Staphylococcus aureus is an important food poisoning bacterium. In food preservation, acidification is a well-known method. Permeant weak organic acids, like lactic and acetic acids, are known to be more effective against bacteria than inorganic strong acids (e.g., HCl). Growth experiments and metabolic and transcriptional analyses were used to determine the responses of a food pathogenic S. aureus strain exposed to lactic acid, acetic acid, and HCl at pH 4.5. Lactic and acetic acid stress induced a slower transcriptional response and large variations in growth patterns compared with the responses induced by HCl. In cultures acidified with lactic acid, the pH of the medium gradually increased to 7.5 during growth, while no such increase was observed for bacteria exposed to acetic acid or HCl. Staphylococcus aureus increased the pH in the medium mainly through accumulation of ammonium and the removal of acid groups, resulting in increased production of diacetyl (2,3-butanedione) and pyrazines. The results showed flexible and versatile responses of S. aureus to different types of acid stress. As measured by growth inhibition, permeant organic acid stress introduced severe stress compared with the stress caused by HCl. Cells exposed to lactic acid showed specific mechanisms of action in addition to sharing many of the mechanisms induced by HCl stress.

Background: High-pressure processing (HPP) is a commonly used technique in the food industry to inactivate pathogens, including L. monocytogenes. It has been shown that L. monocytogenes is able to recover from HPP injuries and can start to grow again during long-term cold storage. To date, the gene expression profiling of L. monocytogenes during HPP damage recovery at cooling temperature has not been studied. In order identify key genes that play a role in recovery of the damage caused by HPP treatment, we performed RNA-sequencing (RNA-seq) for two L. monocytogenes strains (barotolerant RO15 and barosensitive ScottA) at nine selected time points (up to 48 h) after treatment with two pressure levels (200 and 400 MPa). Results: The results showed that a general stress response was activated by SigB after HPP treatment. In addition, the phosphotransferase system (PTS; mostly fructose-, mannose-, galactitol-, cellobiose-, and ascorbate-specific PTS systems), protein folding, and cobalamin biosynthesis were the most upregulated genes during HPP damage recovery. We observed that cell-division-related genes (divIC, dicIVA, ftsE, and ftsX) were downregulated. By contrast, peptidoglycan-synthesis genes (murG, murC, and pbp2A) were upregulated. This indicates that cell-wall repair occurs as a part of HPP damage recovery. We also observed that prophage genes, including anti-CRISPR genes, were induced by HPP. Interestingly, a large amount of RNA-seq data (up to 85%) was mapped to Rli47, which is a non-coding RNA that is upregulated after HPP. Thus, we predicted that Rli47 plays a role in HPP damage recovery in L. monocytogenes. Moreover, gene-deletion experiments showed that amongst peptidoglycan biosynthesis genes, pbp2A mutants are more sensitive to HPP. Conclusions: We identified several genes and mechanisms that may play a role in recovery from HPP damage of L. monocytogenes. Our study contributes to new information on pathogen inactivation by HPP.

The efficacy of applying ultrasound (US) as a system to homogenize emulsions has been widely demonstrated. However, research has not yet shown whether the effect achieved by homogenizing milk with US is modified by subsequent pasteurization treatments that use new processing technologies such as pulsed electric fields (PEF), microwaves (MW), and high hydrostatic pressure (HPP). The aim of this study was, therefore, to optimize the application of US for milk homogenization and to evaluate the effect of PEF, HPP, and MW pasteurization treatments on the sensorial, rheological, and microbiological properties of milk throughout its shelf life. To homogenize whole milk, a continuous US system (20 kHz, 0.204 kJ/mL, 100%, 40 °C) was used, and different ultrasonic intensities (0.25, 0.5, and 1.0 kJ/mL) were evaluated. The optimal ultrasonic treatment was selected on the basis of fat globule size distribution and pasteurization treatments by MW (5800 W, 1.8 L/min), PEF (120 kJ/kg, 20 kV/cm) and HPP (600 MPa, 2 min, 10 °C) was applied. The ultrasound intensity that achieved the highest reduction in fat globule size (0.22 ± 0.02 µm) and the most homogeneous distribution was 1.0 kJ/mL. Fat globule size was smaller than in commercial milk (82% of volume < 0.5 µm for US milk versus 97% of volume < 1.2 µm for commercial milk). That size was maintained after the application of the different pasteurization treatments, and the resulting milk had better emulsion stability than commercial milk. After 28 days of storage, no differences in viscosity (4.4–4.9 mPa s) were observed. HPP pasteurization had the greatest impact on color, leading to higher yellowness values than commercial milk. Microbial counts did not vary significantly after 28 days of storage, with counts below 10² CFU/mL for samples incubated at 15 °C and at 37 °C. In summary, the homogenization of milk obtained by US was not affected by subsequent pasteurization processes, regardless of the technology applied (MW, PEF, or HPP). Further research is needed to evaluate these procedures’ effect on milk’s nutritional and functional properties.

Background: High pressure processing (HPP; i.e. 100–600 MPa pressure depending on product) is a non-thermal preservation technique adopted by the food industry to decrease significantly foodborne pathogens, includingListeria monocytogenes, from food.However, susceptibility towards pressure differs among diverse strains ofL. monocytogenesand it is unclear if this is due to their intrinsic characteristics related to genomic content. Here, we tested the barotolerance of 10 differentL. monocytogenesstrains, from food and food processing environments and widely used reference strains including clinical isolate, to pressure treatments with 400 and 600 MPa. Genome sequencing and genome comparison of the testedL. monocytogenesstrains were performed to investigate the relation between genomic profile and pressure tolerance.Results: None of the tested strains were tolerant to 600 MPa. A reduction of more than 5 log₁₀was observed for all strains after 1 min 600 MPa pressure treatment.L. monocytogenesstrain RO15 showed no significant reduction in viable cell counts after 400 MPa for 1 min and was therefore defined as barotolerant. Genome analysis of so far unsequencedL. monocytogenesstrain RO15, 2HF33, MB5, AB199, AB120, C7, and RO4 allowed us to compare the gene content of all strains tested. This revealed that the three most pressure tolerant strains had more than one CRISPR system with self-targeting spacers. Furthermore, several anti-CRISPR genes were detected in these strains. Pan-genome analysis showed that 10 prophage genes were significantly associated with the three most barotolerant strains.Conclusions: L. monocytogenesstrain RO15 was the most pressure tolerant among the selected strains. Genome comparison suggests that there might be a relationship between prophages and pressure tolerance inL. monocytogenes.