Enteroaggregative Escherichia coli foodborne outbreak in Shandong Province, China (2023): comprehensive epidemiology and genomic resistance profiling

Abstract

Introduction: Enteroaggregative Escherichia coli (EAEC) is an emerging and intricate diarrheagenic bacterial pathogen responsible for acute and persistent diarrhea in children, adults, and travelers. Despite its clinical significance, the global understanding of EAEC outbreaks and pathogen characteristics remains limited due to its complex epidemiological profile.

Methods: This study presented a retrospective analysis of a foodborne diarrheal outbreak caused by EAEC in a county school in Shandong Province, China in 2023 by using epidemiological investigation and comprehensive genomic analysis techniques.

Results: A total of 15 EAEC isolates were identified, including 13 from stool samples, one from sauced beef, and one from flies. Epidemiological and phylogenetic analyses pinpointed the EAEC isolates from sauced beef as the primary causative agent of the outbreak. Genomic comparisons revealed significant genetic consistency across nine outbreak-associated strains, particularly in virulence gene profiles, antimicrobial resistance gene profiles, molecular typing, and genetic evolution.

Discussion: These findings underscored the utility of whole-genome sequencing in elucidating the genetic diversity of EAEC within specific environments and tracing its origins, thereby supporting targeted interventions such as infection control and prevention strategies. Notably, the study also identified an extensively drug-resistant (XDR) EAEC strain from flies in the canteen, harboring both the mcr-1 and bla_CTX-M-132 resistance genes. This unexpected discovery highlighted the critical importance of applying the "One Health" approach, emphasizing the need for continuous surveillance of antibiotic-resistant bacteria in animals and environmental sources to mitigate potential public health risks.

Keywords: Enteroaggregative Escherichia coli (EAEC); antimicrobial resistance; foodborne outbreaks; whole-genome sequencing; “One Health” approach.

Figure 1

Investigation of outbreak epidemics. (A) Epidemic curve for outbreak cases and timeline of key events in the investigation. (B) PFGE patterns of the EAEC strains isolated from individuals involved in the outbreak.

Figure 2

The phylogenetic tree of the 15 EAEC group isolates this study and another 199 EAEC group isolates in the NCBI database.

Figure 3

Pathogenicity of outbreak isolates. (A) BLAST atlas produced using BLAST ring image generator. Comparative BLASTn analysis with 70, 90, and 100% identities are displayed, and gaps in circles represent regions with no identity of genes. The innermost ring represents the EAEC BZ21 used as the reference and its coordinates. The second ring (in black) plots the GC content of the reference, followed by its GC skew (in purple/green). The following rings represent isolated EAEC (in order from inside out: BZ01, BZ03, BZ05, BZ12, BZ15, BZ16, BZ32, BZ18, BZ24, BZ26, BZ27, BZ28, BZ31, BZ37, and BZ21). Outer ring annotation: black, virulence genes; red, plasmid replicons. (B) Adherence phenotype to Hep-2 cells of different EAEC strains. B1 BZ21 (AA), B2 BZ12 (AA), B3 BZ24 (AA), B4 BZ15 (LA).

Figure 4

Drug resistance in outbreak isolates. (A) Phenotype of drug resistance. MICs of 15 antimicrobials belonging to 11 antimicrobial classes were tested. (B) Research on drug resistance genes. The resistance (red block) and susceptibility (blue block) of isolates to the antibiotics were indicated using different colors.

Figure 5

Genetic analysis of plasmid sequences of PLBZ01-2 (this study, BZ01) and pM-132-4972 (MT773671.1). 99% homology are marked by gray shading. Arrows represent direction of transcription. Red open reading frames (ORFs) indicate mcr-1/CTX-M-132, blue ORFs indicate insertion elements, orange ORFs indicate mobile elements, and gray ORFs indicate other proteins or proteins of unknown function.