A comparative evaluation of parasitological tests and a PCR for Trypanosoma evansi diagnosis in experimentally infected water buffaloes

In this study five parasitological methods and a polymerase chain reaction (PCR) were compared for the diagnostic sensitivity for Trypanosoma evansi in experimentally infected water buffaloes over a period of 15 weeks. The combined estimates of sensitivity (CE(se)) of the PCR proved to be highest at 78.2%, closely followed by the mouse inoculation (MI), the micro-haematocrite centrifugation technique (MHCT) and the mini-anion-exchange centrifugation technique (MAECT) with CE(se) of, respectively, 74.0, 69.6 and 62.4%. The CE(se) of the buffy-coat technique (BCT) at 38.6% and the sodium dodecyl sulfate (SDS) clarification technique at 25.1% were considerably lower. PCR detected consistently all buffaloes infected from week 3 post-infection (PI) onwards. For MI this occurred after 5 weeks PI while for MHCT and MAECT these sustainable high levels were reached in the 7th week PI. BCT and SDS never detected all buffaloes infected. The influence of time and temperature on the viability of T. evansi in heparinized blood from water buffalo was also studied. In general we observed that the survival time tends to be longer when blood is kept at 4 degrees C. In samples kept in direct sunlight parasites became undetectable with the MHCT after 30min. After treatment of the water buffaloes with diminazene aceturate, the PCR signal disappeared within 24h.     

水牛伊氏锥虫人工感染抗体规律及其广西地区自然感染状况的初步调查

通过实验感染,研究伊氏锥虫在水牛体内抗体及虫体消长的规律.ELISA检测结果表明,水牛感染伊氏锥虫后第6 d检出抗体,而且在2个多月的监测时间里,抗体都维持在较高水平(OD450nm>0.3),因此可利用ELISA方法对水牛感染伊氏锥虫进行早期诊断和监测;水牛感染伊氏锥虫后,即出现一个2～3周左右的虫血症峰期,然后虫血症峰期迅速下降,在虫血症峰期下降后1个多月的监测时间里,虫血症维持在一个很低的水平(≤1条虫体/视野),水牛感染伊氏锥虫后这种峰期短无明显症状、虫血症水平低的现象,给临床诊断和血液镜检虫体造成了一定的困难.对广西部分地区79份血清样品水牛检测的结果表明,伊氏锥虫抗体阳性率达到29.1%,说明广西各地水牛感染伊氏锥虫的现象普遍存在,而且感染率比较高,为了保证本地区水牛产业的健康发展,有必要对水牛伊氏锥虫感染进行有效的监测.     

Evaluation of the card agglutination test (CATT/T. evansi) for detection of Trypanosoma evansi infection in water buffaloes (Bubalus bubalis) in Egypt

A card agglutination test (CATT/T. evansi) was evaluated for detection of antibodies against Trypanosoma evansi (T. evansi) in experimentally and naturally infected buffaloes. Four calves were inoculated with a strain of T. evansi isolated from a dromedary camel. Parasitological examination of the calves revealed trypanosomes in the blood from days 4 to 9 post-inoculation (PI). General emaciation appeared from day 26 PI and aggravated until the end of the experiment (day 88 PI). Antibodies against T. evansi were detectable from day 8 PI till the end of the experiment. Parasitological examination of 200 water buffalo blood samples obtained from slaughterhouses revealed negative results. Serological examination of these animals showed that 48 (24%) water buffaloes had anti-T. evansi antibodies.     

Trypanosoma evansi infection in buffaloes in North-east Thailand. II. Abortions

Summary By exclusion of other possible aetiological agents, strong circumstantial evidence is presented ofTrypanosoma evansi infection being the cause of late gestation abortion and stillbirth in buffaloes.

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Resumen Se presentan evidencias circunstanciales de infecciones porTrypanosoma evansi, como causa de abortos y momificaciones al final de la gestación en 32 búfalos.

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Résumé Par exclusion de r?le étiologique possible d’autres agents, de fortes preuves indirectes sont présentées qui indiquent que l’infection parTrypanosoma evansi est la cause d’avortements en fin de gestation et de mortinatalités chez 32 bufflesses.

     

Study on the antioxidant status of rats experimentally infected with Trypanosoma evansi

The antioxidant status of rats experimentally infected with Trypanosoma evansi isolated from a camel was studied using established parasitological, haematological and biochemical methods. The results indicated that infections in all rats resulted in a fulminating parasitaemia. Changes in blood parameters in T. evansi-infected rats indicated leukocytosis and a macrocytic hypochromic anaemia. A degree of anisocytosis was also observed. The activities of plasma glucose-6-phosphate dehydrogenase and glutathione peroxidase in whole blood of infected rats were significantly higher (p<0.05 and p<0.001, respectively) compared with control. No statistically significant difference was observed in the activity of superoxide dismutase in infected and control rats. Results obtained indicated that trypanosomosis caused oxidative stress and induced antioxidant enzymes.     

