Antimicrobial activity of suspensions and nanoemulsions of citral in combination with heat or pulsed electric fields.

The application of essential oils in form of nanoemulsions has been proposed as a method to improve their solubility in aqueous solutions, and hence their antimicrobial activity. The objective of this study was to evaluate the antimicrobial activity of citral, applied directly or in combined treatments with heat or pulsed electric fields (PEF), as a function of the inoculation procedure assayed: (i) a simple, vigorous shaking method by vortex agitation (suspension of citral; s-citral) or (ii) the previous preparation of nanoemulsions by the emulsion phase inversion (EPI) method (nanoemulsion of citral; n-citral). n-Citral was more effective in either inhibiting or inactivating Escherichia coli O157:H7 Sakai than s-citral. However, when combined with heat, a greater synergistic effect was observed with s-citral rather than with n-citral, either in lab media (pH 7·0 and 4·0) or apple juice. For instance, while almost 5 log10 cell cycles were inactivated in apple juice after 15 min at 53°C in the presence of 0·1 µl ml-1 of s-citral, the use of n-citral required 30 min. The use of nanoemulsions did not modify the slight synergism observed when citral and mild PEF were combined (150 µs, 30 kV cm-1 ). SIGNIFICANCE AND IMPACT OF THE STUDY: The exploration of different delivery systems of antimicrobial compounds such as citral in aqueous food products aids in the establishment of successful combined treatments for food preservation. While at room temperature, citral in form of a nanoemulsion shows a higher antimicrobial activity; its combination with heat would imply a partial loss of the outstanding synergistic lethal effect achieved when added in suspension form. Therefore, the most suitable procedure to magnify the synergism between heat and citral when processing juices would merely require an intense homogenization step prior to the combined treatment.

Listeria monocytogenes Inhibits Serotonin Transporter in Human Intestinal Caco-2 Cells.

Listeria monocytogenes is a Gram-positive bacterium that can cause a serious infection. Intestinal microorganisms have been demonstrated to contribute to intestinal physiology not only through immunological responses but also by modulating the intestinal serotonergic system. Serotonin (5-HT) is a neuromodulator that is synthesized in the intestinal epithelium and regulates the whole intestinal physiology. The serotonin transporter (SERT), located in enterocytes, controls intestinal 5-HT availability and therefore serotonin's effects. Infections caused by L. monocytogenes are well described as being due to the invasion of intestinal epithelial cells; however, the effect of L. monocytogenes on the intestinal epithelium remains unknown. The main aim of this work, therefore, was to study the effect of L. monocytogenes on SERT. Caco2/TC7 cell line was used as an enterocyte-like in vitro model, and SERT functional and molecular expression assays were performed. Our results demonstrate that living L. monocytogenes inhibits serotonin uptake by reducing SERT expression at the brush border membrane. However, neither inactivated L. monocytogenes nor soluble metabolites were able to affect SERT. The results also demonstrate that L. monocytogenes yields TLR2 and TLR10 transcriptional changes in intestinal epithelial cells and suggest that TLR10 is potentially involved in the inhibitory effect observed on SERT. Therefore, L. monocytogenes, through TLR10-mediated SERT inhibition, may induce increased intestinal serotonin availability and potentially contributing to intestinal physiological changes and the initiation of the inflammatory response.

Environmental and biological factors influencing the UV-C resistance of Listeria monocytogenes.

In this investigation, the effect of microbiological factors (strain, growth phase, exposition to sublethal stresses, and photorepair ability), treatment medium characteristics (pH, water activity, and absorption coefficient), and processing parameters (dose and temperature) on the UV resistance of Listeria monocytogenes was studied. The dose to inactivate 99.99% of the initial population of the five strains tested ranged from 21.84 J/mL (STCC 5672) to 14.66 J/mL (STCC 4031). The UV inactivation of the most resistant strain did not change in different growth phases and after exposure to sublethal heat, acid, basic, and oxidative shocks. The pH and water activity of the treatment medium did not affect the UV resistance of L. monocytogenes, whereas the inactivation rate decreased exponentially with the absorption coefficient. The lethal effect of UV radiation increased synergistically with temperature between 50 and 60 °C (UV-H treatment). A UV-H treatment of 27.10 J/mL at 55 °C reached 2.99 and 3.69 Log10 inactivation cycles of L. monocytogenes in orange juice and vegetable broth, and more than 5 Log10 cycles in apple juice and chicken broth. This synergistic effect opens the possibility to design UV combined processes for the pasteurization of liquid foods with high absorptivity.

