Factors that influence the prevalence of acaricide resistance and tick-borne diseases.

This manuscript provides a summary of the results presented at a symposium organized to accumulate information on factors that influence the prevalence of acaricide resistance and tick-borne diseases. This symposium was part of the 19th International Conference of the World Association for the Advancement of Veterinary Parasitology (WAAVP), held in New Orleans, LA, USA, during August 10-14, 2003. Populations of southern cattle ticks, Boophilus microplus, from Mexico have developed resistance to many classes of acaricide including chlorinated hydrocarbons (DDT), pyrethroids, organophosphates, and formamidines (amitraz). Target site mutations are the most common resistance mechanism observed, but there are examples of metabolic mechanisms. In many pyrethroid resistant strains, a single target site mutation on the Na(+) channel confers very high resistance (resistance ratios: >1000x) to both DDT and all pyrethroid acaricides. Acetylcholine esterase affinity for OPs is changed in resistant tick populations. A second mechanism of OP resistance is linked to cytochrome P450 monooxygenase activity. A PCR-based assay to detect a specific sodium channel gene mutation that is associated with resistance to permethrin has been developed. This assay can be performed on individual ticks at any life stage with results available in a few hours. A number of Mexican strains of B. microplus with varying profiles of pesticide resistance have been genotyped using this test. Additionally, a specific metabolic esterase with permethrin-hydrolyzing activity, CzEst9, has been purified and its gene coding region cloned. This esterase has been associated with high resistance to permethrin in one Mexican tick population. Work is continuing to clone specific acetylcholinesterase (AChE) and carboxylesterase genes that appear to be involved in resistance to organophosphates. Our ultimate goal is the design of a battery of DNA- or ELISA-based assays capable of rapidly genotyping individual ticks to obtain a comprehensive profile of their susceptibility to various pesticides. More outbreaks of clinical bovine babesisois and anaplasmosis have been associated with the presence of synthetic pyrethroid (SP) resistance when compared to OP and amidine resistance. This may be the result of differences in the temporal and geographic patterns of resistance development to the different acaricides. If acaricide resistance develops slowly, herd immunity may not be affected. The use of pesticides for the control of pests of cattle other than ticks can affect the incidence of tick resistance and tick-borne diseases. Simple analytical models of tick- and tsetse-borne diseases suggest that reducing the abundance of ticks, by treating cattle with pyrethroids for example, can have a variety of effects on tick-borne diseases. In the worst-case scenario, the models suggest that treating cattle might not only have no impact on trypanosomosis but could increase the incidence of tick-borne disease. In the best-case, treatment could reduce the incidence of both trypanosomosis and tick-borne diseases Surveys of beef and dairy properties in Queensland for which tick resistance to amitraz was known were intended to provide a clear understanding of the economic and management consequences resistance had on their properties. Farmers continued to use amitraz as the major acaricide for tick control after the diagnosis of resistance, although it was supplemented with moxidectin (dairy farms) or fluazuron, macrocyclic lactones or cypermethrin/chlorfenvinphos.

Characterisation of shops selling veterinary medicines in a tsetse-infested area of Kenya.

In a tsetse-infested area of Kenya, we characterised veterinary-drug outlets based on personnel and business characteristics to assess their capacities to provide clinical veterinary services. Structured questionnaires were administered to the retailers and sought information on the characteristics of the owners, salespersons and businesses. A total of 41 retail outlets (20 agro-veterinary, 11 pharmacy, and 10 general shops) were identified. There was poor response to questions on owner characteristics. Proprietors, who had no more than secondary education owned 15 out of 28 shops. Few shops (4/29) were owned by proprietors, who had professional qualifications (in animal health). Most salespersons had only secondary education but no qualifications. Animal-health assistants (AHAs), veterinarians and manufacturer's package inserts (drug leaflets) were the preferred information sources for the retailers. We concluded that drug retailers were poorly equipped with the technical knowledge necessary for drug dispensation and advice.

Trypanocidal drug resistance in eastern province of Zambia.

