Identification Of Hirschmanniella Spp. (Nematoda: Pratylenchidae) Extracted From Rice Root In Guangdong Province And A Loop-mediated Isothermal Amplification Assay For Detection Of Hirschmanniella Oryzae

The rice root nematodes,Hirschmanniella spp.are widely distributed in the rhizosphere of rice roots,and can affect the growth of rice.Different species of Hirschmanniella spp.can co-parasitize the same rice fields,even the same plants.Hirschmanniella species are also widespread distributed in Guangdong,however systemic species identification have not been reported yet.In this study,using morphological identification and a single nematode PCR identification,species of the genus Hirschmanniella were identified from the rice root nematode samples collected from Ningxi base of South C hina Agriculture University in Zengcheng city,the farm of South China Agricultural University,Q ingyuan,Yangjiang,Jiangmen and Zhaoqing in Guangdong province.In addition,the p hylogenetic trees of Hirschmanniella species were established based on bioinformatic analysis,a rapid detection to identify H.oryzae by Loop-mediated isothermal amplification?LAMP?were also carried out.The main results are shown as follows:1.Nine species of rice root Hirschmanniella species were identidied and described in this study.They are Hirschmanniella areolata,H.behningi,H.anchoryzae,H.caudacrea,H.diversa,H.orycrena,H.oryzae,H.microtyla and H.mucronata.Among them,H.areolata and H.behningi are new record species to China.H.anchoryzae,H.diversa,H.anchoryzae,H.areolata,H.orycrena and H.behningi are new record species to Guangdong.H.oryzae,H.microtyla and H.mucronata are recorded species,.2.Phylogenetic trees of H.oryzae and H.mucronata were constructed based on the ITS and D2-D3 sequences.3.The conexisted nematode species from the 6 different places were identified.The largest number of species from the same field was 7,and that of species from the same rice plant was usually 2 to 3,among all the nematode species,H.oryzae is dominant species.4.The specific primers for LAMP were designed according to the ITS ribosomal DNA?rDNA?sequences of H.oryzae by comparing the related sequences of Hirschmanniella spp.deposited in GenBank.The LAMP products were successfully detected by the agarose gel electrophoresis and SYBR Green I dying,and then the LAMP amplification with highest efficiency were obtained by the optimized conditions of 57?,1.4 mmol/L dN TPs,7 mmol/L MgSO₄,and 45 minutes of extension.In this study,different species and coexisted phenomena of these different species of Hirschmanniella spp.extracted from rice roots in Guangdong province were identified in addition,LAMP rapid identification method of H.oryzae were also established.These results are significant for mastering of the occurrence of Hirschmanniella spp.in Guangdong province,and formulating reasonable prevention and control measures.