Emergence of pulsed electric fields resistance in Salmonella enterica serovar Typhimurium SL1344.

In this investigation we selected and isolated a culture derived from Salmonella enterica serovar Typhimurium SL1344 with stable increased resistance to pulsed electric fields (PEF) after repeated rounds of PEF treatment and outgrowth of survivors. The resulting culture showed a higher resistance to PEF treatments under different treatment conditions. The acquisition of PEF resistance was only observed in stationary phase cells. The cytoplasmic membrane of the resistant variant showed a higher resilience against PEF treatments, since a lower permeabilization degree was observed after PEF treatments, in comparison to the parental strain. Resistance to PEF was also accompanied by a higher tolerance to acidic pH, hydrogen peroxide and ethanol, but not to heat. The occurrence of a PEF resistant variant in S. enterica serovar Typhimurium SL1344 emphasizes the need to further study the mechanisms of inactivation and resistance by PEF for an adequate design of safe treatments.

New insights in mechanisms of bacterial inactivation by carvacrol.

AIMS: To study the mechanism of bacterial inactivation by carvacrol and the influence of genetic and environmental factors in its antimicrobial activity. METHODS AND RESULTS: In general, bacterial inactivation by carvacrol was higher in the Gram-positive Listeria monocytogenes than in the Gram-negative Escherichia coli and at acidic pH. At pH 4.0, 25 µl l(-1) of carvacrol for 5 h inactivated 1 and more than 5 log(10) cycles of E. coli and L. monocytogenes populations, respectively. Genetic and environmental factors also influenced cell resistance to carvacrol: rpoS and sigB deletion decreased carvacrol resistance in E. coli and L. monocytogenes, respectively; a heat shock induced a phenomenon of cross-protection to carvacrol treatments. Repair of sublethal injuries in cell envelopes suggested that carvacrol targets lipid fractions and proteins of these structures. This result was corroborated by attenuated total reflectance infrared microspectroscopy analysis. CONCLUSIONS: This study shows critical genetic and environmental factors, such as rpoS or sigB and heat shocks, and reveals new microbial structures involved in the mechanism of bacterial inactivation by carvacrol. SIGNIFICANCE AND IMPACT OF THE STUDY: A better understanding of the mechanisms of microbial inactivation is of great relevance to design more appropriate carvacrol treatments with high antimicrobial effects.

Development of resistance in Cronobacter sakazakii ATCC 29544 to thermal and nonthermal processes after exposure to stressing environmental conditions.

AIMS: The objective was to study the response of Cronobacter sakazakii ATCC 29544 cells to heat, pulsed electric fields (PEF), ultrasound under pressure (Manosonication, MS) and ultraviolet light (UV-C) treatments after exposure to different sublethal stresses that may be encountered in food-processing environments. METHODS AND RESULTS: Cronobacter sakazakii stationary growth-phase cells (30°C, 24h) were exposed to acid (pH 4·5, 1h), alkaline (pH 9·0, 1h), osmotic (5% NaCl, 1h), oxidative (0·5mmoll(-1) H(2) O(2), 1h), heat (47·5°C, 1h) and cold (4°C, 4h) stress conditions and subjected to the subsequent challenges: heat (60°C), PEF (25kVcm(-1) , 35°C), MS (117µm, 200kPa, 35°C) and UV-C light (88·55mWcm(-2), 25°C) treatments. The inactivation kinetics of C. sakazakii by the different technologies did not change after exposure to any of the stresses. The combinations of sublethal stress and lethal treatment that were protective were: heat shock-heat, heat shock-PEF and acid pH-PEF. Conversely, the alkaline shock sensitized the cells to heat and UV-C treatments, the osmotic shock to heat treatments and the oxidative shock to UV-C treatments. The maximum adaptive response was observed when heat-shocked cells were subjected to a heat treatment, increasing the time to inactivate 99·9% of the population by 1·6 times. CONCLUSIONS: Cronobacter sakazakii resistance to thermal and nonthermal preservation technologies can increase or decrease as a consequence of previous exposure to stressing conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: The results help in understanding the physiology of the resistance of this emerging pathogen to traditional and novel preservation technologies.

