Evaluation of antigen and antibody rapid detection tests for Trypanosoma evansi infection in camels in Kenya

The card agglutination test for Trypanosoma evansi (CATT/T. evansi) for the detection of antibodies, and Suratex for the detection of circulating antigens were compared in a cross-sectional study involving camels in eastern and central parts of Kenya. Of the 2227 camels screened, 2038 were owned by nomadic pastoralists in T. evansi endemic areas in eastern Kenya. A herd of 86 camels were from a ranch in Mugwoni. In Athi River area, 35 camels belonged to Kenya Trypanosomiasis Research Institute, and 68 were slaughter animals. Diagnostic sensitivity estimates were obtained by testing sera from 51 camels that had been found to be parasitologically positive by the haematocrit centrifugation technique, buffy-coat technique and mouse inoculation. Diagnostic specificity was estimated by testing sera from 35 camels known to be trypanosome-free. Positive and negative predictive values (NPVs) were calculated using a range of prevalence values. The sensitivity of CATT/T. evansi (68.6%) was higher than that of Suratex (58.8%), but not significantly. Both tests had equally high specificity (100%). The overall prevalence was 2.3% (51 out of 2227) by parasite detection, 32.2% (327 out of 1017) by CATT/T. evansi and 19.6% (188 out of 961) by Suratex. Overall, there was a positive association between CATT/T. evansi and Suratex though the strength of association was low (McNemar's test=46.12, P=0.001; kappa=0.26, CI: 0.20-0.33). Parasite prevalence ranged from 0% in several herds to 27.8% in a herd in Isiolo. Prevalence was highest in Isiolo with 2.5% (51 out of 2030) by parasitological detection, 38.8% (321 out of 828) by CATT/T. evansi and 21.9% (169 out of 772) by Suratex. In Mugwoni prevalence was 7 and 18% by CATT/T. evansi and Suratex, respectively, and no parasites were detected. In Athi River Suratex detected 2.9% (3 out of 103) positive while CATT/T. evansi and parasitological methods gave negative results. At prevalence values between 10 and 100%, CATT/T. evansi as well as Suratex had infinitely high positive predictive values, whereas Suratex had a lower NPV than CATT/T. evansi. In conclusion, results of this study showed that CATT/T. evansi and Suratex were able to detect aparasitaemic infections rapidly and were more sensitive than parasitological methods in revealing the true extent of trypanosomosis in a herd. The tests effectively complemented parasitological methods in the detection of T. evansi infections in camels.     

Animal-level risk factors for Trypanosoma evansi infection in camels in eastern and central parts of Kenya

Point prevalences and animal-level risk factors for Trypanosoma evansi infection were investigated in a cross-sectional study that involved 2227 camels from eastern and central parts of Kenya. The screening tests used were haematocrit centrifugation technique (HCT), mouse inoculation and latex agglutination (Suratex). All camels were screened with HCT, while 396 and 961 of them were, in addition, screened with mouse inoculation and Suratex tests, respectively. Parasitological and Suratex test results were used in parallel to determine the number of camels exposed to T. evansi infections. Statistical analyses were conducted using Statistical Analysis Systems. Parasitological and Suratex test results in parallel were dependent variables in multivariable logistic regression models that determined risk factors for T. evansi infection. Herd-level clustering was corrected with general estimation equations. The prevalences were 2.3% and 19.6%, using parasitological and Suratex tests, respectively, and 21.7% when both tests were used in parallel. There was a positive association between the screening tests (McNemar's test = 104.8, P = 0.001) although the strength of association was low (Kappa = 0.2; 95% CI: 0.1-0.3). Before accounting for herd-level clustering, dry season (OR = 1.5; 95% CI: 1.0, 2.1) and nomadic pastoralism (OR = 1.8; 95% CI: 1.1, 3.2) were associated with increased odds of a camel being exposed to T. evansi infection compared to wet season and ranching, respectively. Following this correction, only nomadic pastoralism was significantly associated (OR = 3.1; 95% CI = 1.0, 14.4) with T. evansi infection compared to ranching. It is concluded that camels managed under nomadic pastoralism had higher risk of being exposed to T. evansi infections than camels from ranching systems of management.     

