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The land application of animal manure can introduce manure microbiome and resistome to croplands where food crops are grown. The objective of this study was to characterize the microbiome and resistome on and in the leaves of lettuce grown in manured soil and identify the main transmission routes of microbes and antibiotic resistance genes (ARGs) from soil to the episphere and endosphere of lettuce. Shotgun metagenomic results show that manure application significantly altered the composition of the microbiome and resistome of surface soil. SourceTracker analyses indicate that manure and original soil were the main source of the microbiome and resistome of the surface soil and rhizosphere soil, respectively. Manure application altered the microbiome and resistome in the episphere of lettuce (ADONIS p < 0.05), and surface soil accounted for ∼81% of the microbes and ∼62% of the ARGs in episphere. Manure application had limited impacts on the microbiome and resistome in the endosphere (ADONIS p > 0.05). Our results show that manure-borne microbes and ARGs reached the episphere primarily through surface soil and some epiphytic microbes and ARGs further entered the endosphere. Our findings can inform the development of pre-and postharvest practices to minimize the transmission of manure-borne resistome from food crops to consumers.

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