Incidence rates of resistant enterotoxigenic Escherichia coli in fresh vegetables and salads.

  1. Carlos Ramón Vázquez-Quiñones
  2. Monica Rincón-Guevara
  3. Ivan Natividad-Bonifacio
  4. Carlos Vázquez-Salinas
  5. Humberto González-Márquez

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Abstract

1.Abstract Diarrheal diseases remain a significant global health challenge, particularly in developing regions such as Africa, Asia, and Latin America, where they are a leading cause of child mortality. Contaminated food, including raw or undercooked vegetables, is a major transmission route for diarrheal pathogens such as norovirus, Campylobacter, non-typhoid Salmonella, and pathogenic Escherichia coli. This study aimed to assess the prevalence of enterotoxigenic Escherichia coli (ETEC), a key diarrheal pathogen, in fresh produce and prepared salads in Mexico City. A total of 334 samples, including prepared salads (lettuce, carrots, and tomatoes) and unprocessed coriander and lettuce, were analyzed over two years using protocols from the Bacteriological Analytical Manual and the Official Mexican Standard (NOM) SSA 210. Genotyping was performed to detect ETEC-specific virulence genes encoding heat-stable and heat-labile enterotoxins (st and lt) respectively. ETEC was identified in 9.9% of the total samples, representing 51.56% of the confirmed E. coli isolates. Contamination rates varied by food type, with coriander showing the highest prevalence (78.78%), followed by lettuce (9.09%), and prepared salads from La Vicentina Market (9.09%) and La Purísima Market (3.03%). Genotyping revealed that 12.12% of the ETEC-positive samples carried both st and lt genes, while 33.3% and 54.6% carried only the lt or st gene, respectively. In lettuce samples, 9.09% were positive for ETEC, with 3.03% carrying the lt gene, 3.03% the st gene, and 3.03% both genes. Similarly, in coriander, 21.21% were positive for the lt gene, 51.51% for the st gene, and 6.06% for both genes. These findings highlight the widespread presence of ETEC in fresh produce sold in Mexico City, posing a significant public health risk, particularly given the increasing consumption of raw vegetables. The study provides the first reported data on ETEC contamination ratios in Mexico City, emphasizing the urgent need for improved food safety measures, including better hygiene practices during production, handling, and preparation of fresh produce. This research underscores the importance of ongoing surveillance and preventive strategies to mitigate the risk of foodborne diarrheal diseases in urban populations.

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  1. Version published to 10.1099/acmi.0.000957.v2 on Access Microbiology
  2. Access Microbiology

    The reviewers have highlighted major & minor concerns with the work presented. Please ensure that you address their comments. Please provide more detail in the Methods section and ensure that software is consistently cited and its version and parameters included. The reviewers believe the results shown in the manuscript only partially support the conclusions presented - please address this through clearer presentation and description of the results, and/or further experiments. In addition, the paper is poorly structured and written, which, to some extent, has prevented a proper assessment of the research done.

  3. Access Microbiology

    Comments to Author

    Dear author I am writing to your manuscript (Incidence and multidrug resistance of Escherichia coli pathotypes on fresh vegetables and salads). I have some comments and questions that need your answers - Line 112 (and to determine the prevalence of resistance)
, you used the word (incidence) in the title, please rephrase this sentence as there is a difference between the meaning of the two words . Lines 137-139 (Further identification using MALDI/TOF TOF 
 corroborated the presence of other microorganisms within the coliform group, which 
had biochemical characteristics like those of E. coli. Please identify the biochemical reaction tests used and please explain why these reactions couldn't identify these Gram-negative bacilli (E.coli which is motile, indole positive from Klebsiella pneumoniae 
 which is non motile, indole negative organism)? - Line 374-376 (These cultures were streaked massively onto a Mueller-Hinton agar plate where filter paper discs were placed on equidistant zones, each one impregnated with the following antibiotics)
 what do mean by streaked massively, please explain if you used standardized method as 0.5 McFarland. - Your manuscript is quit long and there is redundancy in many parts especially in discussion and recommendation sections Thank you

    Please rate the manuscript for methodological rigour

    Satisfactory

    Please rate the quality of the presentation and structure of the manuscript

    Good

    To what extent are the conclusions supported by the data?

    Partially support

    Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?

    No

    Is there a potential financial or other conflict of interest between yourself and the author(s)?

    No

    If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?

    Yes

  4. Access Microbiology

    Please rate the manuscript for methodological rigour

    Good

    Please rate the quality of the presentation and structure of the manuscript

    Good

    To what extent are the conclusions supported by the data?

