This chapter discusses the epidemiology, transmission, pathology, pathogenesis, diagnosis, control and economic impact of Trypanosoma evansi and T. vivax infections in domestic livestock.

The study investigated the effect of gamma-irradiation on bovine serum samples on the ability of enzyme-linked immunosorbent assay (ELISA) methods to detect trypanosomal antibodies. The serum samples were analysed using two standardised indirect ELISA systems. Higher measurement values were observed for most gamma-irradiated antibody positive and negative test samples. Using cut-off points, determined from the analysis of a non-irradiated trypanosomal antibody-negative population, the gamma-irradiated sera data showed that there was an increased risk of misclassifying samples as false positive or cross-reactive due to increased analytical sensitivity and decreased analytical specificity. The intraplate precision and agreement between tested and expected values of measurements were not altered throughout. The impact on the assays' diagnostic performance was estimated by analysing diagnostic sensitivity, diagnostic specificity and related parameters. The data demonstrated that although there was a bias of higher measurement values after gamma-irradiation, this could be compensated after readjustment of the cut-off points to obtain best separation of antibody-positive and -negative samples. Thus, for each assay, no significant difference of the diagnostic proficiency was found before and after gamma-irradiation. The practical implications are discussed of a serum sterilisation procedure using (60)Co gamma-rays for routine sample testing, assay validation and trypanosomosis monitoring and tsetse-fly control and eradication programmes.

Four indirect enzyme-linked immunosorbent assays (ELISAs) for the detection of antibody against trypanosomes using antigen-precoated plates (Trypanosoma congolense and T. vivax) were used in 15 veterinary diagnostic laboratories in Africa and Europe. The study provided data allowing an evaluation of charting methods with respect to the operational performance of each ELISA. Data from standardised internal quality control (IQC) samples were plotted on charts and used as the assay performance indicators with reference to expected upper and lower control limits. Based on unprocessed (optical density) and normalised absorbance values (calculated as a percentage positivity of a control), dispersion of values from the expected data range was estimated plotting the location and deviation of the values. In addition, assay precision was estimated plotting the distribution of coefficients of variation<10% of the IQCs. Binding ratios of controls were calculated to estimate the assay proficiency with respect to the accuracy of assessing that the IQC samples tested positive or negative in the test proper. The graphical analysis of dispersion of absorbance values in combination with assay precision and proficiency criteria was considered fully satisfactory to evaluate the operational performance of the ELISAs and provided useful decision criteria for plate acceptance and rejection. The establishment of standardised and transparent IQC data charting methods for the indirect ELISAs provided an increased measure of confidence to national laboratories with respect to their reports on disease occurrence. Moreover, the relative assay performances between all laboratories were examined using summary data charts with reference to the performance criteria described. The IQC data were also examined using modified Youden plot analysis demonstrating that indirect ELISA methods can be successfully applied at diagnostic laboratories in the tropics for monitoring trypanosomosis control programmes.

Research was undertaken to improve the antigen-coating step of indirect enzyme-linked immunosorbent assay (ELISA) method through the use of polystyrene 96-well plates precoated with antigenically stabile crude trypanosomal antigens. The plates were precoated with antigens, air dried and sealed before being packed in plastic bags with silica gel desiccant packets. Such plates stored at +4 and +37 degrees C provided an assay performance, which was superior to that of plates freshly coated with antigens from a frozen stock. Antigen-precoated plates consistently proved stable after storage up to +50 degrees C for at least 1 year. The accuracy of the assay was not affected, i.e. trypanosomal antibody-positive sera were clearly discriminated from trypanosomal antibody-negative negative sera. In contrast, lyophilized trypanosomal antigens lacked stability on storage at +37 degrees C for longer than 1 month. It was concluded that the routine use of antigen precoated polystyrene plates for the enzyme immunoassay technique will contribute to improved assay robustness at an acceptable diagnostic proficiency. The modified coating procedure will also provide an improved quality assurance and standardization procedure for the assay, which is required to allow the reliable detection of trypanosomal antibodies and comparison of data from different laboratories.

