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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.

In this study, swine fecal specimens (n = 251) collected from nursing and weaned piglets raised under smallholder production systems were screened for the presence of kobuviruses by RT-PCR. Porcine kobuviruses were detected in 13.1 % (33/251) of the samples. We demonstrated that porcine kobuvirus infections exist in indigenous pigs in Kenya and Uganda and that the prevalence was higher in young piglets than older pigs: nursing piglets (15 %), post-weaning (3-month-old) pigs (17 %), 4-month-old pigs (10 %). Genetic analysis of the partial RNA-dependent RNA polymerase (RdRp) region (690 nt) revealed that kobuviruses circulating in East Africa are diverse, sharing nucleotide sequence identities ranging from 89.7 to 99.1 % and 88 to 92.3 % among them and with known porcine kobuviruses, respectively. The nucleotide sequence identities between our kobuvirus strains and those of human, bovine and canine kobuviruses were 69.4-70.7 %, 73.1-74.4 % and 67-70.7 %, respectively. Additionally, upon sequencing selected samples that showed consistent 720-bp RT-PCR bands while using the same primer set, we detected porcine astroviruses in our samples belonging to type 2 and type 3 mamastroviruses. To our knowledge, this study reports the first detection and molecular analysis of both porcine kobuviruses and astroviruses in an African region. Further studies are required to determine the role of these viruses in gastrointestinal infections of pigs in this region and to determine the genetic diversity of the circulating strains to develop accurate diagnostic tools and implement appropriate control strategies.

Phylogenetic relationships among and genetic variability within 60 goats from two different indigenous breeds in Narok and Isiolo counties in Kenya and 22 published goat samples were analysed using mitochondrial control region sequences. The results showed that there were 54 polymorphic sites in a 481-bp sequence and 29 haplotypes were determined. The mean haplotype diversity and nucleotide diversity were 0.981 ± 0.006 and 0.019 ± 0.001, respectively. The phylogenetic analysis in combination with goat haplogroup reference sequences from GenBank showed that all goat sequences were clustered into two haplogroups (A and G), of which haplogroup A was the commonest in the two populations. A very high percentage (99.90%) of the genetic variation was distributed within the regions, and a smaller percentage (0.10%) distributed among regions as revealed by the analysis of molecular variance (amova). This amova results showed that the divergence between regions was not statistically significant. We concluded that the high levels of intrapopulation diversity in Isiolo and Narok goats and the weak phylogeographic structuring suggested that there existed strong gene flow among goat populations probably caused by extensive transportation of goats in history.

BackgroundStudies 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.FindingsTrypanosome 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.ConclusionThese 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.

The mountain bongo antelope Tragelaphus eurycerus isaaci has rapidly declined in recent decades, due to a combination of hunting, habitat degradation and disease. Endemic to Kenya, mountain bongo populations have shrunk to approximately 100 individuals now mainly confined to the Aberdares mountain ranges. Indirect observation of bongo signs (e.g. tracks, dung) can be misleading, thus methods to ensure reliable species identification, such as DNA-based techniques, are necessary to effectively study and monitor this species. We assessed bongo presence in four mountain habitats in Kenya (Mount Kenya National Park, Aberdare National Park, Eburu and Mau forests) and carried out a preliminary analysis of genetic variation by examining 466 bp of the first domain of the mtDNA control region using DNA extracted from faecal samples. Of the 201 dung samples collected in the field, 102 samples were molecularly identified as bongo, 97 as waterbuck, one as African buffalo and one as Aders' duiker. Overall species-identification accuracy by experienced trackers was 64%, with very high error of commission when identifying bongo sign (37%), and high error of omission for waterbuck sign (82%), suggesting that the two species' signs are easily confused. Despite high variation in the mtDNA control region in most antelope species, our results suggest low genetic variation in mountain bongo as only two haplotypes were detected in 102 samples analyzed. In contrast, the analysis of 63 waterbuck samples from the same sites revealed 21 haplotypes. Nevertheless, further examination using nuclear DNA markers (e.g. microsatellites) in a multi-locus approach is still required, especially because the use of mitochondrial DNA can result in population overestimation as distinct dung samples can potentially be originated from the same individual.

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