    Strongly support

    Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?

    No

    Is there a potential financial or other conflict of interest between yourself and the author(s)?

    No

    If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?

    Yes

  5. Access Microbiology

    Comments to Author

    I have reviewed the MS ACMI D 2400 239 entitled "Incidence and multidrug resistance of Escherichia cole prototypes on fresh vegetables and salads" The authors have successfully identified an important area in public health and food safety; however, the following comments must be addressed. Title: I think the tile needs minor revision, I suggest: Incidence rates of resistant pathotypes of Escherichia coli 1 in fresh vegetables and salads. The Abstract is not well written; authors do not indicate why they analyzed food; they did not specify the specific objective and type of E. coli targeted until later and it was only implying that ETEC was involved which could be highlighted in the title. Please indicate either in abstract or title the target organism(s). The authors mentioned genes and genotyping before mentioning the lineage and types targeted, only E. coli was mentioned early on. Even though we known, the genes are abbreviated should be first written out for broad range readers to avoid confusions could be read as (shiga toxin, labile toxin, soluble toxin) ..what heat stable STa types (called STh and STp, encoded by the estA gene). Please clarify. Understandable, the uidA gene coding beta-glucuronidase is for species level detection or genotyping or both. What are the genotypes codes or how are they coded, and what was the dominant genotypic clone, other than a general pathotypic profile mentioned. If not genotype, then I suggest remove the approach and replace with characterized. Is it a multiplex or single gene PCR, Realtime, or conventional? It is not clear in abstract, please clarify and organize the abstract.They should be written out at first appearance. These points reduced the quality and trust. Abstract needs revision: state the objectives after background info, then methods, and results this seq is important. L 38-39 it is not limited to only these, please revise or finish sentence with …etc or foodborne enteric bacteria. Please do not specify or limit to which types cause diarrhea from these lineages ETEC, EIEC, EHEC, EPAEC…etc. L41, please write out on first occurrence, especially this is a method. L43 44, (ETEC was identified in 51.56% of the confirmed isolates, representing 9.9% of the samples collected over two years), the 10% representation refers to ETEC or the confirmed isolates? L43 confirmed E. coli? L47 48, these lines imply the genes are not stable markers to genotype, or do the authors mean characterization for gene content? L49, please explain and 9.9% was the representative rate in L 44 43 for all and, and here, lattice is a main issue? L56 the Abstract concludes with a positive general statement; however, not sure if only one type of food (salads or raw foods) are the sources since general hygienic conditions were of concern. 59, some of the keywords are present in the title, please avoid L75 EDA Introduction L65, introduction is well written, and paragraphs discuss single relevant topics; however, there is a significant paucity in the review amount of the specific topic in question, different E. coli types are listed in a line or two, but no mention of their frequency rates or how endemic or epidemic they are. L115. Results As mentioned earlier, sample size could be less or justify. 117 123, do these numbers consistent with other tests' results? 141, ruled out, there not the organisms thought they were? Material and Method L 299 Do the sample size reflects the population or the city, how large is Iztapalapa Iztapalapamunicipality in Mexico? It feels sample size could be a limitation needs justification. The data seems a bit old, would it reflect current situation, no improvement anticipated? L329, you used a conventional method for presumptive detection, then used a conventional PCR? For detection uidA gene for species level, then how do you know the "pathotype" at this stage, and what other pathotypes were identified than ETEC? do you mean lineages in general, please be specific to help the reader understand. L331, please mentioned the specific gene names here too, and if they were multiplexed or single. It sounds from PCR conditions, these were single gene PCR, and up until now, other types were not described earlier. I think objectives need to be clear how many and what of these types were studied differently. L341, is this machine was a multigene multiplex or is a name L352, the MALDI Biotyping results should be magnified a bit because it explains how types were detected. L368 what was the version of CLSI L374, please revise (…streaked massively" is a bit confusion, they should be spread evenly not to exceed conc L383 this is the supplement version, please add it above too, does it represent the reference breakpoints in your area for the organism used. Please add some more references and review the list for consistency.

    Please rate the manuscript for methodological rigour

    Satisfactory

    Please rate the quality of the presentation and structure of the manuscript

    Poor

    To what extent are the conclusions supported by the data?

    Partially support

    Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?

    No

    Is there a potential financial or other conflict of interest between yourself and the author(s)?

    No

    If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?

    Yes

  6. Version published to 10.1099/acmi.0.000957.v1 on Access Microbiology