The clinic vs the farm: exploring prevalence and function of CRISPR-Cas in agriculturally relevant niches

Authors: UPRETI, CHAHAT - University Of Texas; Durso, Lisa; PALMER, KELLI - University Of Texas

Submitted to: Annual Wind River Conference on Procaryotic Biology
Publication Type: Abstract Only
Publication Acceptance Date: 5/2/2022
Publication Date: 6/1/2022
Citation: Upreti, C., Durso, L.M., Palmer, K. 2022. The clinic vs the farm: exploring prevalence and function of CRISPR-Cas in agriculturally relevant niches. Annual Wind River Conference on Procaryotic Biology. Available:.

Interpretive Summary: The development and use of antibiotics has revolutionized human health and livestock production. However, due to the use and misuse of antibiotics, we are now facing a global issue of antibiotic resistance. More than a million people each year die from antibiotic-resistant infections. One of the ways in which bacterial pathogens become resistant to antibiotics is via transfer of resistance genes from one bacterium to another, on DNA fragments called plasmids. To protect themselves from foreign genetic elements such as plasmids, bacteria have evolved several defense systems, one of which is called CRISPR-Cas. It has been shown previously that bacteria that possess a functioning CRISPR-Cas system are significantly less likely to be multi-drug resistant. This has several potential applications since we can leverage CRISPR presence or CRISPR-based technologies to tackle antibiotic resistance. However, much of this work has been done on strains from clinical origins. Bacteria from agricultural niches are also exposed to a wide variety of antibiotics, and multi-drug resistance is a problem in these niches, but how prevalent CRISPR-Cas is in them is not well known. In this work, we try to answer this question, by focusing on the bacterium Enterococcus faecalis. E. faecalis is both clinically relevant, as one of the major causes of hospital acquired infections in US, and also agriculturally relevant, since it colonizes most animals and has been implicated in transfer of resistance from food animals to humans. We found a clear similarity in CRISPR type diversity in E. faecalis strains from humans and animals, and also a large overlap between the CRISPR targets in both groups. To test whether CRISPR-Cas is functional in this environment, we tested its ability to block plasmid transfer in manure and found that its efficacy is lower than in lab conditions initially, but surpasses lab conditions after 48 hours. Our work is among the first to explore prevalence and efficacy of CRISPR-Cas in agricultural environments, and it lays the foundation for further CRISPR-based interventions to tackle antibiotic resistance.

Technical Abstract: Global efforts to combat antibiotic resistance highlight the need for a OneHealth approach that includes human, animal, and environmental perspectives. One of the major drivers of resistance transmission is horizontal gene transfer. Based on its role as an adaptive defense mechanism against horizontal gene transfer in bacteria, CRISPR-Cas is a possible tool to address this crisis. Most studies in this area, however, focus on clinically-derived strains. As a result, the prevalence of CRISPR-Cas in bacteria from agricultural sources and its efficacy in those environments is poorly understood. In recent years, Enterococcus faecalis has emerged as an important zoonotic pathogen and is regularly isolated from human and animal guts. Here, we perform a comprehensive analysis of 1,985 publicly available E. faecalis genomes from human and animal sources for CRISPR-Cas content and diversity. We found that the presence of CRISPR-Cas and multidrug resistance were negatively correlated. Genomes from both sources showed similar trends in Type II CRISPR-Cas occurrence. Examination of CRISPR spacer profiles from these different environments revealed reduced redundancy and higher target diversity in spacers from animal-sourced E. faecalis as compared to their human counterpart. While there was a large overlap between spacers with targets across both groups, we found several spacers specifically enriched in animal and human niches. We tested the functional efficacy of CRISPR-Cas in an agricultural environment by evaluating its ability to block plasmid transfer in manure. Efficacy in manure was lower than in solid or liquid media initially but surpassed them after 48 hours. Together, our work shows for the first time a comprehensive picture of CRISPR-Cas prevalence and efficacy in agriculturally relevant niches for the generalist bacterium E. faecalis.