ABSTRACT
BACKGROUND: Studies on the epidemiology of African Animal Trypanosomiasis (AAT) rarely consider the spatial dimension of disease prevalence. This problem is confounded by use of parasitological diagnostic methods of low sensitivity in field surveys. Here we report a study combining highly sensitive and species specific molecular diagnostic methods, and Geographical information system (GIS) for spatial analysis of trypanosome infection patterns, to better understand its epidemiology. Blood samples from 44 and 59 animals randomly selected from Teso and Suba districts respectively were screened for trypanosomes using PCR diagnostic assays. Spatial distribution of the positive cases was mapped and average nearest neighbour analysis used to determine the spatial pattern of trypanosome cases detected. FINDINGS: Trypanosome prevalence of 41% and 29% in Suba and Teso districts respectively was observed. T. vivax infections were most prevalent in both areas. Higher proportions of T. brucei infections (12%) were observed in Suba, a known sleeping sickness foci compared with 2% in Teso. Average nearest neighbour analysis showed the pattern of trypanosome infections as random. An overlay with tsetse maps showed cases lying outside the tsetse infested areas, mostly being cases of T. vivax which is known to be transmitted both biologically by tsetse and mechanically by biting flies. CONCLUSION: These findings suggest a need to design control strategies that target not just the biological vector tsetse, but also the parasite in cattle in order to clear the possibly mechanically transmitted T. vivax infections. There is need to also review the accuracy of available tsetse maps.
ABSTRACT
Currently, several PCR based diagnostic assays have been developed to improve the detection of pathogenic trypanosomes. These tests include use of species specific primers, single and nested PCRs' based on primers amplifying the Internal Transcribed Spacer (ITS) regions of ribosomal DNA. This study compares three PCR based diagnostic assays and assesses the agreement of these three asaays by screening 103 cattle blood samples randomly collected from trypanosome endemic areas in western Kenya. The nested ITS based PCR, the single ITS based PCR and the species specific based PCR detected 28.1%, 26.2% and 10.7% of the samples respectively as positive for trypanosome infection. Nested ITS and single ITS PCRs' picked 3.8% and 1.9% as mixed infections respectively. Cohen kappa statistic used to compare agreements beyond chance between the assays showed highest degree of agreement (0.6) between the two ITS based tests, and the lowest (0.2) between the nested PCR test and the species specific PCR. The single ITS and nested ITS based diagnostic assays detected higher numbers of positive cases, and reduced the number of PCR reactions per sample to one and two respectively, compared to the five PCR reactions carried out using the species specific primers. This significantly reduced the labour, time and the cost of carrying out PCR tests, indicating the superiority of the ITS multi-species detection techniques. Reliable epidemiological studies are a prerequisite to designing effective tsetse and trypanosomiasis control programs. The present study established the suitability of using ITS based PCR assays for large-scale epidemiological studies.
ABSTRACT
We established an in vitro hepatocyte primary culture system from Oreochromis niloticus, a tropical fish species of great economical importance, and evaluated its ability to express albumin, a liver-specific protein, consistently for a period of 3 wk. Serum requirements for fish hepatocyte cultures were assessed. A one-step in situ perfusion of tilapia liver retrogradely followed by collagenase liver dissociation and subsequent washing produced nearly 90% homogenous viable hepatocytes, as shown by trypan blue exclusion test. Mixed primary monolayer and aggregate hepatocyte cultures achieved by 10% fetal calf serum medium supplements expressed consistent levels of albumin. The results of light and electron microscopy showed that the hepatocytes did not significantly proliferate (P<0.05) but remained viable for at least 3 wk. The results of this study show that in vitro cultures of mixed primary hepatocyte monolayers and aggregates established from Nile tilapia may be useful models for studying transient cellular stress induction.