The incidence of trypanosomosis in sentinel cattle on Unguja Island, Zanzibar, was monitored every two to five months in 1994-97 to observe changes in disease transmission attributable logically to the application of insecticides, the release of sterilized tsetse flies (Glossina austeni Newstead) and the consequent decline and eradication of the wild tsetse population. Two parasitological techniques (microhematocrit centrifuge and buffy coat) were used to monitor the disease incidence caused by Trypanosoma congolense Broden and T. vivax Ziemann. T. congolense and T. vivax were detected in 1994 and 1995, but only T. vivax was detected thereafter. By 1997, the incidence of bovine trypanosomosis was only 0.1%. There was evidently no increase in disease incidence due to the release of sterilized isometamidium chloride-treated male tsetse flies.

The study reports the performance of four indirect enzyme-linked immunosorbent assays (ELISAs) for antibody (AB) detection using microtitre plates which were precoated with native or heat/detergent denatured antigens (AGs) from Trypanosoma congolense (T.c.) and T. vivax (T.v.), and stored for between 1 to 206 days at +37 degrees C. Bovine serum samples were obtained by sequential bleeding of 3-months old T.c.-infected bulls and their uninfected cohorts, as well as by a single bleeding of uninfected adult cattle. The first day of AB detection, and observations on samples after this (defined as estimated ELISA sensitivity), depended on the cut-off value in the specific ELISAs. Cut-off values from pre- and early post-infection samples of individual animals demonstrated a seroconversion in all ELISAs on average after 10-15 days post-infection (dpi). The AB detection was delayed in the T.c. native and denatured AG-based ELISAs using cut-off points from uninfected cohort cattle (16.5 dpi, 19.3 dpi) and the adult cattle population (22.1 dpi, 25.0 dpi). The T.v. AG-based ELISAs however lacked crossreactiviy to T.c. ABs. The estimated sensitivity of each T.c. AG-based ELISA was above 96% throughout, but significantly lower for the T.c. native AG-based ELISA (91.1%) when the adult cattle derived cut-off point was used (p<0.01). The sensitivity of the phase contrast buffy coat technique was similar to the T.c. AG-based ELISAs, but significantly lower when the T.c. denatured AG-based ELISA was used at the adult cattle derived cut-off point (p<0.05). The implications of the results and future research aspects on ELISAs to detect trypanosomal ABs and AGs are discussed.

L'incidence de la trypanosomose chez des bovins sentinelles sur l'île d'Unguja à Zanzibar a été contrôlée tous les deux à cinq mois entre 1994 et 1997 afin d'observer les changements dans la transmission de la maladie, logiquement attribuables à l'application d'insecticides, aux lâchers de mouches tsé-tsé stérilisées (Glossina austeni Newstead) et à la diminution consécutive, jusqu'à l'éradication, de la population naturelle de glossines. Deux techniques parasitologiques (centrifugation microhématocrite et examen du buffy coat) ont été utilisées pour surveiller l'incidence de la maladie due à Trypanosoma congolense Broden et à T. vivax Ziemann. T. congolense et T. vivax ont été détectés en 1994 et en 1995 mais, par la suite, seul T. vivax a été observé. En 1997, l'incidence de la trypanosomose bovine n'était plus que de 0,1 p. 100. Elle n'a manifestement pas augmenté après les lâchers de mouches tsé-tsé mâles stérilisées et traitées au chlorure d'isométamidium.

The diagnosis of trypanosomosis in animals with low parasitaemia is hampered by low diagnostic sensitivity of traditional detection methods. An immunodiagnostic method based on a direct sandwich enzyme-linked immunosorbent assay (ELISA), using monoclonal antibodies, has been examined in a number of African laboratories for its suitability for monitoring tsetse control and eradication programmes. Generally, the direct sandwich ELISAs for the detection of trypanosomal antigens in serum samples have proved to be unsatisfactory with respect to diagnostic sensitivity when compared with traditional parasitological methods such as the dark ground/phase contrast buffy-coat technique. Consequently, antigen-detection systems exploiting various other direct, indirect and sandwich ELISA systems and sets of reagents are being developed to improve diagnosis. In addition, an existing indirect ELISA for the detection of antibodies has been improved and is being evaluated in the field in order to detect cattle that are or have been recently infected with trypanosomes. Developments and advantages of other diagnostic techniques, such as dip-stick assay and tests based on the polymerase chain reaction are also considered.

