Food-borne bacteria can cause life-threatening disease, yet it is still unclear why some strains are tolerated by the host. To find an answer, a project led by Professor Rosa del Carmen Rocha-Gracia (Benemérita Universidad Autónoma de Puebla, Mexico) and Professor Manel Camps (University of California Santa Cruz, USA) analysed the genetic diversity of Escherichia coli populations, comparing isolates from food to those from hospitalised patients (clinical strains). Food strains were distinct from clinical strains, but shared with them genes that contribute to virulence and antibiotic resistance. This gene exchange was found to be mediated by a specialised set of plasmids – rings of DNA – and the researchers defined a set of markers to monitor them.
Food poisoning is common, with toxic strains of Escherichia coli (E. coli) bacteria and Salmonella being two of the most frequent culprits. Tackling infections of food-borne bacteria is a top priority for public health bodies, as toxic strains can cause serious illness or even death. E. coli is a species of bacteria that exists naturally in our gut. While many strains are harmless or even beneficial to us, some strains of E. coli cause gastrointestinal problems. But one particularly dangerous subset can even infect organs outside of the intestines, resulting in urinary tract infections, pneumonia, meningitis, and sepsis.
E. coli can enter the body in various ways, including through contaminated food and water, untreated sewage, and person-to-person contact. Food is one of the most common sources of E. coli, and infections with toxic strains can lead to severe complications, including dehydration, sepsis, or organ failure. Worryingly, toxic strains of E. coli and Salmonella are becoming increasingly – and rapidly – resistant to common antibiotics used as first-line medications to treat infections. Therefore, a better understanding of the mechanisms leading to toxicity and antibiotic resistance in these bacteria is urgently needed.
Fortunately, Professor Rosa del Carmen Rocha-Gracia at Benemérita Universidad Autónoma de Puebla in Mexico and Professor Manel Camps at the University of California Santa Cruz in the USA are on the case. An integrated and unifying approach to public health, called One Health, recognises the complex intersection between pathogens and humans, animals, and plants. To meet the threat to public health posed by food-borne infections, Camps emphasises on the need for ‘holistic, multidisciplinary approaches’. The team works to understand how and why food-borne pathogens spread and how they acquire antibiotic resistance.
Understanding E. coli
Pathogens which cause food-borne illness must develop the capacity to occupy multiple niches. These bacteria persist on food or skin surfaces, and expand in the dark, warm, and oxygen-free environment of the intestine. The E. coli strains that cause severe illness have mastered both environments and developed the ability to colonise other organs as well. Understanding E. coli infections involves a holistic and multidisciplinary study of how the bacteria population adapts and changes as it moves between settings and the conditions which facilitate this evolution.
Scientists are working to understand how these pathogens are transmitted and how they cause disease by escaping the intestine while avoiding the human immune response. Virulence factors are proteins which help the bacteria bind to host cells, obtain micronutrients, secrete toxins, or evade the immune response. When the bacteria move into a new space, it is possible that the virulence factors that they carry with them are spreading to the resident bacteria population. Studying exactly how this occurs will help scientists determine how the bacteria population adjusts to its specific niche and the origin and movement of the virulence genes.
E. coli: from friend to foe
Scientists are still trying to figure out exactly how we are exposed to certain toxic strains of E. coli. One theory is that pathogenic (disease-causing) bacteria in the environment enter our bodies via contaminated food.
An alternative hypothesis is that strains of E. coli found in food can share their disease-causing genes with E. coli strains resident in our gut. The transfer of disease-causing properties to resident E. coli can enhance the ability of resident strains to cause disease.
To distinguish between these two hypotheses, María Balbuena-Alonso, a PhD student in Rocha-Gracia’s laboratory, examined the genomes of E. coli strains found in food. Comparing strains isolated from foods to toxic, disease-causing clinical strains isolated in the hospital, she found that the E. coli strains isolated from food sources were more closely related to each other than they were to the clinical strains. This implies that the clinical strains of the (increasingly antibiotic-resistant) bacteria responsible for disease are very different from the ones found in our food. It is therefore unlikely that the toxic bacteria are already abundant in contaminated food and we are simply consuming them in high enough numbers to become sick.
Instead, Balbuena-Alonso found more evidence supporting the second theory: abundant gene transfer between the two populations. This includes a high number of resistance genes. The presence of these genes in the food strains of E. coli is possibly due to the overuse of antibiotics in our food production systems and antibiotic contamination of wastewater. Overall, the researchers concluded that transfer of genes between bacteria cells is a more likely mechanism for the evolution and adaptation of E. coli populations.
