Analysis of the Sequences of 16S rDNA and secY of Cajanus scarabaeoides Witches’-broom Phytoplasma in Yuanmou, Yunnan Province

Purpose To determine the pathogen of Cajanus scarabaeoides witches’-broom disease samples which were collected in Yuanmou of Yunnan Province.

Method Near-full-length phytoplasmal 16S rDNA gene sequences and secY gene sequences were amplified via direct-PCR and nested-PCR using phytoplasma universal primer pairs from DNAs extracted from C. scarabaeoides symptomatic plants. The amplicons were cloned and sequenced. DNA fragments about 1.8 kb of 16S rDNA and 1.7 kb of secY were obtained.

Result Classification of 16S rDNA by using computer-simulated restriction fragment length polymorphism (RFLP) and phylogenetic analysis in iPhyClassifier indicated that the pathogen of C. scarabaeoides witches’-broom is a member of phytoplasma subgroup A, group 16SrII (similarity coefficient 1.00), which related to Candidatus Phytoplasma australasiae. From the homology comparison and the phylogenetic tree based on secY gene sequences showed that genetic relationship of C. scarabaeoides witches’-broom phytoplasma strain (CaSWB-YNym) was closely related with PnWB from Yunnan and Taiwan Province and confirmed that CaSWB-YNym belongs to the member of subgroup A, group 16SrII. The classification result based on secY gene analysis coincided with those of results based on 16SrDNA gene.

Conclusion To our best knowledge, it is the first report of a 16SrII group phytoplasma associated with C. scarabaeoides witches’-broom disease in Yunnan Province. And it is the first study to determine the phytoplasma pathogen of C. scarabaeoides witches’-broom disease in Yunnan Province and identify taxonomic status using molecular biological techniques. These results will provide a theoretical basis for early diagnosis and detection of C. scarabaeoides witches’-broom disease.