Trypanosoma evansi infection in buffaloes in north-east Thailand. I. Field investigations

Clinical, parasitological and serological findings of Trypanosoma evansi infections in buffaloes (Bubalis bubalis) from north-eastern Thailand are reported. The overall infection rate was found to be around 20% with a distinct peak of acute infections during the rainy season. The disease is aggravated by normally well tolerated concomitant infections such as liver fluke infestations and by other stress factors.     

Trypanosoma evansi infection in buffaloes in north-east Thailand. I. Field investigations

Summary Clinical, parasitological and serological findings ofTrypanosoma evansi infections in buffaloes (Bubalis bubalis) from north-eastern Thailand are reported. The overall infection rate was found to be around 20% with a distinct peak of acute infections during the rainy season. The disease is aggravated by normally well tolerated concomitant infections such as liver fluke infestations and by other stress factors.

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Resumen Se discuten los resultados clínicos, parasitológicos y serológicos, de infecciones porTrypanosoma evansi en búfalos (Bubalis bubalis), en el noroeste de Tailandia. La rata de infección general fue del 20%, con aumento de infecciones agudas durante la estación lluviosa. La enfermedad se agrava cuando existen infecciones concominanates, como la fascioliasis y otros factores de estrés.

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Résumé Les observations cliniques, parasitologiques et sérologiques sur les infections àTrypanosoma evansi chez les buffles (Bubalis bubalis) dans le Nord-Est de la Tha?lande sont rapportées. Le taux général d'infection se situe autour de 20 p. 100 avec un pic distinct d'infections aigues pendant la saison des pluies. La maladie est aggravée par des infections concomitantes normalement bien tolérées telles que les distomatoses et par les autres facteurs de stress.

     

Ocular lesions associated with Trypanosoma evansi in experimentally infected goats

Ocular lesions associated with Trypanosoma spp. infection have been described in man and many animal species. However, loss of vision has not been demonstrated in humans presenting Chagas disease or in animals affected by different trypanosome species. In order to assess the possible ocular disorders caused by Trypanosoma evansi infection, six goats were inoculated with 1 x 10(5) T. evansi and maintained for 12 months and four goats were used as control. The inoculated animals became positive at serological and parasitological tests at 1-month post-inoculation and showed a subclinical course of the disease. Unilateral superficial corneal ulceration and retinochoroiditis were observed in two inoculated animals. Data from ocular neurologic examination and electroretinography showed no significant differences between inoculated and non-inoculated goats. It could be concluded that Trypanosoma evansi can produce ocular lesion but without apparent loss of vision in goats.     

Tropical liver fluke-induced stress in experimentally infected and immunised buffaloes

Disease-related stress associated with Fasciola gigantica infection was investigated in 16 male, yearling Murrah buffaloes. The animals were randomly assigned to one of four groups. Animals in groups 1 and 2 were vaccinated subcutaneously with 4.8 mg of excretory-secretory antigen (group 1) or 1300 μg infection-specific antigen (group 2), in three doses. Subsequently, all the animals in groups 1, 2 and 3 were infected orally with 800 viable F gigantica metacercariae (bubaline origin) on week 6 of the experiment. The animals in group 4 served as healthy controls. The clinical progress of the disease with respect to the adrenocortical response to infection was assessed periodically. The infected animals in group 3 developed characteristic signs of the disease and had the highest in situ fluke population (mean [se] 331.8 [19.5] ). One of the four animals in group 3 died on day 110 postinfection (PI). Animals that had been immunised before infection (groups 1 and 2) acquired adequate levels of immunity, were comparatively healthier and had significantly lower (P<0.05) fluke populations (mean [se] 194.3 [11.8] in group 1 and 164.5 [9.2] in group 2). Throughout the course of the disease, the group 3 animals had significantly higher cortisol levels than those in groups 1 and 2 (P<0.01) from week 4 PI onward. Although animals in the immunised groups (groups 1 and 2) had increased hormone levels, they were not significantly different from those in the control animals in group 4. Cortisol levels were higher during the early prepatency phase (weeks 1 to 6 PI) than during the late prepatency and/or patency phases of the disease. Cortisol levels in the healthy control animals in group 4 remained within the normal range.     