A survey to investigate resistance to drugs used in the treatment of bovine trypanosomosis was conducted in the eastern province of Zambia between 1996 and 1998. A cross-sectional study was conducted in three districts (Petauke, Katete, Lundazi) at 34 village sampling sites selected at random from villages that had shown greater than 6% prevalence of bovine trypanosomosis during an earlier survey. A longitudinal study was conducted in same three districts over a 1-year period. The study sites were chosen from the cross-sectional study and included eight sites showing high trypanosomosis prevalence and where no control activities were recorded. Use was made of parasitological methods, tests of resistance in cattle and mice and isometamidium-ELISA. Overall mean prevalence of trypanosomosis was 14.4, with 96% of infections caused by Trypanosoma congolense. The remainder was caused by Trypanosoma vivax (2%) and Trypanosoma brucei (2%). Tests in mice showed that of the stabilates collected, 24 (34%) were resistant to only isometamidium chloride, 8 (11.3%) were resistant to only diminazene aceturate, 1 (1.4%) was resistant to both drugs, and 38 (53.5%) were sensitive to both drugs. At least 2 out of 27 stabilates tested in cattle appeared to be resistant to trypanocidal drugs, 1 to isometamidium and 1 to diminazene. Isometamidium could be detected in only 63 (4.1%) of 1526 serum samples from cattle in the study. Only 6 (2.8%) of 212 serum samples from trypanosome-infected cattle had serum levels of the drug above 0.4 ng isometamidium per ml serum which is indicative for drug resistance in the infecting parasite population. Although some drug resistance is apparent, diminazene aceturate and isometamidium chloride can still be expected to be effective as a sanative pair in this area in most cases, since not more than 1 stabilate of 71 investigated showed evidence of resistance to both drugs.

Standardised tests in mice and cattle for the detection of drug resistance in tsetse-transmitted trypanosomes of African domestic cattle.

Resistance to the drugs used to control African animal trypanosomosis is increasingly recognised as a constraint to livestock production in sub-Saharan Africa. The most commonly used tests for detection of trypanocidal drug resistance are tests using mice or ruminants, but these suffer from lack of standardisation and hence it may be difficult to compare the results of different investigators. Tests in mice are less expensive than tests in ruminants, but while tests in mice they may be useful as a general guide to resistance in a geographic area they should not be extrapolated to cattle on an individual trypanosome level. Moreover, the commonly used protocols are too laborious for their application to large number of trypanosome isolates on an area-wide basis. This paper presents guidelines for standardised testing of trypanocidal drugs in vivo, and introduces a simplified single-dose test for use in mice, which is convenient for use in areas with limited laboratory facilities. The single-dose test is appropriate for characterisation of geographic areas in terms of trypanocidal drug resistance using large numbers of trypanosome isolates, for making comparisons between areas, and for monitoring changes in trypanocidal drug resistance over time. Multiple-dose tests may be used to determine the degree of resistance of individual stabilates to be determined precisely in mice are also described, but for logistical reasons these will rarely be conducted on more than a few stabilates, and testing of a larger number of stabilates in the single-dose test will generally provide more useful information. Finally, we describe tests in cattle that may be used to determine the efficacy of recommended curative doses of trypanocidal drugs for the treatment of infection with individual trypanosome isolates, including Trypanosoma vivax, which is rarely infective for mice.

Antibody-ELISA seroprevalence of bovine trypanosomosis in the Eastern Province of Zambia.

A longitudinal study was conducted over a 1-year period in six selected villages in Petauke and Katete districts in the Eastern Province of Zambia. Starting in November 1997, 50 animals were sampled at random at each village every 2 months. The parasitological prevalence of trypanosomosis was determined by the haematocrit centrifugation buffy-coat technique, supplemented with thick and thin Giemsa-stained blood films. Serum samples also were collected for anti-trypanosomal antibody determination by indirect enzyme-linked immunosorbent assay. Parasitological prevalence was highly variable between villages and between visits (range: 0-28.6%; median: 3.1%). Seroprevalence was also variable between villages (range: 0-80.8%; median: 50%), but was less variable between visits. Average annual parasitological prevalence and average annual seroprevalence for each village were highly correlated [R(2)(adjustedford.f.)=0.89, p<0.01]. Seroprevalence measured on any single visit to a study village was better than parasitological prevalence as a predictor of average annual parasitological prevalence.

Adaptation and validation of antibody-ELISA using dried blood spots on filter paper for epidemiological surveys of tsetse-transmitted trypanosomosis in cattle.

The indirect enzyme-linked immunosorbent assay (ELISA) for the detection of anti-trypanosomal antibodies in bovine serum was adapted for use with dried blood spots on filter paper. Absorbance (450 nm) results for samples were expressed as percent positivity, i.e. percentage of the median absorbance result of four replicates of the strong positive control serum. The antibody-ELISA was evaluated in Zambia for use in epidemiological surveys of the prevalence of tsetse-transmitted bovine trypanosomosis. Known negative samples (sera, n = 209; blood spots, n = 466) were obtained from cattle from closed herds in tsetse-free areas close to Lusaka. Known positive samples (sera, n = 367; blood spots, n = 278) were obtained from cattle in Zambia's Central, Lusaka and Eastern Provinces, diagnosed as being infected with Trypanosoma brucei, T. congolense, or T. vivax using the phase-contrast buffy-coat technique or Giemsa-stained thick and thin blood smears. For sera (at a cut-off value of 23.0% positivity) sensitivity and specificity were 86.1 and 95.2%, respectively. For bloodspots (at a cut-off value of 18.8% positivity) sensitivity and specificity were 96.8 and 95.7%, respectively. The implications of persistence of antibodies following treatment or self-cure are discussed.