Ready-to-eat street food was obtained in the streets of the city of Ambato, Ecuador, and processed the same day. A sharp sterile blade was used to cut the samples on sterile surfaces. 10 g of each sample was placed in sterile brain heart infusion broth (BHIB) (Merck, Darmstadt, Germany) in 90 ml, shaken on a rotator for 8-10 min, and incubated for 24 h at 37 °C. A large amount of broth was inoculated on MacConkey agar plates (Merck, Darmstadt, Germany), Cromocult Coliforms Agar (Merck KGaA, Darmstadt, Germany), and CHROMagar mSuperCARBA were incubated overnight at 37 °C under aerobic conditions. Further purification was performed on Macconkey agar.

The isolates were amplified by polymerase chain reaction (PCR), analysed using agarose gel electrophoresis and visualised with Sybr Safe DNA Gel Stain. ³ For the identification of the isolated Enterobacterales, biochemical tests such as catalase, oxidase, TSI agar, Simmons citrate, lactose test, indole production, urea agar, methyl red test, and Voges-Proskauer were carried out and their interpretation was performed based on Bergey's manual. ⁴ Additionally, the software for Automated Biometric Identification Systems (ABIS) was used to confirm the biochemical identification results.

Agar disk diffusion assays (Thermo Scientific Oxoid and Bioanalyse) on Mueller-Hinton Agar (Thermo Scientific Oxoid) were performed. Antibiograms tests were based on the measured diameter of the zones of inhibition and interpreted as sensible, intermediate or resistant by referring to CLSI breakpoints. ⁵

The PCR test was performed according to the standardized protocol of the UTA RAM One Health research group ^{6 , 7} : 2.5 μL of DNA from each sample and 22.5 μL of PCR mix containing 12.5 μL DreamTaq PCR Master Mix (ThermoFisher Scientific, USA), 9 μL Nuclease-free water, 0.5 μL Primer 1 and 0.5 μL Primer 2 (final concentration of primers: 0.5 μM) were mixed to run PCR. The PCR conditions are reported in Supplementary Table S4. PCR products were analyzed by 1.2% agarose gel electrophoresis stained by Sybr Safe DNA Gel Stain (ThermoFisher Scientific, USA).

Hierarchical clustering

Hierarchical clustering was performed using the Euclidean correlation method and clustered by affinity. ² The MeV Multiexperiment Viewer software version 4.8.1 was used in this study.

The data presented show the frequency of isolation of Enterobacterales in 151 samples of ready-to-eat street food in Ambato, Ecuador ( Figure 1). The specific characteristics (date of sampling, type of street food, location) of the samples were reported in Supplementary Table S1. A total of 145 isolates were analyzed, and the results of the biochemical tests were reported in Supplementary Table S2. Among them, 86 isolates corresponded to E. coli and 59 isolates to other Enterobacterales.

To visualize the relative similarity of the antimicrobial resistance patterns of the isolates, a hierarchical cluster analysis was performed using the results of the antibiograms, where the phenotypes ‘resistant’, ‘intermediate’, and ‘susceptible’ were observed as red, white, and blue colors respectively. Dendrograms and clustered data were assembled using the complete linkage method through Pearson correlation and sample leaf organization. ⁷ For this purpose, the MeV Multiexperiment Viewer software version 4.8.1 was used. ⁸ Figures 2 and 3 represent the resistance profiles and the hierarchical clustering of E. coli and the rest of Enterobacterales, respectively. The complete information is shown in Supplementary Table S3.

The presence of diarrheagenic E. coli pathotypes present in ready-to-eat food was assessed in this study through the analysis of virulence genes related to the pathotypes. Only one isolate (C2.1c) was positive for the eae gene, suggesting the potential presence of enteropathogenic E. coli (EPEC) or enterohemorrhagic E. coli (EHEC). The β-lactamase resistance genes of Enterobacterales isolated in this study are reported in Table 1. Mobile colistin resistance genes or quinolone resistance genes were not found in the Enterobacterales isolates. The complete information about virulence genes and antibiotic resistance genes are available in Supplementary Table S5. The information about primers and PCR conditions were shown in Supplementary Table S4. The gel electrophoresis images are available at Supplementary figure S6. The disk difussion assays figures were shown at Supplementary figure S7.

This collection contains the following extended data:

Supplementary table S1. Characteristics (Sample type, date, treatment type, location, coordinates) of the ready-to-eat food samples. figshare. Dataset. https://doi.org/10.6084/m9.figshare.19579108.v1 ¹²

Supplementary table S2. Biochemical tests performed on Enterobacterales isolates from Ready-to-eat Street Food in Ambato, Ecuador. figshare. Dataset. https://doi.org/10.6084/m9.figshare.19579177.v1 ¹³

Supplementary table S3. Antibiogram of Enterobacterales isolated from ready-to-eat Street food of Ambato, Ecuador. figshare. Dataset. https://doi.org/10.6084/m9.figshare.19579189.v1 ¹⁴

Supplementary table S4. Primers used in this study and PCR conditions. figshare. Dataset. https://doi.org/10.6084/m9.figshare.19579198.v1 ¹⁵

Supplementary table S5. Antibiotic resistance genes and virulence genes harbored by Enterobacterales isolates from ready-to-eat street food in Ecuador. figshare. Dataset. https://doi.org/10.6084/m9.figshare.19579207.v1 ¹⁶

Supplementary figure S6. PCR results (positive electrophoresis images). figshare. Figure. https://doi.org/10.6084/m9.figshare.19729618.v1 ¹⁷

Supplementary figure S7. Disk diffusion assay images-Antibiotic resistance evaluation of Enterobacterales isolated from ready-to-eat street food of Ambato, Ecuador. figshare. Figure. https://doi.org/10.6084/m9.figshare.19729630.v1 ¹⁸

Data are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).