of trypanosomiasis in cattle, buffalo and horses on the island of Madura, a survey was conducted to determine the prevalence of Trypanosoma evansi in the region. Blood samples were collected from 130 cattle and 147 buffalo in 5 villages, whole blood was examined for trypanosomes by the microhaematocrit (MHCT) method and serum samples were subjected to an enzyme linked immuno absorbent assay to detect T. evansi antibodies. T. evansi was detected by MHCT in 50% of the buffalo examined and 13% of the cattle, while antibodies were detected in 47% of the buffalo and 30% of the cattle. The outbreak was brought under control following the administration of Nagano (Suramin, Bayer) to infected animals and animals considered to be at risk of infection.
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The first reported human case of trypanosomiasis caused by Trypanosoma evansi was treated using suramin. Patient follow-up indicates that the drug and specific regimen used were well tolerated. Clinical, serological and parasitological investigations at 6 months indicate complete cure of the patient. Suramin should be considered in the treatment of other cases of human T. evansi infection, if they occur.
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Nifurtimox
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Trypanosoma evansi is a parasite that causes surra in a variety of wild and domestic animals and is mainly transmitted by biting flies in Africa, Asia and Latin-America. Horses infected by Trypanosoma evansi present a chronic weight loss, icterus, oedema, anemia, abortions and neurological troubles. Due to this parasite, cases of human trypanosomiosis have been reported in different countries by contacting with infected animals. In this study, 206 healthy equines (177 horses and 29 donkeys) from El-Bayadh district, located in southwest Algeria, were tested for the presence of parasites in blood using Giemsa-stained blood films and for the presence of antibodies against T. evansi using CATT /T. evansi. While none of the equines showed detectable parasites in the blood, the individual seroprevalence of T. evansi was found to be 46.6% (CI 95%, 40.7-54.4%). Out of 98 positives samples, 56.1% (55/98) were shown at level 1 (+), 27.5% (27/98) at level 2 (++) and 16.3% (16/98) at level 3 (+++). The results show that out of 177 tested horses, 80 were seropositive to T. evansi, 45.2% (CI 95%, 37.8-52.5%) and out of 29 tested donkeys, 18 were seropositive to T. evansi, 62.1% (CI 95%, 44.4-79.7%). A questionnaire for the owners, targeted to associate risk factors for surra in horses, showed that environmental factors that are favorable for Tabanids, such as water and vegetation, but also promiscuity with dromedaries were positively associated with the seroprevalence rate in the horses. El-Bayadh district is a highly endemic region for surra in Algeria.
Trypanosoma evansi
Equidae
Equus asinus
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Trypanosoma evansi is a pathogenic African animal protozoan, affecting livestock and wild animals worldwide and Iran. The present study was carried out to detect the infection of T. evansi in one-humped camels (Camelus dromedarius) of southeastern Iran. Over six months, a total numbers of 370 dromedary camels were randomly selected from three sub-areas located in southeastern Iran in 2015. Blood samples were taken from jugular vein and examined by using micro-hematocrit centrifugation (MHCT) and PCR techniques. Genomic DNA was extracted and PCR was performed to amplify a fragment of the mini-chromosome satellite DNA TBR1/2 of T. evansi. The overall prevalence was 31.35% (116/370). The highest T. evansi infection was significantly in adult camels (24.05%, 89/370) with chronic clinical signs (11.89%, 44/370). There was significant difference between prevalence and sex (9.46% male and 21.89% female). Only 19.19% (71 out of 370) of the infected camels were from the plain areas. The highest T. evansi infection rate was significantly recorded in the camels of north (19.19%, 71/370) part of the region. The molecular analysis was uncovered high level of infection with T. evansi in camel herds which can help to establish effective control programs in the region.
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