BAPC-assisted CRISPR/Cas9 System: Targeted delivery into adult ovaries for heritable germline gene editing (Arthropoda: Hemiptera)

Authors: Hunter, Wayne; Gonzalez, Maria; TOMICH, JOHN - KANSAS STATE UNIVERSITY

Submitted to: bioRxiv
Publication Type: Research Technical Update
Publication Acceptance Date: 12/8/2018
Publication Date: 12/10/2018
Citation: Hunter, W.B., Gonzalez, M.T., Tomich, J. 2018. BAPC-assisted CRISPR/Cas9 System: Targeted delivery into adult ovaries for heritable germline gene editing (Arthropoda: Hemiptera).bioRxiv. 451559.
DOI: https://doi.org/10.1101/478743

Interpretive Summary: First successful gene knockout in Asian citrus psyllid, that caused significant reduction in egg production, with shortened lifespan of adults. Innovative strategies are often limited in effectiveness in insects due to problems associated with successful delivery. In insects like psyllids microinjection into eggs fails due to causing 100 percent mortality after treatments. The Asian citrus psyllid, Diaphorina citri, is the vector for a pathogenic bacterium, Liberibacter asiaticus, Las, which is devastating the United States citrus industries. The disease called, Huanglongbing, HLB, (aka. Citrus greening disease) is transmitted during psyllid feeding. The economic losses are estimated in the billions of dollars, in United States, Brazil, and China. Reducing psyllid populations is one strategy being developed to prevent the spread of the disease. Breakthroughs that are very specific to the psyllid, will benefit other beneficial insects and organisms in environment. The method used a protein called 'BAPC'- to deliver suppression molecules into psyllid eggs while inside adult female psyllids. This method provides one avenue to suppress psyllid populations that would be safe, and not affect other organisms in the field. Development of these treatments will reduce insect vectors and stop pathogen transmission providing sustainable citrus and fruit crops in the United States.

Technical Abstract: Innovative gene targeting produced the first successful gene knockout in Asian citrus psyllid, Diaphorina citri, (Hemiptera: Liviidae), resulting in significant egg reduction, and shortened lifespan. This innovative method bypasses embryonic microinjection using a peptide-assisted delivery of gene targeting molecules. In hemipterans, embryonic injections fail due to 100 percent embryo mortality. The psyllid is the vector of the pathogenic bacterium, Candidatus Liberibacter asiaticus, CLas, which is devastating the United States citrus industries. The economic losses are estimated in the billions of dollars, in United States, Brazil, and China. Innovative technologies aimed at reducing psyllid populations using ribonucleic acid, RNA suppression, like RNA interference, or gene-editing, have potential to reduce insect vectors in a highly specific manner. This breakthrough improves insect suppression using a peptide call 'BAPC', to assist in the delivery of molecules into nymphs and adult psyllids by injection. These components when injected near ovaries produced heritable germline edits in subsequent generations. This method opens the potential of suppression strategies against insect vectors of disease. The treatment can reduce insect vectors, helping to stop pathogen transmission, and aiding efforts for sustainable citrus and fruit crop industries.