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

During 2016 and 2017, glassy-winged sharpshooters (GWSS; Homalodisca vitripennis ) collected from vineyards with a known history of Pierce’s disease or nearby citrus orchards in Kern County, California were assessed for presence of Xylella fastidiosa by quantitative PCR using total DNA samples extracted from GWSS heads. Conventional PCR products for three X. fastidiosa housekeeping genes ( petC , leuA , and holC ) were cloned from a subset of X. fastidiosa -positive GWSS DNA samples. 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 clones were sequenced: 1499 (74.8%) were genotyped as subspecies fastidiosa and 491 (24.5%) as subspecies multiplex . Among 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 pathogen acquisition events can lead to co-colonization of GWSS; 3) competitive exclusion of X. fastidiosa in the foregut is weak or not operating; and 4) the vector foregut is a potential arena for exchange of genetic material among sympatric X. fastidiosa subspecies.