Good and bad lettuce leaf microbes? Unravelling the genetic architecture of the microbiome to inform plant breeding for enhanced food safety and reduced food waste

Lettuce is a high value food crop, consumed raw around the world. Engineering of the leaf microbiome could provide significant benefits for enhanced crop yield and stress resistance and help to reduce food waste caused by microbial spoilage. Lettuce leaves also act as a vector for human pathogens, implicated in several high-profile food-borne disease outbreaks. Since host genotype helps determine microbiome composition, we hypothesize that leaf surface traits can be defined that associate with ‘good’ bacterial microbiomes providing benefits to the crop and that ‘bad’ microbiomes, where spoilage organisms and human pathogens are abundant, can also be associated to underlying leaf genetics, providing key targets for future crop breeding. Using a Recombinant Inbred Line (RIL) population, we show that cultivated and wild parental genotypes differ with reduced bacterial diversity, larger leaves and fewer, larger stomata, smaller epidermal cells and more hydrophilic leaf surfaces found in the cultivated compared to wild lettuce. Functional analysis of the associated microbiome revealed increased abundance of genes associated with disease virulence for the cultivated lettuce genotype, suggesting domestication has had broad impacts on leaf and associated bacterial microbiome traits. We defined the core lettuce bacterial microbiome from 171 RILs, comprised of 45 taxa in the phyla Proteobacteria, Actinobacteria, Firmicutes, Chloroflexi and Deinococcus-Thermus. Leaf surface characteristics important in influencing bacterial diversity and abundance were identified as stomatal size (length and width), epidermal cell area and number and leaf surface hydrophobicity of the abaxial leaf surface. Quantitative trait loci (QTL) for leaf surface traits, frequently mapped alongside those for the extended phenotype of bacterial taxa abundance, including for human pathogensCampylobacterspp.,Escherichia-Shigellaspp.,Clostridiumspp. (LG 4, 5 and 6) and spoilage bacteria,Pseudomonasspp. (LG 1, 3, 4, 6 and 9). Candidate genes underlying these QTL were enriched in GO terms for cell wall assembly and modification, defence response, hormone-mediated signalling and biosynthesis and anatomical structure development. This work provides the first insight into the genetic architecture of host surface traits in a leafy crop alongside the mapped genetic architecture of bacterial communities and has identified areas of the lettuce genome as important targets for future microbiome engineering.