DIFFERENTIAL DIAGNOSIS OF AFRICAN SWINE FEVER IN SUSPECTED OUTBREAK SAMPLES IN DOMESTIC PIGS IN WESTERN KENYA USING MOLECULAR TOOLS Obange F.A 1, Okoth E. 2, Obiero G.O. 1 Centre for Biotechnology and Bioinformatics (CEBIB)- University of Nairobi 1 International Livestock Research Institute (ILRI) 2 Expected Results Methods Introduction CENTRE FOR BIOTECHNOLOGY AND BIOINFORMATICS (CEBIB), UNIVERISTY OF NAIROBI African Swine Fever (ASF) is a contagious, hemorrhagic viral disease of pigs caused by the African Swine Fever Virus (ASFV) that is transmitted by the tick vectors Ornithodoros moubata and Ornithodoros sonrai. It has a mortality rate of up to 100 percent leading to decreased productivity and great economic losses for pig farmers in several parts of the world; most notably in Sub-Saharan Africa where it persists. While ASF is lethal to domestic pigs, it presents with similar symptoms to those observed in other haemorragic pig diseases of lower morbidity and mortality including Classical Swine Fever (CSF), Erysipelas, Salmonellosis, Thrombocytopaenia Purpura, Porcine Circovirus, Porcine Reproductive and Respiratory Syndrome and Warfarin poisoning. Accurate diagnosis of a pig suspected to have ASF is important in order to avoid misdiagnosis and consequent erroneous ‘stamping out’ of pig populations suspected to have ASF which could lead to great economic losses for pig farmers. This study sets forth with the objective of using molecular tools to determine the presence, prevalence and molecular characteristics of the microbial differential diagnoses of ASF in domestic pig samples collected during suspected ASF outbreaks in western Kenya. This information will be used to inform surveillance systems on diseases to be included in outbreak investigations following reports of outbreaks of ASF- like diseases. This will further inform development of control strategies for these ASF differential diagnoses in Kenya. 1.Confirmation of the presence of ASF or the differential diagnoses of ASF in domestic pigs in western Kenya 2.An assessment of the number of coinfections present with these diseases African Swine Fever infected pig Source: Objectives Hypothesis There are no diseases of similar clinical presentations to ASF in suspected ASF-like cases in Western Kenya 1.To determine microbial presence, diversity and relative species abundance in pig samples collected during suspected African Swine Fever outbreaks 2.To genetically characterize detected microbial communities to establish associations with similar microbes 3.To analyse the temporal patterns of the detected microbial communities over time Ornithodoros moubata Source: Data analysis Similarity analysis using MG-RAST® pipeline Phylogenetic trees construction using MEGA® Sorting using R Statistical Software to generate disease temporal patterns Sequencing MiSeq (Illumina®) platform using sequence runs of 2×150 paired-end reads De novo assembly of contigs using the Meta-IDBA® assembler Library Construction RNA library construction using TruSeq® RNA Sample Preparation kit v2 DNA library construction using and KAPA® Hyper Prep Kit DNA/RNA purity and integrity measurement Spectrophotometry Gel electrophoresis DNA/RNA extraction and purification Serum aliquoting DNase buffer added to each aliqout Host DNA degradation with DNase Viral DNA/RNA extraction Sampling Grouping of samples according to outbreaks Random sampling to generate sample ID numbers for analysis using R statistical software African Swine Fever Virus Source: