DETECTION, IDENTIFICATION AND CHARACTERIZATION OF PHYTOPHTHORA spp. INFECTING CITRUS IN INDIA

摘要

Phytophthora species that cause root rot, foot rot, gummosis and decline of citrus in India were characterized by morphological and molecular means. P. nicotianae was found to be the predominant species followed by P. palmivora and P. citrophthora in the 28 nurseries and 174 orchards of five major citrus growing states of the country viz. Maharashtra, Madhya Pradesh, Punjab, Andhra Pradesh and Sikkim. Morphological characterization of 100 Phytophthora spp. isolates revealed 12 different types of colony pattern on V8 agar and 9 isolates of P. nicotianae were observed as A2 mating type and rest all the 91 isolates of Phytophthora spp. were of A1 mating type, while no self-fertile isolate was observed. The species P. nicotianae, P. palmivora, and P. citrophthora could all be clearly identified by PCR amplification of the nuclear ribosomal internal transcribed spacer (ITS) region followed by restriction analysis with MspI, AluI and RsaI. Sequencing and phylogenetic analyses of the ITS region, beta tubulin gene, translation Elongation factor 1 alpha and the region containing the mitochondrial cytochrome c oxidase subunit 2 gene fragment also confirmed the identity of these species. The phylogenetic groups did not correlate with geographic origin of the isolates. Additionally, isolates of P. nicotianae and P. palmivora were identified using species-specific PCR primers. The identification tools described in this study accurately detected the species of Phytophthora affecting citrus in nurseries and orchards in India.