Investigating a role for nitric oxide in the control of biofilm and zoonotic pathogen colonisation of the spinach phylloplane

Abstract

Outbreaks of food poisoning associated with zoonotic pathogen contamination of RTE fresh products such as leafy salads are on the increase and new strategies for the control of pathogen contamination are required. Salmonella spp. are of particular concern as they remain the main cause of fresh produce associated food poisoning outbreaks in the U.S. and Europe. Efficient colonisation of spinach leaves by S. Thompson and S. Typhimurium, and the formation of bacterial biofilms, was demonstrated using EDIC/EF microscopy. The novel biofilm dispersal agent, nitric oxide, did not induce detachment of Salmonella spp from the phylloplane but did influence attachment and colonisation of spinach leaves by Salmonella spp. Factors influencing the attachment to and colonisation of surfaces by S. Typhimurium, (temperature and flagellar expression) were investigated and found to influence, but not be essential for surface colonisation and biofilm formation. Nitric oxide treatment also did not induce dispersal of S. Typhimurium biofilms in an abiotic surface model system. However, a proteomic investigation demonstrated that nitric oxide signaling is active in S. Typhimurium and that proteins involved in signaling and in energy production consistent with existence outside of a biofilm state were up-regulated. Nitric oxide treatment was also investigated for its potential to induce biofilm dispersal for indigenous phylloplane populations for the improvement of product quality and was not found to induce significant removal of indigenous bacteria from leaves. Together these investigations demonstrate that there remains much more to be understood about the interactions of zoonotic pathogens and indigenous phylloplane populations with fresh produce surfaces for development of novel decontamination strategies such as the use of nitric oxide for biofilm dispersal.

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Identifiers

ORCID for Charles W. Keevil: orcid.org/0000-0003-1917-7706

ORCID for Jeremy S. Webb: orcid.org/0000-0003-2068-8589

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