Individual field-collected glassy-winged sharpshooter vectors harbor sequences from two Xylella fastidiosa subspecies

Authors: Stenger, Drake; Burbank, Lindsey; Krugner, Rodrigo; Sisterson, Mark

Submitted to: European Journal of Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/4/2019
Publication Date: 4/15/2019
Citation: Stenger, D.C., Burbank, L.P., Krugner, R., Sisterson, M.S. 2019. Individual field-collected glassy-winged sharpshooter vectors harbor sequences from two Xylella fastidiosa subspecies. European Journal of Plant Pathology. 155:329-388. https://doi.org/10.1007/s10658-019-01742-x.
DOI: https://doi.org/10.1007/s10658-019-01742-x

Interpretive Summary: Xylella fastidiosa is an invasive bacterial plant pathogen of significant economic impact worldwide. The bacterium exists as distinct subspecies that differ in ability to cause disease in certain hosts. Subspecies fastidiosa causes Pierce’s disease in grapevine; subspecies multiplex causes leaf scorch diseases in a variety of landscape trees and perennial crops but does not cause Pierce’s disease. Both subspecies occur in California and are transmitted to host plants by the glassy-winged sharpshooter. An epidemic of Pierce’s disease occurring in the Southern San Joaquin Valley of California was monitored during 2016-2017. Incidence of X. fastidiosa infection of glassy-winged sharpshooters collected from vineyards and adjacent citrus was determined by polymerase chain reaction (PCR) for three genes. Nucleotide sequence analysis of X. fastidiosa present in field-collected glassy-winged sharpshooters revealed that individual insects often harbor sequences of both subspecies, suggesting mixed infection in the insect vector. As two subspecies may occur in close proximity in co-infected glassy-winged sharpshooters, the results indicate that insect vectors are a plausible arena for genetic exchange among X. fastidiosa subspecies. Such genetic exchange increases the risk of emergence of new variants of X. fastidiosa with enhanced and/or altered potential to cause disease of increased severity and/or expansion of pathogen host range.

Technical Abstract: During 2016 and 2017, glassy-winged sharpshooters (GWSS) collected from vineyards or nearby citrus orchards in Kern County, California were assessed for presence of Xylella fastidiosa infection by quantitative PCR using total DNA samples extracted from GWSS heads. A subset of X. fastidiosa-positive samples was subjected to multi-locus sequence typing (MLST) to determine X. fastidiosa genotypes associated with GWSS in this region. Conventional PCR products for three genes (petC, leuA, and holC) were cloned from GWSS DNA samples and sequenced. Cloned sequences were assigned to X. fastidiosa subspecies based on Single Nucleotide Polymorphism (SNP) signature barcodes (7-12 polymorphic sites per amplicon that differentiate reference genomes of subspecies fastidiosa and multiplex). A total of 2005 MLST clones were sequenced: 1499 (74.8%) were genotyped as subspecies fastidiosa and 491 (24.5%) as subspecies multiplex. Of cloned subspecies fastidiosa sequences, 95.4% had SNP barcodes identical to the corresponding gene present in subspecies fastidiosa strains cultured from Pierce’s disease-affected vines in the same region sampled in 2016 and 2017. Presence of SNP barcodes representing both subspecies in one or more cloned genes was commonly observed within individual GWSS. This observation indicates that populations of X. fastidiosa in some insects consists of two subspecies or of two genotypes of one subspecies in which a proportion of the population was derived from a lineage with a history of horizontal gene transfer and homologous recombination. These conclusions suggest that 1) individual GWSS may visit multiple host species (inoculum sources); 2) sequential acquisition events can lead to co-infection of GWSS; 3) competitive exclusion of X. fastidiosa in the foregut is weak or not operating; and 4) the vector foregut represents a potential arena for exchange of genetic material among sympatric X. fastidiosa subspecies.