Molecular Identification of Phytophthora sp. From Indonesian Cocoa Using Phylogenetic Analysis

Abstract

Background and Objective: Diseases caused by Phytophthora species cause widespread damage worldwide and are troubling cocoa farmers in Indonesia. The specific species causing disease in an area can be ascertained by characterizing its rDNA fragments. This study aimed to identify Phytophthora sp., samples from cocoa plantations in Southeast Sulawesi, Indonesia, based on phylogenetic analysis of rDNA fragments. Materials and Methods: Identification of rDNA fragments of Phytophthora sp., done by amplifying rDNA fragments using PCR (Polymerase Chain Reactions) techniques with the specific primer of Phytophthora (Phy-F and Phy-R) which can amplify regions of ITS1, 5.8S rRNA and ITS2. The rDNA fragments are then sequenced and analyzed using: The BLAST (Basic Local Alignment Search Tools) provided by NCBI (National Center for Biotechnology Information) via (www.ncbi.nlm.nih.gov/blast) to analyze the local alignment of DNA sequences with Genbank DNA data and Mega 7.0.26 software is used to construct the phylogenetic tree. Results: The DNA sequencing results showed the rDNA measuring 786 bp consisted of complete sequences of ITS 1 (210 bp), 5.8S rRNA (162 bp) and ITS 2 (414 bp). Based on phylogenetic tree analysis using the maximum likelihood method with 1000 bootstrap replications showed that the rDNA of Phytophthora sp., isolates and 29 comparator isolates formed 2 large groups. Phytophthora sp., formed a subgroup with Phytophthora palmivora with a bootstrap value of 99%. Conclusion: The type of Phytophthora spreading in cocoa plantations in Southeast Sulawesi, Indonesia, is 1 group with Phytophthora palmivora.