Molecular Detection and Characterization of Pasteurella Multocida Infecting Camels in Marsabit and Turkana Counties, Kenya

Abstract

Pasteurella multocida is considered to be one of the normal flora in the respiratory tract of camels and other animals. It becomes pathogenic and causes Pasteurellosis when the resistance of the camel body is diminished by harmful environmental influences such as sudden changes in environmental conditions, poor dietary nutrition, walking for long distances and infestation by parasites such as trypanosomiasis. P. multocida is a small, gram-negative, nonmotile, non–spore-forming coccobacillus with bipolar staining features. The bacteria typically appear as single bacilli on Gram stain; however, pairs and short chains can also be seen. Close herding, overwork, limited food supply, and wet climatic conditions are stresses that seem to speed the spread of the infection. A study done in northern Kenya noted that in Africa P. multocida infections causing death in camels (Camelus dromedarius) have been existing since 1890 though the real cause of this disease remains elusive and needs further study. P. multocida capsular type B is the possible cause of respiratory disease in camels, its role as the causative agent for Pasteurellosis in camels’ remains unclear and needs further investigation. Polymerase chain reaction if done using proper primer design can enhance identification of P. multocida at different specificity viz; genus, strain, species, or all members of a domain. The study was done to detect, characterize and analyze P. multocida capsular types in Marsabit and Turkana Counties by application of PCR. The study was a retrospective study that used samples collected for diagnosis. P. multocida was detected, capsular typed and genetic diversity determined using PCR. The study was done on EDTA blood samples and nasal swabs brought to central veterinary laboratories (CVL)-Kabete collected from camels in Marsabit and Turkana Counties in the month of November 2018 and April 2019 respectively. The blood and nasal swabs preserved at -80°C were used for DNA extraction for subsequent molecular analysis. P. multocida KMT1 gene and capsular groups’ capA (hyaD-hyaC), capB (bcbD), capD (dcbF), capE (ecbJ) and capF (fcbD) were targeted marker genes. Capsular typing data was used to determine the genetic diversity of P. multocida. The genomic DNA was extracted from 132 samples (102 whole blood and 30 nasal swabs) for Molecular detection of P. multocida and capsular types by PCR assays. A total of 16% (21/132) samples for the two counties showed amplification product with KMT1SP6 and KMT1T7 specific primers. Marsabit County showed the highest P. multocida detection with 24.6% (15/61) and Turkana with the lowest detection of 8.5% (6/71). Based on the sample type the highest P. multocida detection was recorded in EDTA blood with 18.6% (19/102) and 6.7% (2/30) from nasal swabs. Capsular groups were determined using multiplex PCR. Among 21 DNA identified as P. multocida, capsular group E was detected by Multiplex PCR in twenty (21) P. multocida positive samples. Sequence analysis of the P. multocida positive samples matched capsular group E for gene ecbJ with accession number AF302466. The study concludes that PCR analysis based on KMT1T7 and KMT1SP6 primers work well for species confirmation of field strains irrespective of capsular types. The study recommends further characterization of P. multocida and capsular groups to provide definitive answers to the real cause of camel Pasteurellosis. Monitoring the capsular types of P. multocida strains circulating in a specific geographical region may be important in the formulation of vaccines. The P. multocida strain circulating in Marsabit and Turkana Counties is of capsular group E, there are not many capsular groups present in the two counties, and PCR assay targeting KMT1 gene is recommended for large-scale surveillance of P. multocida.