Anti-infective activities of long-chain fatty acids against foodborne pathogens

Abstract

Free fatty acids (FFAs) have long been acknowledged for their antimicrobial activity. More recently, long-chain FFAs (>12 carbon atoms) are receiving increased attention for their potent antivirulence activity against pathogenic bacteria. In the gastrointestinal tract, foodborne pathogens encounter a variety of long-chain FFAs derived from the diet, metabolic activities of the gut microbiota, or the host. This review highlights the role of long-chain FFAs as signaling molecules acting to inhibit the infectious potential of important foodborne pathogens, including Salmonella and Listeria monocytogenes. Various long-chain FFAs interact with sensory proteins and transcriptional regulators controlling the expression of infection-relevant genes. Consequently, long-chain FFAs may act to disarm bacterial pathogens of their virulence factors. Understanding how foodborne pathogens sense and respond to long-chain FFAs may enable the design of new anti-infective approaches.

Keywords: Listeria monocytogenes; Salmonella; antimicrobial activity; antivirulence activity; foodborne pathogens; long-chain free fatty acids.

Figure 1.

Long-chain FFAs inhibit the expression of SPI-1 virulence genes in Salmonella. The regulatory proteins HilD, HilC, and RtsA form a feed-forward loop controlling the virulence regulator HilA, which activates expression of SPI-1 genes. Long-chain FFAs, such as oleic acid (C18:1) and the DSF compound c2-HDA (Table S1, Supporting Information), interfere with the regulatory activity of HilD. Furthermore, c2-HDA promotes Lon-mediated degradation of HilD and inhibits the regulatory activities of HilC and RtsA.

Figure 2.

Model of how medium- and long-chain FFAs interfere with AraC-like transcriptional regulators. AraC family members, such as HilD in Salmonella, ToxT in V. cholerae, and Rns in EHEC, are known to activate the expression of virulence genes. Specific medium- and/or long-chain FFAs inhibit the stability, dimerization and/or DNA-binding activity of AraC-like regulators.

Figure 3.

Long-chain unsaturated FFAs inhibit the PhoP/PhoQ two-component system in Salmonella. The histidine kinase PhoQ autophosphorylates in the presence of an activating signal, such as Mg²⁺ limitation, cationic antimicrobial peptides (CAMPs), low pH, and high osmolarity. The phosphoryl group is transferred to the response regulator PhoP, which acts as a transcription regulator of virulence genes. Long-chain unsaturated FFAs, such as palmitoleic acid (C16:1) and linoleic acid (C18:2) (Table S1, Supporting Information), interfere with the autokinase activity of PhoQ, thereby preventing phosphorylation of the cytoplasmic response regulator PhoP. Consequently, PhoP-dependent regulation of virulence genes is repressed. OM: outer membrane. IM: inner membrane.

Figure 4.

Lack of GlcNAc glycosylation on WTAs promotes FFA tolerance in L. monocytogenes. In the wildtype situation, L. monocytogenes serotype 1/2a contains two modifications of the WTAs: GlcNAc and rhamnose. Antimicrobial medium- and long-chain FFAs are expected to target the bacterial membrane (left). FFA-tolerant strains lacking GlcNAc glycosylation on WTAs are characterized by a more hydrophilic cell surface and reduced binding of FFAs (right). Consequently, the FFAs are repulsed from the bacterial surface, thereby protecting the mutant strain from FFA toxicity. PG: peptidoglycan layer. CM: cytoplasmic membrane.

Figure 5.

Medium- and long-chain FFAs inhibit PrfA-mediated expression of virulence factors required for the intracellular lifestyle of L. monocytogenes. (A) The internalins InlA and InlB promote bacterial entry into nonphagocytic host cells. The pore-forming toxin LLO, and the phospholipases PlcA and PlcB, enable bacterial escape from the primary vacuole. Inside the cytosol, the pathogen multiplies and spreads to adjacent cells using the actin assembly-inducing protein ActA. Finally, LLO, PlcA, and PlcB promote bacterial escape from the secondary vacuole formed upon entry of L. monocytogenes into the neighboring cell. (B) The transcriptional regulator PrfA activates transcription of virulence genes required for the intracellular lifestyle of L. monocytogenes. The level or activity of PrfA is affected by various signals from the environment, such as high temperature, carbohydrates, and glutathione (GSH). Furthermore, exposure to medium-chain saturated and long-chain unsaturated FFAs prevent PrfA from activating transcription of virulence genes encoding InlA, LLO, ActA, and so on.

Figure 6.

Model of how medium- and long-chain FFAs interfere with the virulence regulator PrfA. Specific medium-chain saturated and long-chain unsaturated FFAs are known to prevent PrfA from binding to DNA. Furthermore, exposure to FFAs inhibit PrfA-dependent activation of virulence genes in L. monocytogenes. These observations support a model where specific medium- and long-chain FFAs inhibit the dimerization and/or DNA-binding activity of PrfA.