Biofilm formation ability of Salmonella enterica serovar Typhimurium acrAB mutants.

Recent studies offer contradictory findings about the role of multidrug efflux pumps in bacterial biofilm development. Thus, the aim of this study was to investigate the involvement of the AcrAB efflux pump in biofilm formation by investigating the ability of AcrB and AcrAB null mutants of Salmonella enterica serovar Typhimurium to produce biofilms. Three models were used to compare the ability of S. Typhimurium wild-type and its mutants to form biofilms: formation of biofilm on polystyrene surfaces; production of biofilm (mat model) on the air/liquid interface; and expression of curli and cellulose on Congo red-supplemented agar plates. All three investigated genotypes formed biofilms with similar characteristics. However, upon exposure to chloramphenicol, formation of biofilms on solid surfaces as well as the production of curli were either reduced or were delayed more significantly in both mutants, whilst there was no visible effect on pellicle formation. It can be concluded that when no selective pressure is applied, S. Typhimurium is able to produce biofilms even when the AcrAB efflux pumps are inactivated, implying that the use of efflux pump inhibitors to prevent biofilm formation is not a general solution and that combined treatments might be more efficient. Other factors that affect the ability to produce biofilms depending on efflux pump activity are yet to be identified.

Resistance to essential oils affects survival of Salmonella enterica serovars in growing and harvested basil.

The number of outbreaks of food-borne illness associated with consumption of fresh products has increased. A recent and noteworthy outbreak occurred in 2007. Basil contaminated with Salmonella enterica serovar Senftenberg was the source of this outbreak. Since basil produces high levels of antibacterial compounds the aim of this study was to investigate if the emerging outbreak reflects ecological changes that occurred as a result of development of resistance to ingredients of the basil oil. We irrigated basil plants with contaminated water containing two Salmonella serovars, Typhimurium and Senftenberg, and showed that Salmonella can survive on the basil plants for at least 100 days. S. Senftenberg counts in the phyllosphere were significantly higher than S. Typhimurium, moreover, S. Senftenberg was able to grow on stored harvested basil leaves. Susceptibility experiments demonstrated that S. Senftenberg is more resistant to basil oil and to its antimicrobial constituents: linalool, estragole and eugenol. This may indicate that S. Senftenberg had adapted to the basil environment by developing resistance to the basil oil. The emergence of resistant pathogens has a significant potential to change the ecology, and opens the way for pathogens to survive in new niches in the environment such as basil and other plants.

Inactivation of foodborne pathogens in ground beef by cooking with highly controlled radio frequency energy.

The consumer demand for fresh tasting, high quality, low salt, preservative-free meals which require minimal preparation time magnifies the safety concern and emphasizes the need to use innovative technologies for food processing. A modern technique to uniformly heat and cook foods is based on a combination of convection and controlled radio frequency (RF) energy. However any advantage conferred on meat cooked by this method would be lost if application of the technology results in decreased safety. Our main goal was to study the inactivation efficacy of this method of cooking against pathogens in ground meat in comparison to standard convection cooking. Meat balls were artificially inoculated with GFP expressing Escherichia coli, Salmonella Typhimurium and Listeria monocytogenes as well as spores of Bacillus cereus and Bacillus thuringiensis and cooked by convection heating (220°C, 40 min), by using energy generated from frequencies in the RF bandwidth (RF cooking, 7.5 min) or by combined heating (5.5 min), until the center temperature of each sample reached 73°C. The mean reductions in total indigenous bacteria obtained by RF and convection were 2.8 and 2.5 log CFU/g, respectively. Cooking of meat balls with convection reduced the E. coli population (8 log CFU/g) by 5.5 log CFU/g, whilst treatment with RF reduced E. coli population to undetectable levels. The mean reductions of S. Typhimurium obtained by RF and convection were 5.7 and 6.5 log CFU/g, respectively. The combined treatment reduced the Salmonella population to undetectable levels. In contrast, L. monocytogenes was poorly affected by RF cooking. The mean reduction of L. monocytogenes obtained by RF energy was 0.4 log CFU/g, while convection cooking resulted in undetectable levels. Interestingly, the combined treatment also resulted with undetectable levels of Listeria although time of cooking was reduced by 86%. One-step cooking had negligible effects on the Bacillus spores and therefore a 2-step treatment of RF or convection was applied. This 2-step treatment proved to be efficient with 4.5 log CFU/g reduction for both RF and convection. In conclusion, here we show that combination of RF with convection cooking resulted in similar or even better effects on selected foodborne pathogens compared to convection only, while the time required for safe cooking is cut down by up to 86%. The equal or better results in the levels of all investigated pathogens using RF with convection compared with convection only suggest that this technology looks promising and safe for ground beef cooking.

