De Novo Analysis And Functional Classification Of The Transcriptomes Of Heterodera Avenae And The Pathotype Characterization

The cereal cyst nematode (CCN) is worldwide and caused the yield losses range from10%to40%in winter wheat, in some particularly heavily infested fields could to80%, even have no harvests. Chinais the world‘s largest producer of wheat, with more than120Mt produced per year, with average yieldof about4t/ha. The distribution of H. avenae now includes16provinces and autonomous regions fromthe central, north, northwest and east of China. The wheat growing area of these16provinces representsmore than1/6of the total wheat in China and H. avenae is affecting4,000,000ha of the major wheatproducing regionsPathotype characterizations of two populations of Heterodera avenae China isolated from Beijingand Qinghai were identified using23standard International Test Assortment, and one local cultivars,Wenmai19was added as susceptible control. The results showed the two populations of H. avenae wereavirulent to Ortolan (Ha1). The barley Ortolan, Siri, Morocco, were resistant to the two populations.The six oats were all resistant to the two tested Heterodera avenae populations except the IGV.H72-646in the Beijing population. The wheat cultivars, except the cv. Iskamisch K-2-light in Beijing population,the others were all susceptible. But between the two tested nematode hosts, there‘s just only4cvsHerta、Harlan43、IGV.H72-646、Iskamish-K-2-light show different results with each other. Accordingto the characterization item, so we classified them as the new pathotypes named Ha91.Little is known about the molecular mechanism of H. avenae parasitism. In this study, weinvestigated the transcriptomes of H. avenae from Taian, at different developmental stages by HiSeq2000. Over49,46and47million high-quality reads were obtained from preparasitical second-stagejuveniles (J2s), fourth-stage juvenile (J4) females and adult females, respectively. Resulting in66962transcripts with average transcript length of1546bp,43953transcripts with average transcript lengthof1029bp and62697transcripts of average transcript length of1068bp with Trinity assemblers,respectively.45.42%,46.64%and43.10%of transcripts of preparasitical J2s, J4and adult females weresuccessfully annotated in Nt, KEGG, Swiss-Prot and Nr databases, respectively.According to the comparisition of RPKM, for the pair of preparasitical J2s and J4females,220of1101upregulated genes were clustered into73KEGG pathways, whereas499of1894downregulatedgenes were assigned to69KEGG pathways. For the pair of preparasitical J2s and adult females,728of2905upregulated genes were assigned to144KEGG pathways, whereas427of1689downregulatedgenes were clustered into57KEGG pathways. While only38of285upregulated genes and4of123downregulated genes were assigned to21and3KEGG pathways for the pair of J4females and adultfemales. When we compared the3libraries,836differentially expressed transcripts were found, with697transcripts were expressed in preparasitical J2s without gene expression in J4females and adultfemales,17genes in J4females without gene expression in preparasitical J2s and adult females,122genes in adult females without gene expression in preparasitical J2s and J4femalesGene Ontology (GO) analysis showed that5671,6244and6961differentially expressed genes were found in preparasitical J2s, J4females and adult females, respectively. Swiss-Prot analysis showedthat4181,5170and5571differentially expressed genes were found in preparasitical J2s, J4females andadult females, respectively. KEGG analysis indicated that6645unigenes were clustered into5014Biological process,929Cellular components and2012Molecular function pathways.3libraries KEGGanalysis respectively, indicated1824unigenes were assigned to239KEGG pathways in preparasiticalJ2s.2029unigenes were assigned to244KEGG pathways in J4females, whereas in adult females,2275unigenes were clustered into250KEGG pathways.The cDNA or protein sequences of30selected plants and animals were downloaded from NCBIdatabase, respectively. cDNA sequences of free-living nematode C. elegans was downloaded fromWormbase. All the unigenes from the three H. avenae transcriptomes of preparasitical J2s, J4femalesand adult females were used to search for homologous sequences in these30plants and animals and C.elegans by tBLASTX or BLASTX. In addition,471putative effectors that were similar to knownphytonematode effectors and117putative novel effectors that had no match sequence to knownphytonematode effectors were identified in H. avenae transcriptomes. Further227transcripts withoutorthologous sequence to30selected plants and animals were found to have putative lethal RNAiphenotypes. These data lay the foundation for future study of H. avenae-cereal interactions and H.avenae parasite control.