Identification Of Several Kinds Of Important Pathogenic Nematodes In Citrus

China is one of the key citrus producing country in the world.Plant-parasitic nematodes are the important pathogens of citrus spp..There were reports that the root knot nematode,Meloidogyne spp.caused serious damage to citrus in some citrus-producing orchards in Guangdong Province,China,but species of the root knot nematodes were uncertain.Surveys have been done for citrus root knot nematode during the year from 2014 to 2015 in Guangdong.The root knot nematode diseases of citrus were found in Zhaoqing region in Guangdong Province,and species of the Meloidogyne were identified.Phylogenetic relationships among citrus root knot nematode populations collected from Fujian,Guangdong,Guangxi Province were analyzed based on phylogenetic tree.The four newly recorded nematodes species parasitizing on citrus in China or Fujian were identified based on morphological and molecular characteristics.The main results obtained are as follows:1.Citrus root-knot nematode were collected form the roots of Citrus reticulate Blanco.cv.Shatangju(Poncirus trifoliate root stock)in the Heerkou town and Jinzhuang town,Zhaoqing region in Guangdong province.Fibrous roots were infected by the nematode,typical symptoms of root infected by the nematode was curly roots with galls.The species of Meloidogyne were identified as Meloidogyne mingnanica by the morphological characteristics of female’s perineal pattern;male;second-stage juveniles.The 18S;rDNA-ITS;28S D2-D3 region of Populations of Meloidogyne mingnanica from Guangdong province,Meloidogyne mingnanica from Guangxi province and Meloidogyne sp.collect citrus root form Shunchang county,Fujian province were sequenced.Phylogenetic trees were constructed inferred from the sequences of 18S,rDNA-ITS or 28S D2-D3 region with the corresponding regions of other Meloidogne species available in GenBank based on Maximum Likelihood(ML)and Bayesian analysis.28S D2-D3,18S based ML or Bayesian analysis revealed higher resolution within the group containing sequences of populations of citrus root knot nematode collected from Guangdong and Guangxi Province,within another group containing sequences of the populations collected from Shunchang country,Fujian Province.The results have clarified taxonomic status of species of root knot nematode parasitizing on citrus in Guangdong,Guangxi,Fujian Province.2.The species-specific primers were designed for the first time to diagnosis M.mingnanica based on the sequences of 28S D2-D3 expansion region.The primers were sensitive to amplified the expected band size(399bp)from DNA templates of a single second-stage juvenile(J2)and no band were amplified from populations of M.javanica,M.enterolobii,M.incognita,M.graminicola,M.arenaria and Meloidogyne sp.collected from citrus root from Shunchang country.Specificity and reliability of the primers were validated.3.Based on the morphological and molecular characters(the sequence of the D2-D3 expansion region of 28S and rDNA-ITS region).Paratylenchus lepidus collected form roots of citrus in Fujian province was identified,and it is a new record in Mainland China.Pratylenchus vulnus and Hoplolaimus seinhorsti were identified from roots of Poncirus trifoliate.P.vulnus is a newly recorded species in Fujian province and H.seinhorsti is first report on P.trifoliate.