PCR amplification of RoTat 1.2 VSG gene in Trypanosoma evansi isolates in Kenya

A direct card agglutination test for Trypanosoma evansi, CATT/T. evansi based on the predominant variable antigen-type (pVAT) RoTat 1.2 was evaluated previously in the field in Isiolo District, Kenya. Sixteen out of 51 (31.4%) parasitologically positive camels were negative by the antibody detection test. In the present study, trypanosomes isolated from the camels were analysed in an attempt to determine the cause of the false negative results of CATT/T. evansi. A total of 20 field isolates comprised 16 stocks from camels that were negative by CATT/T. evansi, and 4 from CATT/T. evansi-positive camels. In addition, 15 known T. evansi and four T. brucei were used as reference. Purified DNA samples were tested using an established RoTat 1.2-based polymerase chain reaction (PCR) that yields a 488 bp product for the specific detection of T. evansi. Antibodies to RoTat 1.2 variant surface glycoprotein (VSG) were used in Western blotting to detect RoTat 1.2 VSG linear epitopes. Results of PCR and Western blot showed that the 16 stocks isolated from CATT/T. evansi-negative camels fell into three groups. In Group 1, both the RoTat 1.2 VSG gene and the VSG were absent in three stocks. In five trypanosome stocks in Group 2, the RoTat 1.2 VSG gene was detected, but Western blot was negative indicating absence of the expressed VSG. Five other stocks containing the RoTat 1.2 VSG gene were also in this group. The RoTat 1.2 VSG gene was detected and Western blot was positive in all four trypanosome stocks in Group 3. All four stocks from CATT/T. evansi-positive camels contained the RoTat 1.2 VSG gene and the expressed VSG. The reference T. evansi KETRI 2479 lacked the RoTat 1.2 VSG gene and there was no immune reactivity detected by Western blot. The rest of the reference T. evansi stocks examined contained the RoTat 1.2 VSG gene. All the four T. brucei samples examined were negative by PCR and Western blot. In conclusion, this study showed that the RoTat 1.2 VSG gene was absent from some T. evansi trypanosomes in Kenya.     

Prevalence and distribution of Trypanosoma evansi in camels in Somaliland

Tropical Animal Health and Production - The prevalence and distribution of Trypanosoma evansi (T. evansi) infection on camels in Somaliland were studied using the card agglutination test (CATT/T....     

First outbreak of Trypanosoma evansi in camels in metropolitan France

The first outbreak of trypanosomosis caused by Trypanosoma evansi in camels in France was reported on a farm in the Aveyron Department. Five camels were imported from the Canary Islands to the farm in early July 2006, and trypanosomes were observed on a stained blood smear from one of them, which died in October. On further investigations, trypanosomes were observed in the blood of five camels, three of them indigenous to the farm and two that had been imported. On the basis of microscopical examination (morphological criteria and measurements) and serological results based on the card agglutination T evansi test and PCR typing, the parasites were identified as T evansi. After treatment with melarsomine, the infected camels rapidly became negative by parasitological tests and were negative two to four months later by serological tests. The parasite was probably transmitted by tabanids and Stomoxys calcitrans, which were abundant in July to September 2006. No parasites were observed in other animals on the farm or on neighbouring farms, but some of the sheep on these farms were positive by PCR or serology.     

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.     

Prevalence and infection pattern of Trypanosoma evansi in camels in mid-eastern Sudan

The antigen detection enzyme immunoassay (AgELISA) in conjunction with parasitological examination of blood were used to study the enzootic situation of cameline trypanosomiasis in mid-Eastern Sudan. A one year survey showed that the infection is endemic among pastoral camels with a prevalence of 5.4% based on parasitological examination and 31.3% based on AgELISA. The infection rate was higher during the dry period (November to May) than the wet season. Young camels had a much lower infection rate as detected by parasitological techniques, but not with AgELISA. A lower prevalence of infection was detected by buffy coat technique (BCT) in herds of camels raised by nomads compared with those kept by agropastoralists and in camels located in the southern districts of mid-Eastern Sudan.     