A case-control study of calves under 3 months of age was carried out by weekly visits to 15 farms in the canton of Tilarán, Costa Rica. Most farms were dedicated to beef or dual-purpose (DP) production. Faecal samples were collected over a 6-month period from a total of 194 calves with clinical signs and from 186 animals without clinical signs of diarrhoea as assessed by a scoring system. The samples were investigated for the presence of viruses, bacteria and parasites. Torovirus was detected for the first time in Costa Rica and was present in 14% of calves with diarrhoea and in 6% of the controls. Coronavirus and Rotavirus were less frequently encountered in either one of the groups (in 9 and 7% of scouring calves and in 1 and 2% of controls, respectively). Escherichia coli was detected in 94% of all the faecal samples, but isolates from only three samples from calves with diarrhoea contained the K99 antigen. Similarly, Salmonella was found only in scouring calves. Cryptosporidium oocysts were detected in animals with signs of diarrhoea, while other coccidia oocysts, Strongylida and Strongyloides eggs were frequently found in animals both with and without diarrhoea. A conditional logistic regression (CLR) analysis to compare healthy and scouring calves showed a significant difference with regard to the presence of Torovirus, Rotavirus and Coronavirus.

Samples of bovine serum from uninfected and African trypanosomes-infected animals were tested before and after gamma-irradiation, using three sandwich enzyme-linked immunosorbent assays (ELISA). Each test system utilized a different monoclonal antibody, reputedly allowing the specific detection of conserved-invariant cytoplasmic antigens of trypanonosomes, T. congolense, T. vivax, and T. brucei, respectively. Results have identified two groups of samples. The first contained samples where there were unequivocal ELISA results indicating positivity and negativity, for non-irradiated samples. In this group, irradiation had no effect on the diagnostic sensitivity of the assays. All samples shown to be positive before irradiation remained positive and those shown to be negative, remained negative. There was, however, a statistically significant reduction in signal in each of the ELISAs following irradiation. The second group contained samples identified before irradiation as flanking the diagnostic negative/positive threshold of OD > or =0.05. These showed a negative bias after irradiation of the order of OD -0.01, which was shown to be statistically significant by paired t-statistics. Without correction of the given diagnostic negative/positive threshold, bovine sera with OD values around the threshold were expected to deliver more false negative test results upon irradiation. This was confirmed when serological data were compared with parasitological findings; where three times more false negative test results were found from irradiated serum samples. Consequently, for this group of irradiated bovine samples tested by ELISA, the re-adjustment of the diagnostic negative/positive threshold of the ELISAs using defined irradiated serum samples is recommended; otherwise, the frequency of false negative results might be increased.

Des facteurs et des moments critiques influant sur la qualité hygiénique du lait ont été examinés au Costa Rica dans une coopérative de petits éleveurs laitiers qui livraient tous leur lait à une laiterie. Le lait de toutes les vaches en lactation a été examiné avec le CMT (California Mastitis Test). Le lait positif au CMT a subi un examen bactériologique (EB). En outre, des échantillons de lait ont été prélevés dans les cuves de stockage pour le comptage des cellules somatique (CCS) et EB. Huit pour cent des échantillons positifs au CMT ont révélé la présence de S. aureus. Vingt-huit pourcent des échantillons prélevés dans les cuves de stockage contenaient S. aureus et 79 % des échantillons contenaient E. coli. Tous les échantillons pris dans les cuves de stockage de la laiterie avaient un nombre de CFU supérieurà 2.106 cellules/ml. Ce lait était fortement contaminé par des agents bactériologiques provenant de l'environnement. Le refroidissement du lait était inadéquat. La méthode de préparation des mamelles, les appareils de traite insuffisamment aseptisés et l'eau de nettoyage étaient les principales sources de contamination du lait.

