Molecular Detection of Anaplasma phagocytophilum in Small Mammals and Infesting Ticks in Laikipia County, Kenya.

Anaplasmosis is a set of disease conditions of various mammals caused by bacteria species of the genus Anaplasma. These are sub-microscopic, Gram-negative, obligate intracellular pathogens that infect both vertebrate and invertebrate hosts. Significant species that infect domestic and wildlife animals include Anaplasma marginale, Anaplasma ovis, Anaplasma mesaeterum, Anaplasma platys, and Anaplasma phagocytophilum. Although A. phagocytophilum has a widespread distribution, there are only a few epidemiological reports from sub-Saharan Africa. This study focused on molecular detection and characterization of A. phagocytophilum in small mammals and their infesting ticks in Laikipia County, Kenya. A total of 385 blood and 84 tick archival samples from small mammals (155 females and 230 males) were analyzed. The blood samples were subjected to a nested PCR-HRM melt analysis using species-specific primers to amplify the 16S ribosomal RNA genes. The ticks were also subjected to nested PCR-HRM involving 16S rRNA gene primers. Anaplasma phagocytophilum DNA was detected in 19 out of 385 samples using species-specific 16S rRNA gene primers giving a prevalence of 4.9% for A. phagocytophilum. Analysis of the tick's samples using 16S rRNA gene species-specific primers also detected A. phagocytophilum in 3 samples from Haemaphysalis leachi ticks (3/84) equivalent to prevalence of 3.6%. Sequencing of 16S rRNA PCR products confirmed A. phagocytophilum in small mammals and ticks' samples. Phylogenetic analysis of the haplotype from this study demonstrated a close ancestral link with strains from Canis lupus familiaris, Alces alces, Apodemus agrarius, and ticks (Haemaphysalis longicornis) reported in Europe, China, and Africa. Comparison was also made with a known pathogenic A. phagocytophilum variant HA and a nonpathogenic variant 1 that were clustered into a distinctive clade different form haplotypes detected in this study. All the haplotype sequences for A. phagocytophilum from this study were submitted and registered in GenBank under the accession numbers OQ308965-OQ308976. Our study shows that small mammals and their associated ticks harbor A. phagocytophilum. The vector competence for H. leachi in A. phagocytophilum transmission should further be investigated.

The etiology and public health significance of mycobacteriosis of cattle in Kenya.

Background: Zoonotic tuberculosis, commonly referred to as bovine, tuberculosis is mainly caused by Mycobacterium bovis subsp. bovis, a member of the mycobacterium tuberculosis complex (MTBC). Infection in humans resembles that caused by Mycobacterium tuberculosis. Mycobacteria Other Than Tuberculous (MOTTs) are also increasingly being associated with tuberculosis humans but reports of infections in animals are scarce, their zoonotic potential being largely ignored. The study aimed to determine the aetiology and public health importance of mycobacteriosis in slaughter cattle in Kenya. Methods: Routine postmortem meat inspection was performed on a subpopulation of 1000 meat carcasses selected randomly from among 7,564 in a municipal abattoir, between January and July, 2015. Carcasses were examined for tuberculous lesions which were then examined for acid-fast bacilli, (AFB), cultured for isolation of mycobacteria and the isolates characterized by DNA analysis. Results: Of the carcasses examined, 218 (21.8%) had lesions in various parts of the carcasses. Acid-fast bacilli were observed in 63/218 of the lesions and suspected mycobacteria isolated from 35 of them. The isolates were identified as M. fortuitum (12), M. bovis subsp. bovis (3), M. shimoidei (2) M. asciaticum, M. interjectuctum, M. szulgai, M. celatum and M. kansaasii at one (1) each. Thirteen (13) of the isolates could not be speciated. Conclusion: The MOTTs identified in the study have been liked to various types of mycobacteriosis in humans. The study therefore highlights the pathogenic and zoonotic potential of MOTTs, indicating that zoonotic tuberculosis should not be restricted to Mycobacterium bovis subsp.bovis.

Prevalence of bovine tuberculosis in slaughter cattle in Kenya: a postmortem, microbiological and DNA molecular study.

A study to determine the presence and prevalence of bovine tuberculosis in slaughter cattle in Kenya was carried out in two abattoirs from July to November 2009. Routine postmortem meat inspection was performed on a subpopulation of 929 cattle selected randomly from among 4,984. Carcases were inspected for gross tuberculous lesions which were then examined for acid-fast bacilli, (AFB), cultured for isolation of mycobacteria and the isolates characterised by DNA molecular analysis. Of the carcases examined, 176 (18.95 %, 95 % CI) had lesions suggestive of tuberculosis. AFB were observed in 63/176 of the lesioned cattle and mycobacteria were isolated from 64 of them. The isolates were identified as Mycobacterium bovis (19/64), Mycobacterium tuberculosis, (2/64) and mycobacteria other than tuberculosis (43/64). The prevalence of M. bovis by molecular analysis was 2.05 % (95 % CI). This study documents for the first time the presence of bovine tuberculosis among slaughter cattle in Kenya. There is therefore a need to formulate and implement control programmes in order to minimise transmission among animals and to humans. Isolation of M. tuberculosis from cattle underscores the risk tuberculous humans pose to animals.

Assessing the effectiveness of intubation as a challenge model in contagious bovine pleuropneumonia vaccine experiments.

A study was carried out to assess the effectiveness of a bronchoscope in administering a pathogenic field strain of Mycoplasma mycoides subsp. mycoides (MmmSC) in cattle challenge experiments. Out of 16 animals inoculated using the bronchoscope, 10 (62.2%) showed clinical disease as evidenced by fever and 15 (93.8%) displayed typical lesions of CBPP from which MmmSC was isolated. Serum samples collected weekly were tested by Complement Fixation Test (CFT) and competitive enzyme-linked immunosorbent assay (c-ELISA). Antibodies to MmmSC were detected in 10 out of the 16 animals by the CFT and 11 out of the 16 animals by c-ELISA. The onset of clinical disease was as early as 2 days post-inoculation, and most of the animals developed clinical disease 2 to 3 weeks post-infection. These results clearly demonstrate that nasotracheal inoculation of pathogenic strain of MmmSC with the aid of a bronchoscope can lead to early onset of clinical disease; similar to previous studies but with higher numbers of animals showing clinical disease. This is in contrast with previous studies where early clinical disease was observed in as little as 15% of inoculated animals. This nasotracheal inoculation method using a bronchoscope can, therefore, be adopted for use in experimental challenge infections of cattle. This method is found to be a better replacement to the contact transmission method whose drawback includes extra cost of donor animals and unpredictable rate and timing of transmission from intubated to challenge animals.