Camel trypanosomosis in the Canary Islands: assessment of seroprevalence and infection rates using the card agglutination test (CATT/T. evansi) and parasite detection tests

Trypanosomosis due to Trypanosoma evansi (surra) is a major enzootic disease of the dromedary camel. Thus, the purpose of the present study was to assess seroprevalence and infection rates in the Canary Islands using antibody(-card agglutination test-CATT/T. evansi) and parasite detection tests (micro-Haematocrit Centrifugation technique, Giemsa stained blood smears, microscopic examination of lymph node aspirates and mouse inoculation). PCV was also determined. 745 dromedary camels (483 females and 262 males) were examined. Trypanosomes were detected in seven animals. 36 animals yielded CATT positive results while 709 animals were negative. All parasitologically positive animals were also CATT positive. Results showed a good correlation between CATT positive and low PCV and a higher seroprevalence in older animals. Trypanocidal drugs have not been registered in Spain and, consequently, if vigilance is not exercised the prevalence could be increased in the future.     

Evaluation and improvement of parasitological tests for Trypanosoma evansi infection.

Research was undertaken to critically evaluate parasitological tests for the detection of Trypanosoma evansi in blood. The relative sensitivity of mouse inoculation (MI), the haematocrit centrifugation technique (HCT) and a modified miniature anion-exchange centrifugation technique (MAECT) were compared using blood and buffy coat. The effect that storage of blood prior to inoculation into mice has on the reliability of the MI test was also evaluated. The tests may be ranked in increasing order of sensitivity: HCT, MAECT with whole blood, MI with whole blood, MAECT with buffy coat and MI with buffy coat. The latter was able to detect 1.25 T. evansi per 4ml of blood. The reliability of the MI test was not reduced with storage of blood containing at least 25 T. evansi per ml for up to 21h prior to inoculation into mice. These results demonstrate that sensitivity of the MI and MAECT are increased approximately 10-fold through the use of buffy coat in place of whole blood. Although, the MI is marginally more sensitive MAECT is better suited to field use.     

Prevalence of Trypanosoma evansi infection and associated risk factors in camels in eastern Chad

A cross-sectional study was conducted to estimate the prevalence of Trypanosoma evansi infection (Surra) in herds of camels from the eastern area of Chad. The risk factors associated with disease were also identified. From August 1997 to April 1998, a random sample of 2831 camels from 136 herds was selected. Blood samples were collected and examined for the presence of T. evansi using an antibody (card agglutination test-CATT/T. evansi) and a parasite detection test (buffy-coat technique-BCT). Standardized questionnaires with information about the host and management practices were collected and evaluated for their association with seroprevalence (model 1) and parasitological prevalence (model 2) as indications of host sensitivity. In both models, risk factors were selected using ordinary logistic regression (OLR) and herd effect was evaluated using a generalized estimating equations (GEE) model. The apparent prevalence was 5.3% using BCT and 30.5% with CATT. Real prevalence was estimated at 16.9% +/- 1.4 (alpha = 5%). Overall, 27.9% (BCT) and 94.9% (CATT) of the herds had a least one-positive animal. Real herd prevalence was estimated at 42.6 +/- 8.3% (alpha = 5%). Camels of the large transhumants had the highest prevalence (estimated to 30.3% +/- 2.5; 62.9 +/- 12.0 in herds). Risk factors associated with seroprevalence were age, ethnic group, length of seasonal migration and longitude of pasture area in the dry season. Risk factors associated with BCT prevalence were age, length of seasonal migration, longitude of pasture area in the dry season, latitude of pasture area in the rainy season and season of sampling.     

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.     