Milk quality is determined by parameters of composition and hygiene. The compositional quality of milk is mainly influenced by nutritional, managerial and genetic factors. Furthermore, it is affected by proteolytic enzymes. These are either inherent to milk secretion, associated with leukocytes in mastitic milk, or synthesized by psychrotrophic bacteria that contaminate milk (10, 26). Proteolytic activity seems to be partly related to elevated somatic cell counts of raw milk (8, 23, 24). The hygienic quality of milk is influenced by pathogenic organisms, saprophytic microorganisms, residues, and other contaminants (18). Hygienic control measures are necessary in order to achieve a “clean, safe,

Five dairy farms, situated on the slopes of the volcano Poas in the pre-mountain cloud forest ecological zone, Alajuela and Heredia Provinces, Costa Pica, provided data for estimation of the production losses due to sub-clinical mastitis. Within the same farm, cows with proven sub-clinical mastitis were marched with cows without signs of sub-clinical mastitis, according to breed, lactation number and days in lactation. A total of 529 cows were detected with sub-clinical mastitis, of which only 200 could be paired with control cows free of mastitis; each pair was used only once.Crude milk production losses per cow with sub-clinical mastitis were estimated at 1.56 kg day-1 for daily milk yield. Milk production loss per affected quarter due to sub-clinical mastitis was estimated to be 17.6% on average. The decrease in milk production in heifers with sub-clinical mastitis did not differ significantly from the decrease in production in older cows. No significant difference in milk production loss was detected when the data were stratified on parity or the number of quarters affected.

The effects of trypanosome and helminth infections on health and production parameters in 2000 village N'Dama cattle were assessed periodically. Blood examination showed Trypanosoma congolense and Trypanosoma vivax to be prevalent, while strongylid-type eggs were those most frequently encountered in faecal samples. A distinct seasonal fluctuation was detected for both blood levels of trypanosomes and helminth egg output. Strongylid burden and trypanosome infection had significant negative effects on packed red cell volume levels and body weights mainly in animals of 2-3 years old. Clear indications of an increased susceptibility to trypanosomosis were found in animals affected by helminths. Similarly, animals infected with trypanosomes were more frequently infested with strongyles and egg counts were higher than in cattle in which no trypanosomes were detected.

A computerized herd health and production management program developed in the Netherlands has been applied in Costa Rica to assist farmers in optimizing farm management. During the past five years the farm community became acquainted with the computer program in pilot projects initiated by the School of Veterinary Medicine of the National University. Apart from its value in extension work, the program proved to be useful for teaching and research purposes and as a basis for decisions by policy makers.

The productivity of trypanotolerant N'Dama cattle, kept under traditional management conditions in The Gambia, West Africa, was assessed by the regular, monthly collection of health and production parameters in two study areas. The study areas were selected because of differences in tsetse challenge. Performance traits were used to build up an index to estimate the productivity of village N'Dama cattle. The productivity index per 100 kg cow maintained per year varied from 37.2 kg in the study area of Keneba village (with a low tstse challenge) to 21.4 kg for cattle kept near the villages of Tuba and Sambelkunda, an area which had a high tsetse challenge. Average age at first calving was 4.5 or 5.0 years depending on the study area, calving intervals were 623 or 703 days and there was an average 12% loss of body weight in adult females during the dry season. The productivity indices of village N'Dama cattle in The Gambia compare favourably with similar indices for trypanotolerant and trypanosusceptible breeds elsewhere in Africa, and show that even under harsh conditions and with high tsetse challenge, they are able to effectively produce milk and meat for the rural population.

Four experiments were carried out in which supplements of locally available proteinaceous by-products were given to young, mainly pre-weaned calves that ranged in initial weight from 30 to 90 kg. The experiments were part of a programme designed to establish practical and economical nutrition interventions for the improved productivity of N'Dama cattle under traditional village husbandry in a sub-humid agro-ecological environment in West Africa. The supplements included groundnut cake, sesame cake, and cotton seed, supplied during both the dry and wet seasons. While significant increases in growth rate were obtained to both dry and wet season supplementation, the responses were both inconsistent and much smaller than those obtained in other studies with older weaned N'Dama cattle. It is concluded that feed supplementation of young cattle should probably be restricted to animals whose liveweight exceeds 100 kg.