Role of general stress-response alternative sigma factors σ(S) (RpoS) and σ(B) (SigB) in bacterial heat resistance as a function of treatment medium pH.

This investigation aimed to determine the role of general stress-response alternative sigma factors σ(S) (RpoS) and σ(B) (SigB) in heat resistance and the occurrence of sublethal injuries in cell envelopes of stationary-phase Escherichia coli BJ4 and Listeria monocytogenes EGD-e cells, respectively, as a function of treatment medium pH. Given that microbial death followed first-order inactivation kinetics (R(2)>0.95) the traditional D(T) and z values were used to describe the heat inactivation kinetics. Influence of rpoS deletion was constant at every treatment temperature and pH, making a ΔrpoS deletion mutant strain approximately 5.5 times more heat sensitive than its parental strain for every studied condition. Furthermore, the influence of the pH of the treatment medium on the reduction of the heat resistance of E. coli was also constant and independent of the treatment temperature (average z value=4.9°C) in both parental and mutant strains. L. monocytogenes EGD-e z values obtained at pH 7.0 and 5.5 were not significantly different (p>0.05) in either parental or the ∆sigB deletion mutant strains (average z value=4.8°C). Nevertheless, at pH 4.0 the z value was higher (z=8.4°C), indicating that heat resistance of both L. monocytogenes strains was less dependent on temperature at pH 4.0. At both pH 5.5 and 7.0 the influence of sigB deletion was constant and independent of the treatment temperature, decreasing L. monocytogenes heat resistance approximately 2.5 times. In contrast, the absence of sigB did not decrease the heat resistance of L. monocytogenes at pH 4.0. The role of RpoS in protecting cell envelopes was more important in E. coli (4 times) than SigB in L. monocytogenes (1.5 times). Moreover, the role of σ(S) in increasing heat resistance seems more relevant in enhancing the intrinsic resilience of the cytoplasmic membrane, and to a lesser extent, outer membrane resilience. Knowledge of environmental conditions related to the activation of alternative sigma factors σ(S) and σ(B) and their effects on heat resistance would help us to avoid and/or identify situations that increase bacterial stress resistance. Therefore, more efficient food preservation processes might be designed.

Inactivation of Cronobacter sakazakii by manothermosonication in buffer and milk.

The inactivation of Cronobacter sakazakii by heat and ultrasound treatments under pressure at different temperatures [manosonication (MS) and manothermosonication (MTS)] was studied in citrate-phosphate pH 7.0 buffer and rehydrated powdered milk. The inactivation rate was an exponential function of the treatment time for MS/MTS treatments (35-68 °C; 200 kPa of pressure; 117 µm of amplitude of ultrasonic waves) in both media, and for thermal treatments alone when buffer was used as heating media. Survival curves of C. sakazakii during heating in milk had a concave downward profile. Up to 50 °C, the lethality of ultrasound under pressure treatments was independent of the treatment temperature in both media. At temperatures greater than 64 °C in buffer and 68 °C in milk, the inactivating effect of MTS was equivalent to that of the thermal treatments alone at the same temperature. Between 50 and 64 ºC for buffer and 50 and 68 °C for milk, the lethality of MTS was the result of a synergistic effect, where the total lethal effect was higher than the lethal effect of heat added to that of ultrasound under pressure at room temperature. The maximum synergism was found at 60 °C in buffer and at 56 °C in milk. A heat treatment of 12 min (60 °C) or 4 min of an ultrasound under pressure at room temperature treatment (35 °C; 200 kPa; 117 µm) would be necessary to guarantee the death of 99.99% of C. sakazakii cells suspended in milk. The same level of C. sakazakii inactivation can be achieved with 1.8 min of a MTS treatment (60 °C; 200 kPa; 117 µm). Damaged cells were detected after heat treatments and after ultrasound under pressure treatments at lethal but not at non-lethal temperatures.

