Recombinant production of a diffusible signal factor inhibits Salmonella invasion and animal carriage

ABSTRACT

The complex chemical environment of the intestine is defined largely by the metabolic products of the resident microbiota. Enteric pathogens, elegantly evolved to thrive in the gut, use these chemical products as signals to recognize specific niches and to promote their survival and virulence. Our previous work has shown that a specific class of quorum-sensing molecules found within the gut, termed diffusible signal factors (DSF), signals the repression of Salmonella tissue invasion, thus defining a means by which this pathogen recognizes its location and modulates virulence to optimize its survival. Here, we determined whether the recombinant production of a DSF could reduce Salmonella virulence in vitro and in vivo. We found that the most potent repressor of Salmonella invasion, cis-2-hexadecenoic acid (c2-HDA), could be recombinantly produced in E. coli by the addition of a single exogenous gene encoding a fatty acid enoyl-CoA dehydratase/thioesterase and that co-culture of the recombinant strain with Salmonella potently inhibited tissue invasion by repressing Salmonella genes required for this essential virulence function. Using the well characterized E. coli Nissle 1917 strain and a chicken infection model, we found that the recombinant DSF-producing strain could be stably maintained in the large intestine. Further, challenge studies demonstrated that this recombinant organism could significantly reduce Salmonella colonization of the cecum, the site of carriage in this animal species. These findings thus describe a plausible means by which Salmonella virulence may be affected in animals by in situ chemical manipulation of functions essential for colonization and virulence.

Plain Language Summary

Despite our best efforts, infections of agricultural animals with Salmonella persist, posing threats to food safety. Few, if any, measures have proven effective in reducing Salmonella carriage in animals used for food, a major source of this pathogen. Antibiotics are ineffective at curtailing infection and have served only to exacerbate the global crisis of antimicrobial resistance. The alternative then is to seek novel means to reduce Salmonella disease and carriage by preventing its colonization of livestock and poultry. Here we describe an approach targeting invasion, a function essential for Salmonella carriage and disease in animals. We show that a potent chemical inhibitor of invasion, the diffusible signal factor cis-2 hexadecenoic acid, can be produced by recombinant E. coli strains capable of stably colonizing the animal intestine, providing a means to directly affect the virulence of Salmonella within an animal host. These studies may thus provide a route to reduce the carriage of this pathogen in production animals and thus the spread of disease to humans.

KEYWORDS:

• Salmonella
• invasion
• diffusible signal factor
• cis-2-hexadecenoic acid
• rpfF
• virulence
• E. coli Nissle 1917
• chickens

Acknowledgments

We are grateful to the members of the Yung-Fu Chang lab and the Diel de Amorim lab for helping us with techniques and resources. The authors would also like to acknowledge the flow cytometry facility, and the proteomics and metabolomics facility at the Cornell Institute of Biotechnology for their resources.

Author contributions

CA, MA, and RC conceptualized the work; MA, RC, PPB, and CA conducted the experiments; MA, RC, PPB, and CA analyzed and curated the data; MA, RC, and PPB visualized the results; CA acquired funding and administered the study; CA wrote the draft manuscript; MA, RC, PPB, and CA reviewed and edited the manuscript.

Disclosure statement

No potential conflict of interest was reported by the authors.

Data availability statement

The data that support the findings of this study are openly available in Cornell University Library’s institutional repository, eCommons (https://ecommons.cornell.edu), for preservation and access (https://doi.org/10.7298/gv1c-b060). Datasets will be available via the world-wide web without restriction. eCommons provides each item with a persistent identifier and is committed to preserving the binary form of the digital object.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/19490976.2023.2208498.

Additional information

Funding

This project was supported by Agriculture and Food Research Initiative Competitive Grant no. 2021-08162 from the USDA National Institute of Food and Agriculture, and by NIH/NIAID grant R01 AI162944-01 to C.A.