Identification And Pathogen Classification Of Jujube Witches' Broom Disease In Dalian

Phytoplasmas are phytopathogenic prokaryotes which lacking cell walls and inhibit phloem sieve elements in infected plants. Phytoplasma have been associated with diseases in several hundred plant species, and it may cause the plant exhibiting yellows, witches broom, phyllody and others symptoms. In recent years, this disease has the tendency of causing more loss.According to the symptoms of phytoplasmas, we investigated diseases of phytoplasma and collected samples around the country. Suspicious samples of phytoplasma were detected and identified by nested-PCR assay, in which the universal primers for 16S rRNA of phytoplasma were used. The jujube witches'broom phytoplasma after identified were analyzed by sequencing and computer-simulated RFLP. It is the first time to identify and classify phytoplasma of JWB in Dalian, Liaoning Province. Main results and conclusion of this research are as follows:1.Suspicious samples of phytoplasma being collected from Liaoning, Henan, Hubei and other provinces were detected and identified by nested-PCR method, in which universal primers(R16mF2/R16mR1and R16F2/R16R2) of phytoplasma were used to amplifying 16S rRNA gene. As a result, only in DNA samples from Dalian of Liaoning Province, specific bands were amplified by PCR method, and which was identified as jujube witches'-broom phytoplasma disease subsequently.2.The incidence and disease index of JWB in Yingchengzi district in Dalian were investigated. Ultra structure of young tissues affected by phytoplasma were observed using ultra-thin section of electronic microscopy technique, the typical phytoplasmas bodies were observed in the phloem of diseased plant of jujube witches'broom.3.The PCR-amplified DNA segment of JWB-DL was sequenced. Results show that, the length of JWB-DL is 1851bp, G+C% is 44%, GenBank Accession No. is EF661582. Homologic analysis of nucleotide is 98% between JWB-DL and elm yellows phytoplasma (GenBank Accession No.AF189214). Therefore, the jujube trees should be infected by a type of phytoplasma which is belonging to the elm yellows group (16SrV). Analyzing sequence variations of 16S rDNA among JWB-DL and other JWB strains, partial rRNA operon in JWB-DL was different from others, indicating that JWB-DL is a distinct strain. The nucleotide sequence of the JWB-DL phytoplasma rplV-rpsC (GenBank Accession No. EF661581) is more identical to that of hemp fiber witches'broom phytoplasma (GenBank Accession No.EF029093) than those in JWB phytoplasma strains described previously. To our knowledge, this is the first report of a JWB disease in northeastern China, and JWB-DL represents a new, distinct'Candidatus Phytoplasma ziziphi'-related strain.