Environmental factors influencing the inactivation of Cronobacter sakazakii by high hydrostatic pressure.

The effect of High Hydrostatic Pressure (HHP) on the survival of Cronobacter sakazakii was investigated. Deviations from linearity were found on the survival curves and the Mafart equation accurately described the kinetics of inactivation. Comparisons between strains and treatments were made based on the time needed for a 5-log(10) reduction in viable count. The ability of C. sakazakii to tolerate high pressure was strain-dependent with a 26-fold difference in resistance among four strains tested. Pressure resistance was greatest in the stationary growth phase and at the highest growth temperatures tested (30 and 37 °C). Cells treated in neutral pH buffer were 5-fold more resistant than those treated at pH 4.0, and 8-fold more sensitive than those treated in buffer with sucrose added (a(w)=0.98). Pressure resistance data obtained in buffer at the appropriate pH adequately estimated the resistance of C. sakazakii in chicken and vegetables soups. In contrast, a significant protective effect against high pressure was conferred by rehydrated powdered milk. As expected, treatment efficacy improved as pressure increased. z values of 112, 136 and 156 MPa were obtained for pH 4.0, pH 7.0 and a(w)=0.98 buffers, respectively. Cells with sublethal injury to their outer and cytoplasmic membranes were detected after HHP under all the conditions tested. The lower resistance of C. sakazakii cells when treated in media of pH 4.0 seemed to be due to a decreased barostability of the bacterial envelopes. Conversely, the higher resistance displayed in media of reduced water activity may relate to a higher stability of bacterial envelopes.

Inactivation of Cronobacter sakazakii by ultrasonic waves under pressure in buffer and foods.

The objective of this research was to characterize the resistance of Cronobacter sakazakii to ultrasonic waves under pressure (manosonication, MS). The D(MS) value (decimal reduction time value) of C. sakazakii in standard conditions (35°C, 117 µm, 200 kPa, citrate-phosphate buffer pH 7.0) was 0.41 min. This value was higher than that of Yersinia enterocolitica (D(MS)=0.19 min) and lower than those of Salmonella enterica serovar Enteritidis (D(MS)=0.61 min), Listeria monocytogenes (D(MS)=0.86 min), and Enterococcus faecium (D(MS)=1.2 min). Strain studied (ATCC 29544, NCTC 8155, 9238, and 9529), growth temperature (10, 20, 30, and 37°C), and pH of the treatment media (4.0, 5.0, 6.0, and 7.0) did not significantly change C. sakazakii MS resistance. Conversely, entry into stationary growth phase, decreasing water activity of the treatment media (0.98, 0.96, and 0.94), and treatment in food products (apple and orange juices, chicken and vegetable soups, and rehydrated powdered milk) resulted in up to a 1.6-, 3.9-, and 2.5-fold maximum change in D(MS) values, respectively. Whereas an exponential relationship between the amplitude of ultrasonic waves and D(MS) values was found, the relationship between static pressure and D(MS) values was better described by a quadratic equation. The energy transferred into the medium determined the lethality of the ultrasonic waves regardless of the combination of pressure (0, 50, 100, 200 and 300 kPa) and amplitude (34, 62, 90, 117 and 145 µm) applied. There was an exponential relationship between D(MS) values and the power input: an increase of 134 W increased the inactivation rate ten times regardless of the treatment medium. No C. sakazakii cells with sublethally injured cytoplasmic membrane or with sublethal oxidative damage occurred after MS treatments, but the results indicated that damage to the outer membrane preceded microbial death.

Pulsed electric fields cause sublethal injuries in the outer membrane of Enterobacter sakazakii facilitating the antimicrobial activity of citral.