Trypanosoma evansi infection in cattle, buffaloes and horses in Indonesia

Cattle, buffaloes and horses in several areas of Indonesia were examined for evidence of infection with Trypanosoma evansi by the microhaematocrit centrifugation technique (MHCT) and an enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to T. evansi. Evidence of infection was found in animals at each sampling site although differences were seen in prevalence rates between sites. Prevalence rates in buffalo were usually higher than in cattle in the same area while in horses they were much lower than in cattle or buffalo. An age-dependent prevalence rate was seen in buffalo and cattle with the highest rates seen in animals older than 2 years. These results concur with the view that T. evansi infection is widespread throughout most of the livestock-producing areas of Indonesia. The apparent lack of any obvious disease owing to T. evansi infection in the sampled animals suggests that a form of stability exists in most endemic areas which serves to ameliorate the effect of T. evansi infection and has an immunological basis linked to the parasite's limited antigenic diversity.     

Serological responses to Neospora caninum in experimentally and naturally infected water buffaloes (Bubalus bubalis)

The water buffalo (Bubalus bubalis) is an important natural host for Neospora caninum. Serologic responses to N. caninum were studied in experimentally and naturally infected water buffaloes in Brazil. Antibodies were assayed by the indirect fluorescent antibody test (IFAT) using a cut off value of 1:25. Six buffaloes were each inoculated subcutaneously with 5 x 10(6) live culture-derived tachyzoites of the cattle Illinois strain of N. caninum, and two calves were kept as uninoculated controls. Post-inoculation (p.i.) blood samples were collected weekly for 8 weeks and then monthly until 1 year p.i. All inoculated buffaloes developed IFAT titers of 1:100 or more between 7 and 11 days p.i. and the titers remained elevated until 7 weeks p.i. Antibody titers peaked to 1:1600 in 1, 1:800 in 3 and 1:400 in 2, usually by 3 weeks p.i. Antibody titers declined to 1:25 or 1:50 in all the six buffaloes by 12 months p.i. IFAT titers to N. caninum remained at an undetectable level (< 1:25) in both control uninoculated buffaloes. To follow the dynamics of N. caninum antibodies, sera from 29 buffaloes and their calves were collected for 1 year and assayed for N. caninum antibodies; 23 of 29 calves were seropositive (IFAT of 1:100 or more) at 1-2 day of age. Of these 23 calves, 17 remained seropositive during the study, while six became seronegative at four (two calves), six (one calf) seven (two calves) and eight (one calf) months of age. These findings suggest a high rate of neonatal transmission of N. caninum in buffaloes.     

Preliminary evaluation of diagnostic tests using horses experimentally infected with trypanosoma evansi

Seven surra negative horses were intravenously inoculated with 3 x 10(6)Trypanosoma evansi parasites derived from a camel. One horse was maintained as an uninfected negative control. Three antigen and three antibody detection tests were evaluated for diagnosis of infection in horses. The microhaematocrit centrifugation test (MHCT) was the most sensitive, first detecting parasites between one and three days (x 2.4) post infection (p.i.). The antigen (ag)-ELISA detected antigen between three and ten days (x 6.6) p.i. The latex agglutination test (LAT) first gave positive results on day 3 (x 3.0) p.i. Following the treatment of horses with trypanocidal drugs, the MCHT and the mouse inoculation test (MIT) became negative. Antigen levels using LAT declined and reached pre-infection levels in five out of six horses during the period of observation (92-279 days). Antigen levels using the ag-ELISA declined as well but did not reach pre-infection levels in any of the six horses.Three antibody detection techniques, ab-ELISA, card agglutination test (CATT), and immunofluorescent antibody test (IFAT) detected antibodies in the blood of all seven infected horses but not in the uninfected control. However, the ab-ELISA did not discriminate clearly between sera from infected and uninfected horses because unacceptably high ELISA background readings were detected in 15% of the surra negative horses shipped to the UAE from the UK. The ELISA antibody increased above pre-infection levels in the six horses experimentally infected, but not in one horse. In this horse the ELISA antibody level exceeded the cut-off level only after the reoccurrence of the T. evansi infection. The IFAT detected antibodies 15.7 days p.i. in all infected horses.     

Expression of regulatory dendritic cell-related cytokines in cattle experimentally infected with Trypanosoma evansi

Trypanosoma evansi causes wasting disease in many livestock. T. evansi infection gives rise to inflammatory immune responses, which contribute to the development of inflammation-associated tissue injury. We previously reported that regulatory dendritic cells (DCs), which act as potential regulators of inflammation, were activated in infected mice and transfer of regulatory DCs to infected mice prolonged their survival. However, the kinetics of regulatory DCs in cattle, which are natural hosts of T. evansi, remained unclear. In this study, we report that the expressions of CCL8 and IL-10, which promote the development of regulatory DCs, were up-regulated in cattle experimentally infected with T. evansi. This finding is potentially useful for studying the control strategy of T. evansi infection in cattle.     