Presence and persistence of Salmonella enterica serotype typhimurium in the phyllosphere and rhizosphere of spray-irrigated parsley.

Salmonella enterica is one of the major food-borne pathogens associated with ready-to-eat fresh foods. Although polluted water might be a significant source of contamination in the field, factors that influence the transfer of Salmonella from water to the crops are not well understood, especially under conditions of low pathogen levels in water. The aim of this study was to investigate the short- and long-term (1 h to 28 days) persistence of Salmonella enterica serotype Typhimurium in the phyllosphere and the rhizosphere of parsley following spray irrigation with contaminated water. Plate counting and quantitative real-time PCR (qRT-PCR)-based methods were implemented for the quantification. By applying qRT-PCR with enrichment, we were able to show that even irrigation with water containing as little as ∼300 CFU/ml resulted in the persistence of S. Typhimurium on the plants for 48 h. Irrigation with water containing 8.5 log CFU/ml resulted in persistence of the bacteria in the phyllosphere and the rhizosphere for at least 4 weeks, but the population steadily declined with a major reduction in bacterial counts, of ∼2 log CFU/g, during the first 2 days. Higher levels of Salmonella were detected in the phyllosphere when plants were irrigated during the night compared to irrigation during the morning and during winter compared to the other seasons. Further elucidation of the mechanisms underlying the transfer of Salmonella from contaminated water to crops, as well as its persistence over time, will enable the implementation of effective irrigation and control strategies.

A comparative study assaying commonly used sanitizers for antimicrobial activity against indicator bacteria and a Salmonella Typhimurium strain on fresh produce.

With increased concerns over failures in vegetable and fruit sanitation, evaluating the efficacy of widely approved chemicals is ever more important. The purpose of this study was to determine whether sanitation treatments are equally effective against indicator bacteria and human enteric pathogens on cucumber and parsley. We provide here an experimental overview on the efficacy of common sanitation methods, which are based on peracetic acid-hydrogen peroxide, sodium dichloroisocyanurate, and the quaternary ammonium compound didecyldimethylammonium chloride. The sanitizers were tested for their activity against natural populations of total aerobic microorganisms, enterococci, and coliforms, and against the enteric pathogen Salmonella Typhimurium ATCC 14028 (which was added artificially). Results revealed that compared with washing parsley and cucumbers with water, treatments with all three sanitizers were not effective, resulting in a maximal reduction of only 0.7 log CFU of Salmonella Typhimurium. These sanitizers were also not effective in removal of natural bacteria from parsley (maximal reduction was 0.7 log CFU). Sanitation of cucumber was more successful; peracetic acid showed the most effective result, with a reduction of 2.7 log in aerobic microorganisms compared with cucumbers washed with water. Still, removal of natural bacteria from cucumbers proved more efficient than the removal of Salmonella Typhimurium. This may create a debate about the necessity of the sanitation and its contribution to safety, because sanitation of some contaminated vegetables may result in an increased likelihood of foods that, although they are given good hygienic ratings due to low microbial counts, harbor pathogens.