Identification Of The Alfalfa Root Rot And Spot Disease Pathogen In Shaanxi Province

Alfalfa(Medicago sativa) belong to Rosales, legume, herbs of the genus, is one of the earliest and most widely cultivated legumes in the world, and is also one of the largest economic value legumes in China. However in recent years several typical diseases of alfalfa have been observed in Shaanxi Province, mainly phenomenon is that the leaves become yellow, withered plants, leaf distortion and other symptoms, and this reduces the alfalfa economic value and feeding value. In order to understand the pathogenesis and species of Alfalfa disease, in this study, the incidence of Alfalfa disease in different areas of Shaanxi province was investigated, counted the incidence of Alfalfa diseases, and finish the isolation and identification of pathogenic fungi in the laboratory. In order to provide theoretical basis for the prevention of Alfalfa diseases.The main results of this study are as follows:1. Between May and September in 2015, we survey the grassland of test sentinel in Dingbian County, Yuyang District, Suide County, Yangling District, Bin County, Dali County in Shaanxi Province, and counted the incidence of alfalfa disease. The results showed that brown leaf spot was the most common disease, in the whole growth season of alfalfa this disease incidence rate stay high, and the highest reached 62.14%; alfalfa leaf spot disease is relatively common, the incidence rate of around 10%, the highest reached 14.06%; and alfalfa rust occurred at different levels in 8-9 months. At the same time, some of the lower incidence Alfalfa diseases such as root rot, black stem disease, powdery mildew and so on were observed.2. The root rot pathogen of Medicago sativa separated from Shaanxi Province Dingbian County pasture were studied by morphological observation, EF-1α sequence analysis identification and inoculation test. The results indicated that: Through the morphological observation of the pathogen in the single colony in PDA culture medium and the observation of the mycelium and spores under microscope, this pathogen belong to Fusarium which caused the root rot of Medicago sativa; extracting the pathogen genome DNA by CTAB method, taking the translation elongation factor region for PCR amplification, gel recycling and purification, cloning, sequencing and build phylogenetic tree by Mega, analyzed the genetic relationship with known species in Genbank, the result showed that this pathogen have the closest relationship with Fusarium acuminatum, the reliability reached 99%; finally, through the root inoculation test, the symptoms of inoculated strain root are consistent with the root rot disease of Medicago sativa in field, verified that this pathogen we studied which caused root rot disease of Medicago sativa in Shaanxi Province is Fusarium acuminatum.3. The leaf spot disease pathogen of Medicago sativa separated from Shaanxi Province Bin County pasture were studied by morphological observation, OPA2-1 sequence analysis identification, extracting the pathogen genome DNA, PCR amplification, gel recycling and purification, cloning, sequencing and build phylogenetic tree by Mega, analyzed the genetic relationship with known species in Genbank. the result showed that this pathogen have the closest relationship with Alternaria alternata, and the reliability reached 100%; finally, through the leaf inoculation test, the symptoms of inoculated strain root are consistent with the leaf spot disease of Medicago sativa in field, verified that this pathogen we studied which caused leaf spot disease of Medicago sativa in Shaanxi Province is Alternaria alternata.4. By comparing the EF-1α and rDNA-ITS sequence analysis method in the identification of alfalfa root rot pathogen, the results showed that the rDNA-ITS sequence could not identify the level of Fusarium species, while EF-1α sequence analysis classify the Fusarium root rot of Alfalfa. By comparing the OPA2-1 and rDNA-ITS sequence analysis method in the identification of alfalfa leaf spot pathogen, the results showed that the rDNAITS sequence could not identify the level of Alternaria species, while OPA2-1 sequence analysis classify the Alternaria leaf spot of Alfalfa.