Susceptibility of Trypanosoma evansi to propolis extract in vitro and in experimentally infected rats

Current therapy of Trypanosoma evansi infections is not effective for the vast majority of animals with relapsing parasitemia and clinical signs. Recently, attention is being focused on the antiparasitic activity of propolis. This study evaluated the susceptibility of T. evansi to propolis extract in vitro and in vivo. A dose-dependent trypanocidal activity of propolis extract was observed in vitro. All trypomastigotes were killed 1h after incubation with 10μgmL(-1) of the extract. In vivo, the concentrations of 100, 200, 300 and 400mgkg(-1) administered orally for 10 consecutive days showed no curative effect, and the rats died from the disease. However, rats treated with the two highest concentrations of propolis extract showed higher longevity than the other groups. Based on these data, we concluded that T. evansi is susceptible to propolis in vitro. Despite the lack of curative efficacy observed in vivo at the concentrations tested, the propolis extract can prolong life in rats infected with the protozoan.     

山羊、水牛感染肝片吸虫后淋巴细胞自然杀伤活性动态变化

自然杀伤 (NK)活性是在细胞水平反映机体天然免疫力的重要指标之一。本实验以SP2 / 0细胞为靶细胞 (即效应细胞 ) ,采用乳酸脱氢酶(LDH)短程释放法测定人工感染肝片吸虫的水牛和山羊的外周血液中淋巴细胞的NK活性动态变化。从测定结果的相对值来看 ,发现山羊在感染肝片吸虫后NK活性有 2次高峰 ,推测与虫体的发育或感染山羊的免疫状态有关 ;2个感染组 (Ⅰ ,2 0 0个囊蚴 /只 ;Ⅱ ,50 0个囊蚴 /只 )的变化趋势表明 ,感染量与NK活性无相关关系。感染水牛的NK值仅略高于对照组 ,无明显的高峰。而绝对值结果又反映出不同动物的NK活性有差异 ,水牛的NK活性比山羊高 ,说明水牛对肝片吸虫的天然免疫力比山羊强     

Gametogony and sporogony of Sarcocystis fusiformis of buffaloes in the small intestine of experimentally infected cats

The gametogonous and sporogonous stages of Sarcocystis fusiformis were studied in the small intestine of 10 cats each fed 350 macroscopic sarcocysts and killed 12 h, 3, 5, 7, 9, 11, 13, 15 and 17 days post infection. The macrogamonts were observed from 12 h up to 9 days post infection, whereas microgamonts were seen only on Day 3 and were very few in number. Developing oocysts were seen from Day 5 to Day 11 post infection, and from Day 13 fully sporulated oocysts and sporocysts were found in increasing numbers.     

Comparison of serological tests for Trypanosoma evansi natural infections in water buffaloes from north Vietnam

In the present study, a collection of 415 water buffalo serum samples originating from the north of Vietnam was used for evaluation of different diagnostic antibody detection methods available to detect infections with Trypanosoma evansi. The diagnostic sensitivity and specificity of a direct card agglutination test (CATT/T. evansi), an indirect card agglutination test (LATEX/T. evansi) and a newly developed antibody detection ELISA (ELISA/T. evansi) was calculated on the basis of parasitological results, obtained by mouse inoculation, and compared for all assays. The immume trypanolysis assay with the predominant T. evansi RoTat 1.2 variable antigen type was used as reference test for antibody presence. All parasitologically confirmed animals (n=8) were positive in all tests. Diagnostic specificity was highest in CATT/T. evansi (98%) followed by the ELISA/T. evansi (95%) and the LATEX/T. evansi (82%). Concordance of the variant specific immune trypanolysis test with the other tests was calculated and revealed that few (1-8%) false positive results were actually due to a specific reactions, and that LATEX/T. evansi and ELISA/T. evansi detected more immune trypanolysis positives than the CATT/T. evansi. It was concluded that, apart from the immune trypanolysis test, which is not generally applicable, ELISA/T. evansi with a 30% positivity cut-off and LATEX/T. evansi, thanks to their superior capacity of detecting T. evansi specific antibodies, would be suitable as epidemiological tools detecting both active infections and persisting T. evansi specific antibodies. The ELISA/T. evansi with a 50% positivity cut-off and the CATT/T. evansi on the other hand, seem more appropriate to detect true infected water buffaloes.     

The effect of albendazole on nematode parasites in experimentally infected lambs.

Thirty-five complete oestrous cycles were studied in five nonpregnant camels (Camelus dromedarius) over a period of 15 months. The oestrous cycle did not have a luteal phase. During the cycle of 28 days the ovarian activity was strictly follicular. Follicles matured in six days, maintained their size for 13 days and regressed in eight days. Ovulation was non-spontaneous and required the stimulus of coitus. Manual stimulation of the cervix for 15 minutes did not induce ovulation. The external manifestations of the oestrous cycle as well as the changes in the ovaries and the genital tract were also studied.