Longitudinal study of an outbreak of Trypanosoma evansi infection in equids and dromedary camels in Israel

An outbreak of trypanosomoasis caused by Trypanosoma evansi involving horses, camels and donkeys occurred in a farm in Israel. A longitudinal study of two outbreak phases was conducted which included clinical monitoring, blood smears, packed cell volume (PCV), serology and polymerase chain reaction (PCR) followed by reverse dot blot (RDB) for the molecular detection of infection. This was the first reported T. evansi outbreak in domestic animals in Israel. Most of the camels on the farm (8/10; 80%) were diagnosed with T. evansi infection whereas infection was less prevalent in the horses (3/7; 43%) and donkeys (6/13; 46%). Clinical disease was evident in 4 camels and 1 horse exhibiting characteristic clinical signs, anemia and parasitemia detected on blood smears and by positive RDB. Six other animals were diagnosed as asymptomatic latent carriers by positive RDB and 6 additional animals were only seropositive and were considered suspected carriers. A significant difference was found in the mean PCV between symptomatic and latent carriers with severe anemia observed only in the symptomatic animals. An anaphylactic-like reaction, fatal in one case, was observed in 2 camels diagnosed with severe trypanosome parasitemia immediately following treatment with melarsenoxide cysteamine. Furthermore, recurrence of infection was documented in one camel 4 months post treatment.     

Characterization of Trypanosoma (Trypanozoon) evansi from dromedary camels in Kuwait by isoenzyme electrophoresis

Blood from 115 camels in Kuwait was examined for blood parasites. Two camels of a local herd (1.7%) were found to be infected with Trypanosoma (Trypanozoon) evansi and three camels (2.6%) with microfilarial nematodes. The Trypanosoma stocks isolated from these two camels were screened for isoenzyme patterns of 10 enzymes using thin-layer starch-gel electrophoresis. The results revealed that these two stocks were identical to camel stocks of T. evansi from certain countries in Africa, as well as to two stocks isolated from dogs in Kuwait. This is the first record of Trypanosoma (Trypanozoon) evansi isolates and microfilariae from camels in Kuwait.     

Evaluation of loop-mediated isothermal amplification (LAMP), PCR and parasitological tests for detection of Trypanosoma evansi in experimentally infected pigs

Six surra negative piglets (6-week-old) were infected with Trypanosoma evansi and two uninfected piglets were used as negative controls. Detection performances of various diagnostic tests (LAMP, PCR and parasitological tests) were compared by analysing blood samples collected weekly over a period of 11 weeks. With a two by two analysis without a gold standard, all methods were 100% specific. MI had the highest sensitivity of 65%, while LAMP, PCR, MHCT and TBS had sensitivities of 45, 33, 38 and 24%, respectively. However, when the analysis was done using MI as a gold standard, the sensitivity of MHCT was the highest at 53% followed by LAMP, PCR and TBS at 49, 44 and 35%, respectively. All methods gave high specificity above 60%. This study validates LAMP as an alternative method for the diagnosis of surra.     

伊氏锥虫和媾疫锥虫蛋白组分的电泳分析

应用SDS—PAGE和IEF电泳对伊氏锥虫和媾疫锥虫的蛋白组分进行了分析.在SDS—PAGE中,伊氏锥虫有21条带.媾疫锥虫有19条带,两种虫体的蛋白质区带在分子量40000～90000之间存在差异,尤其表现在表面糖蛋白上,伊氏锥虫为43000,媾疫锥虫为60000.在IEF电泳中,伊氏锥虫出现26条带,媾疫锥虫出现33条带,两种虫体的蛋白质区带在等电点4.8～5.6之间相同,而在5.6～7.0之间存在差异.本实验还对马媾疫锥虫的蛋白质进行了双相电泳分析,显示出86个多肽斑点.     

The ability ofStomoxys calcitrans and mechanical means to transmitTrypanosoma (brucei) evansi from goats to camels in Kenya

Stomoxys calcitrans failed to transmitTrypanosoma (b.) evansi from infected goats to other goats or camels, but the trypanosomal infection was transmitted by needle prick from infected goats to camels.Abbreviations HCT haematocrit concentration technique     

应用斑点酶联免疫吸附试验检测水牛伊氏锥虫抗体

应用斑点酶联免疫吸附试验(Dot—ELISA)检测水牛伊氏锥虫IgG抗体,并与间接血凝试验(IHA)进行了比较。检测160份伊氏锥虫疫区水牛血清160份,Dot—ELISA和IHA的阳性率分别为55.63％和53.75％;其中42份虫检阳性血清的阳性率分别为100％和92.86％,两种试验的一致率为95.63％。两种试验的滴度呈显著正相关(r=0.8842)。10份非疫区健康水牛血清两种试验均为阴性。