|SHIH, H.T. - Council Of Agriculture|
|SU, C.C. - Council Of Agriculture|
|CHANG, C.J. - University Of Georgia|
Submitted to: CDFA Pierce's Disease Control Program Research Symposium
Publication Type: Abstract Only
Publication Acceptance Date: 12/1/2019
Publication Date: N/A
Technical Abstract: Kolla paulula (Hemiptera: Cicadellidae) is a plant sap-feeding insect and vector transmitting Xylella fastidiosa subsp. fastidiosa causing Pierce disease (PD) of grapevine in Taiwan. The insect is found in Taiwan and other regions in Asia but not in the Americas. Little is known about the biology of K. paulula. One important research area encouraged by the recent next generation sequencing (NGS) technology is insect microbiota / microbiome. Manipulation of microbiota in insect vectors such as glassy-winged sharpshooter (GWSS, Homalodisca vitripennis) has been proposed as a potential strategy for PD control in California, USA. Members of the plant sap-feeding insect suborder Auchenorrhyncha (Hemiptera) have at least two obligate bacterial symbionts for essential amino acids (EAA) synthesis: a highly conserved symbiont “Candidatus Sulcia muelleri” and a second symbiont species varying among different leafhopper species. This research project explored the use of NGS technology to study endosymbionts of K. paulula. A colony of K. paulula was established and maintained on Commelina diffusa under laboratory conditions. Adult insects were collected and preserved in 70% alcohol solution. DNA was extracted from a single insect using a DNeasy blood and tissue kit (Qiagen, Valencia, CA). Whole-genomic DNA was amplified using an illustra GenomiPhi version 2 DNA amplification kit (GE Healthcare, Waukesha, WI) and sequenced using Illumina MiSeq format after preparation using the Illumina TruSeq DNA PCR-free library prep kit. A total of 28,327,432 reads (151 bp each) were generated. De novo assembling was performed with CLC Genomics Workbench software. The top 20 largest contigs were initially used for BLASTn search against GenBank nr database. Sequences of “Ca. Sulcia muelleri” were detected and collected to assemble the draft genome sequence (designated as Strain KPTW1). The KPTW1 strain has a genome size of 253,942 bp, GC content of 22.7%, 237 predicted protein-coding genes, and 34 RNA genes. Interestingly, no taxonomy identity could be assigned to the largest De novo contig (251,844 bp) according to GenBank nr database. Sequence annotation was performed and a 1,530 bp 16S rRNA gene sequence was identified. According the GenBank 16S rRNA sequence database, the DNA sequence was similar (99% query coverage and 80% identity) to that of Caedimonas varicaedens, an endosymbiont bacterium of the Ciliate Paramecium biaurelia. It is suggested that the Caedimonas-like bacterium could be the second symbiont of K. paulula.