Vegetable phylloplane microbiomes harbour class 1 integrons in novel bacterial hosts and drive the spread of chlorite resistance

Bacterial hosts in vegetable phylloplanes carry mobile genetic elements such as plasmids and transposons that are associated with integrons. These mobile genetic elements and their cargo genes can enter human microbiomes via consumption of fresh agricultural produce, including uncooked vegetables. This presents a risk of acquiring antimicrobial resistance genes from uncooked vegetables. To better understand horizontal gene transfer of class 1 integrons in these compartments, we applied epicPCR, a single-cell fusion-PCR surveillance technique, to link the class 1 integron integrase (intI1) gene with phylogenetic markers of their bacterial hosts. Ready-to-eat salads carried class 1 integrons from the phyla Bacteroidota and Pseudomonadota, including four novel genera that were previously not known to be associated with intI1. We whole-genome sequenced Pseudomonas and Erwinia hosts of pre-clinical class 1 integrons that are embedded in Tn402-like transposons. The proximal gene cassette i..., Emulsion, paired-isolation and concatenation PCR (epicPCR)
Approximately 150,000 bacterial cells were resuspended in 75 µL of PCR reagents containing GC buffer (1x), dNTP mix (0.4 mM), Phusion High-Fidelity DNA polymerase (0.05 U/µL) (Thermo Scientific, United States), bovine serum albumin (1 µg/µL) (Promega, United States), Lucigen Ready-Lyse lysozyme (500 U/µL) (LGC Biosearch Technologies, United States), and three primers: R926 (2 µM), intI1_outer_2 (1 µM), and R519-HS458RC bridge primer (0.04 µM) with the final concentrations of each reagent in parentheses. For each of the six phylloplane microbiome sample, three technical replicates of epicPCR were performed, one of which was spiked with a class 1 integron-free Synechococcus species CC9311 strain at a population frequency of 10% to test if false associations can be observed between the 16S rRNA markers of Synechococcus CC9311 and class 1 integrons in each sample.
Each PCR suspension was added to 425 µL of ABIL oil and was agitated ..., , # Vegetable phylloplane microbiomes harbour class 1 integrons in novel bacterial hosts and a chlorite-resistance-conferring gene cassette

[https://doi.org/10.5061/dryad.f1vhhmh4k](https://doi.org/10.5061/dryad.f1vhhmh4k)

The eighteen FASTQ files that are available for download are as follows:

S1_1, S1_2, S1_3, S2_1, S2_2, S2_3, S3_1, S3_2, S3_3, ML1_1, ML1_2, ML1_3, ML2_1, ML2_2, ML2_3, ML3_1, ML3_2, ML3_3

## Abbreviations

S = spinach leaf salad; ML = mixed leaf salad

The first digit after the letters denotes biological replicates. Each biological replicate represents a packaged salad product purchased from the supermarket on a different day and was processed for microbiome extraction independently. The second digit after the underscore denotes the technical replicates.

## Code/Software

The epicPCR Nanopore reads in the FASTQ files were filtered to ensure that they were flanked by the nested forward and reverse primers in the correct orientation using pychopper (v2.7.9) ([https:...