Morphological Characteristics And Transcriptomics Of Camellia Oleifera Infected With Exobasidium Gracile(Shirai)syd

Camellia oleifera Abel.is a unique oil species originating in China,which has important economic value,and its derivatives are widely used in chemical industry,light industry,food,feed and other industries.However,the disease of Camellia oleifera is becoming more and more serious with the change of the natural eco-environments,which greatly affects the utilization of Camellia oleifera.Hence,we made effort to understand the resistance mechanism of Camellia oleifera to Exobasidium gracile(Shirai)Syd.in order to provide a theoretical basis for better breeding of Camellia oleifera.In this study,both morphological structure and ultra-structure were observed of the infected leaves of Camellia oleifera with Exobasidium gracile(Shirai)Syd.by scanning electron microscopy and transmission electron microscopy.We also performed high-throughput transcriptome sequencing analysis and a set of genes involved in the pathogenesis of Camellia oleifera was identified,further providing a wealth of genetic resources for cloning of the disease-related genes and their functional validation.The main conclusions of this study were as follows:(1)Exobasidium gracile(Shirai)Syd.leads to abnormal development of young leaves of Camellia oleifera.It was found that the thickness of infected leaves was 3 to 5 times in comparision with normal leaves,the cell size was expanded 3 to 8 times,the cell number was increased 1 to 2 times,and palisade and sponge tissue was disappeared while accompanied by appearance of parenchyma cell with large-size.(2)The mycelium occurred in the gap of the 4 ~ 7th cell layer of the lower epidermis of Camellia oleifera leaves following the infection of Exobasidium gracile.As the mycelium grew rapidly,the leaf cells on both sides were gradually separated,and dehydration and abscission of the lower epidermis occurred in the final infection stage accompanied by the exposure of the hymenium with a thickness of 50 to 150 ?m.(3)The ultra-structure of leaf cells was observed by transmission electron microscopy(TEM).It was found that the cytoplasmic distribution was uneven and the organelles began to deform at the illness-health interface of the abnormal leaves.With the exacerbation of leaf swelling,the chloroplast was becoming less or even completely disappeared.We also observed several other abnormal phenomena such as the breakdown of chloroplast membranes,the decrease or even totally disappearance of chloroplast grana,and the breakdown of the other sub-cellular organelles including mitochondria.In addition,we only observed small debris and sac-like structure nearby cell-wall without intact sub-cellular organelles in some cells.(4)High-througput transcriptome sequencing of six samples generated 35.44 Gb clean data and 143014 unigenes were subsequently obtained from 315,771 transcripts.Of them,1160 unigenes were up-regulated while 1674 down-regulated.By the significant enrichment analysis of GO and KEGG,the cell components and the biological processes involved in DNA replication,cell cycle,DNA or chromosome conformation were significantly up-regulated,which was consistent with leaf cell proliferation caused by the pathogen infection.At the same time,up-regulation of genes involved in the synthesis of phenylpropanoid compounds and glutathione or in the metabolism of cytochrome P450,which was most likely tightly linked to the activation of defense responses of Camellia oleifera triggered by Exobasidium gracile.Taken all dada together,identification of the set of differential expression genes provides the fundamental basis to further explore the early biological marker for the disease-resistance-breeding of Camellia oleifera to Exobasidium gracile.