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Infectivity of cloned cacao swollen shoot associated badnaviral genome in Nicotiana benthamiana

Nomatter Chingandu: School of Plant Sciences, University of Arizona

<div>A severe rapid decline symptom phenotype was recently observed in cacao (<em>Theobroma cacao</em>) trees affected with cacao swollen shoot disease (CSSD) in West Africa. At least four badnavirus species, <em>Cacao swollen shoot virus</em> (CSSV), <em>Cacao swollen shoot Togo A virus</em>, <em>Cacao swollen shoot CD virus </em>(CSSCDV) and Cacao red vein virus (CRVV), have been associated with CSSD. To investigate whether the rapid decline phenotype is related to infection by a particular species, the full-length (monomer) genome, and partial dimer of CSSV-G75 isolate from Ghana were cloned and inoculated on 12 <em>Nicotiana benthamiana</em> seedlings using mechanical and agro-infiltration methods, respectively. Symptoms were observed on three of the 12 plants inoculated with the monomer clone, at 20-d post-inoculation (dpi), but not on mock-inoculated, or on the agro-infiltrated plants. Total DNA was isolated from the inoculated plants at 30-dpi and used as template for PCR amplification with CSSV-specific primers, resulting in amplicons of the expected size from monomer-inoculated and partial dimer-inoculated plants. DNA sequences indicated 88-100 % shared nucleotide identity with the cloned CSSV G75 genome. These results show that the cloned CSSV isolate G75 was biologically active and infectious in <em>N. benthaminana</em> plants by mechanical transmission and agro-infiltration. Experiments are underway to test infectivity for CSSCDV and CRVV clones using <em>T. cacao</em> seedlings to fulfill Koch’s postulates.</div>