AIMS: The objective was to evaluate the relation of sublethal injury in the outer membrane of Enterobacter sakazakii to the inactivating effect of the combination of pulsed electric fields (PEF) treatments and citral. METHODS AND RESULTS: The occurrence of sublethal injury in the outer membrane was measured using selective recovery media containing bile salts. Loss of membrane integrity was measured by the increased uptake of the fluorescent dye propidium iodide (PI). PEF caused nonpermanent and permanent envelope permeabilization of Ent. sakazakii at pH 4·0. After PEF, most surviving cells showed transient cell permeabilization and sublethal injury in their outer membranes. The simultaneous application of a mild PEF treatment (100 pulses, 25 kV cm(-1) ) and 200 µl l(-1) of citral to cells suspended in pH 4·0 buffer at a final concentration of 10(7) cells per ml showed an outstanding synergistic lethal effect, causing the inactivation of more than two extra log(10) cycles. CONCLUSIONS: Our results confirm that the detection of sublethal injury in the outer membrane after PEF may contribute to the identification of the treatment conditions under which PEF may act synergistically with hydrophobic compounds such as citral. SIGNIFICANCE AND IMPACT OF THE STUDY: Knowledge about the mechanism of microbial inactivation by PEF will aid the establishment of successful combined preservation treatments.

sigB absence decreased Listeria monocytogenes EGD-e heat resistance but not its Pulsed Electric Fields resistance.

This study shows the behaviour of Listeria monocytogenes under heat and Pulsed Electric Fields (PEF) treatments, as well as the influence of sigB in bacterial resistance and recovery. Absence of the sigB gene resulted in a decreased heat tolerance, showing that sigma(B) influences L. monocytogenes heat survival. Heat treatments at 60 degrees C (40s) caused sublethal membrane injuries in 99.99% of survivors. The repair of heat damage required energy production and lipid, protein, and RNA synthesis, and it lasted 6h. Furthermore, deletion of sigB did not affect the heat injuries repair. PEF resistance at pH 4.0 and 7.0 was not influenced by sigB. Sublethal damage after PEF treatments was only detected when PEF-treated cells had previously been heat-shocked (45 degrees C/1h). The membrane repair only required energy production, and it was independent of sigB. Although both heat and PEF treatments have an effect on cellular membrane, the repair of the sublethal damages suggests different membrane targets, and thus we propose a different mechanism of inactivation by these food preservation technologies.

Development of stress resistance in Staphylococcus aureus after exposure to sublethal environmental conditions.

The ability of Staphylococcus aureus to develop stress resistance responses was investigated. Exponential growth phase cells of S. aureus CECT 4459 were exposed to sublethal conditions (acid and alkaline pH, hydrogen peroxide, and heat) and then the acquisition of resistance to acid (pH 2.5), alkali (pH 12.0), hydrogen peroxide (50mM), and heat (58 degrees C) was determined. Conditions resulting in the maximum development of homologous resistance (tolerance to the same stress), while preventing lethal effects in the population, were pH 4.5 (2h), pH 9.5 (30 min), 0.05 mM H(2)O(2) (30 min), and 45 degrees C (2h). Under these adaptation conditions, times for the first decimal reduction (TFDC) to a lethal treatment at acid pH, alkaline pH, hydrogen peroxide, and heat were increased by a factor of 1.6, 2, 2, and 6, respectively. The presence of chloramphenicol or rifampicin in the adaptation medium completely abolished the increase in homologous resistance to acid pH and to hydrogen peroxide. By contrast, the development of homologous resistance to alkaline pH resulted independently of the presence of either chloramphenicol or rifampicin. S. aureus heat resistance increased in the presence of the inhibitors during the heat shock, but only partially. In some cases, the exposure to a given stress induced cross-protection against other agents. Protective combinations of sublethal stress and lethal agents were: acid pH-heat, acid pH-hydrogen peroxide, alkaline pH-hydrogen peroxide, heat-acid pH, and heat-hydrogen peroxide. These combinations of agents applied sequentially should be avoided in food-processing environments.