Plasmids and horizontal gene transfer
Bacteria can share genes by exchanging circular rings of DNA called plasmids. Plasmids are self-replicating genetic material that can leave the cell to be taken up by another bacterium. This process is known as horizontal gene transfer.Pathogens which cause food-borne illness must develop the capacity to occupy multiple niches.
Many species of bacteria, including those commonly associated with food poisoning, can share plasmids via a specialised process of horizontal transfer called conjugation. During conjugation, two bacterial cells link together and one of them copies its plasmid and passes it to the other one. Note that the mating bacteria do not need to be the same species. Thus, the genes which help bacteria survive and infect a new host can be shared throughout a new bacteria population – once toxic and resistant bacteria introduce them via plasmid conjugation.
Traders of toxic genes
Plasmids that can facilitate gene-sharing between bacteria by conjugation can be identified because they carry a complete set of genes required for transfer, known as mobilisation (MOB) genes. The range of bacterial species these plasmids can transfer DNA to can be estimated using a software method called COPLA, which tracks plasmids across different microorganisms to see how many different hosts the bacteria could infect and classifies them according to the breadth of their mobility. Combining MOB and COPLA analysis, the researchers estimated that the plasmids from food strains of E. coli could move across a broad range of species within their ecological niche.
Tracking shared gene content between plasmids from food vs clinical plasmids, the researchers discovered that the plasmids that mediate and regulate the exchange of genes between E. coli populations represent a small fraction of all mobilisable ones. Thus, these specialised shuttle plasmids appear to facilitate the transfer of genes between bacteria originating in two vastly different ecosystems: food vs host. These shuttle plasmids are arguably a crucial factor facilitating access to virulence and antibiotic resistance for clinically toxic E. coli.
Genes behind virulence and antibiotic resistance
The researchers revealed that genes responsible for antibiotic resistance and virulence normally associated with clinical E. coli populations were also found in the food-borne E. coli. Specifically, resistance to the antibiotics carbapenems and colistin was already present in food-borne strains. This is especially worrying because carbapenems and colistin are usually used as a last resort when other, more common antibiotics have proven ineffective. On the virulence front, the team found that some of the plasmids identified in the food-borne E. coli strains were highly enriched for genes responsible for adhesion factors. These factors increase the bacteria’s ability to stick to food.
Analysing the gene content of these E. coli populations also provided evidence that food-borne E. coli provides a reservoir of virulence and antibiotic resistance genes, which can then be shared with clinical strains. These genes were found to readily jump between members of two vastly different ecological niches due to the significant role of shuttle plasmids.
Future genetic markers
Camps argues that ‘we need to understand how food pathogens are transmitted and persist in the environment and how their virulence factors and antibiotic resistance genes move across the microbiome of a given environmental niche’ to help develop a One Health approach for tackling infections like Salmonella and E. coli. The team’s research has contributed much towards this shared goal for global public health.
Recognising the importance of the newly identified shuttle plasmids, Camps, Rocha-Gracia, Balbuena-Alonso, and their team have identified genetic markers to help monitor them. These markers will be an essential tool to help monitor the evolution and spread of antibiotic resistance and virulence of E. coli populations. Thanks to the researchers, the new understanding of gene transfer between bacteria will enable new epidemiological surveillance tools to aid in the fight against disease outbreaks.
Personal ResponseAre shuttle plasmids a common feature of all horizontal gene transfer between bacteria species, or are they specific to pathogenic bacteria?
Shuttle plasmids are not restricted to pathogenic bacteria, although the horizontal process of DNA transference has contributed to the spread of virulence and antibiotic resistance genes among pathogenic bacteria. Whether the genetic traffic between two ecological niches is generally mediated by a specific subset of conjugative plasmids is still being determined because large-scale genomic information and tools needed to conduct these studies have not been available until very recently.
What inspired you to conduct this research?
We were inspired by the availability of enough fully sequenced strains to be able to explore the location of virulence and resistance factors of strains of E. coli from different sources. We focused on food strains because the question of how frequently they cause disease is still open. We reasoned that being able to see which genes were located on plasmids and to classify these plasmids according to mobilisation ability and host range allowed us to estimate the potential that food strains have to contribute to the spread of resistance and virulence to clinical strains.