Salmonella enterica Typhimurium is a facultative intra-cellular pathogen. This means that Salmonella is not only living in the host gut, but it can also infiltrate and survive in the tissues of mammals for long periods (Figure 1). Antibiotics can help to limit such life-threatening systemic dissemination in patients at risk and eventually eradicate Salmonella from the gut lumen. However, antibiotics rarely sterilize the host tissues where the pathogen forms slow growing or dormant persisters [1, 2].
After the end of the antibiotic therapy, persisters can wake up, reseed and occasionally bloom in the gut lumen . Thus, the ability to form persisters makes Salmonella diarrhea hard to treat with antibiotics. We found that this has two major evolutionary consequences.
Figure 1. Cryosection of an infected mouse cecum showing Salmonella Typhimurium (green) in a lamina propria cell (blue; α-ICAM-1; white: DAPI; red: epithelium). Image by Andreas Müller, ETH Zürich.
Promoting the transmission of virulent clones
Virulence factors are necessary to invade the host tissues. Therefore, intra-cellular persisters are mainly virulent clones that have the entire armoury of virulence factors. In contrast, attenuated clones that lose some key virulence factors tend to emerge naturally in the gut lumen. This happens because virulence factor expression is costly and represents a non-essential burden as long as gut inflammation is caused by some means [4, 5]. We call such attenuated mutants “defectors”, as they get a free ride on the gut inflammation that is triggered by S. Typhimurium cells that express the virulence factors. Upon antibiotic treatments, virulent intra-cellular persisters survive, while gut luminal defectors are eliminated. This de facto selects for Salmonella virulence .
Promoting resistance by horizontal gene transfer
After the disappearance of symptoms, i.e., when the intestinal inflammation and diarrhoea has ceased, the host microbiota usually re-colonizes the gut. In this situation, the full re-colonization of the gut lumen by S. Typhimurium from the host tissues might be difficult as the luminal niche is filled with competitors . Nevertheless, persisters can initiate a very efficient transfer of genetic information to their competitors. Selfish mobile genetic elements like conjugative plasmids, detected in many strains of Salmonella, can jump from reseeding ex-persisters cells to other Enterobacteriaceae occupying the gut lumen . These plasmids often harbour virulence and antibiotic resistance genes. Strikingly, this spread of the plasmids is very efficient even without selection for their genetic payload (Figure 2).
Figure 2. Schematic recapitulating how Salmonella persisters promote the spread of plasmids
These works show that infection of mice with the archetypal entero-pathogen S. Typhimurium offers the opportunity to understand the evolution of virulence and resistance in a relevant and tractable ecological context. From this, we also infer preventive and therapeutic strategies to fight pathogenic bacteria while keeping evolution on our side.
We discovered a way to prevent persister-related effects. Tissue invasion, mucosal disease and horizontal gene transfer can be reduced by high avidity Immunoglobulin A generated after oral vaccination with killed bacteria [7-9]. This is because specific IgA excreted into the intestinal lumen of vaccinated hosts enchains growing S. Typhimurium into clumps . This drastically limits infiltration of the tissues and gives a competitive edge to the competing microbiota thus ensuring elimination of Salmonella without the detrimental effects associated with classical antibiotic therapies.
We are currently asking how we can fortify the protection as an efficient means to prevent the spread of antibiotic resistance plasmids and the emergence of new pathogen variants.
1. Claudi, B., et al., Phenotypic variation of Salmonella in host tissues delays eradication by antimicrobial chemotherapy. Cell, 2014. 158(4): p. 722-733.
2. Kaiser, P., et al., Cecum lymph node dendritic cells harbor slow-growing bacteria phenotypically tolerant to antibiotic treatment. PLoS Biol., 2014. 12(2): p. e1001793.
3. Diard, M., et al., Antibiotic treatment selects for cooperative virulence of salmonella typhimurium. Curr. Biol., 2014. 24(17): p. 2000-2005.
4. Stecher, B., et al., Salmonella enterica serovar typhimurium exploits inflammation to compete with the intestinal microbiota. PLoS Biol., 2007. 5(10): p. 2177-2189.
5. Diard, M., et al., Stabilization of cooperative virulence by the expression of an avirulent phenotype. Nature, 2013. 494(7437): p. 353-356.
6. Lam, L.H. and D.M. Monack, Intraspecies competition for niches in the distal gut dictate transmission during persistent Salmonella infection. PLoS Pathog, 2014. 10(12): p. e1004527.
7. Bakkeren, E., et al., Salmonella persisters promote the spread of antibiotic resistance plasmids in the gut. Nature, 2019.
8. Moor, K., et al., High-avidity IgA protects the intestine by enchaining growing bacteria. Nature, 2017. 544(7651): p. 498-502.
9. Diard, M., et al., Inflammation boosts bacteriophage transfer between Salmonella spp. Science, 2017. 355(6330): p. 1211-1215.