Effect of citral on the thermal inactivation of Escherichia coli O157:H7 in citrate phosphate buffer and apple juice.

Inactivation and sublethal injury of Escherichia coli O157:H7 cells induced by heat in citrate phosphate buffer and apple juice (both at pH 3.8) were studied, and the effect of a combined preservation treatment using citral and heat treatments was determined. Heat resistance of E. coli O157:H7 was similar in both treatment media; after 27 min at 54°C, 3 log units of the initial cell population was inactivated in both treatment media. However, under less harsh conditions a protective effect of apple juice was found. Whereas inactivation followed linear kinetics in the citrate phosphate buffer, when cells were treated in apple juice the survival curves were concave downward. Heat treatment caused a great degree of sublethal injury; 4 min at 54°C inactivated less than 0.5 log CFU/ml but sublethally injured more than 3 log CFU/ml. The addition of 18 and 200 ppm of citral to the treatment medium acted synergistically with heat at 54°C to inactivate 3 × 10(4) and 3 × 10(7) CFU/ml, respectively. Addition of citral thus reduced the time needed to inactivate 1 log unit of the initial E. coli O157:H7 population from 8.9 to 1.7 min. These results indicate that a combined process of heat and citral can inactivate E. coli O157:H7 cells and reduce their potential negative effects.

Organic acids make Escherichia coli more resistant to pulsed electric fields at acid pH.

Stationary growth phase cells of Escherichiacoli were more pulsed electric fields (PEF) resistant in citrate-phosphate McIlvaine buffer at pH 4.0 than at pH 7.0. The greater PEF resistance was also confirmed in fruit juices of similar acid pH. In this work we studied whether the higher PEF resistance of E. coli at acid pH was due to the low pH itself or to the interaction of the components of the treatment medium with the cells. The protective effect on E. coli cells was due to the presence of organic acids such as citric, acetic, lactic or malic at pH 4.0. The protective effect of citric acid at pH 4.0 depended on its concentration. A linear relationship was observed between the Log(10) of the citric acid concentration and the degree of inactivation. Organic acids contained in laboratory treatment media (citrate-phosphate buffer) or in fruit juices did not sensitize E. coli cells to PEF but, on the contrary, they induced a protective effect that made E. coli cells more resistant at pH 4.0 than at neutral pH. This work could be useful for improving food preservation by PEF technology and it contributes to the knowledge of the mechanism of microbial inactivation by PEF.

Inactivation of Escherichia coli by citral.

AIMS: The aim was to evaluate (i) the resistance of Escherichia coli BJ4 to citral in a buffer system as a function of citral concentration, treatment medium pH, storage time and initial inoculum size, (ii) the role of the sigma factor RpoS on citral resistance of E. coli, (iii) the role of the cell envelope damage in the mechanism of microbial inactivation by citral and (iiii) possible synergistic effects of mild heat treatment and pulsed electric fields (PEF) treatment combined with citral. METHODS AND RESULTS: The initial inoculum size greatly affected the efficacy of citral against E. coli cells. Exposure to 200 microl l(-1) of citral at pH 4.0 for 24 h at 20 degrees C caused the inactivation of more than 5 log(10) cycles of cells starting at an inoculum size of 10(6) or 10(7) CFU ml(-1), whereas increasing the cell concentration to 10(9) CFU ml(-1) caused <1 log(10) cycle of inactivation. Escherichia coli showed higher resistance to citral at pH 4.0 than pH 7.0. The rpoS null mutant strain E. coli BJ4L1 was less resistant to citral than the wild-type strain. Occurrence of sublethal injury to both the cytoplasmic and outer membranes was demonstrated by adding sodium chloride or bile salts to the recovery media. The majority of sublethally injured cells by citral required energy and lipid synthesis for repair. A strongly synergistic lethal effect was shown by mild heat treatment combined with citral but the presence of citral during the application of a PEF treatment did not show any advantage. CONCLUSIONS: This work confirms that cell envelope damage is an important event in citral inactivation of bacteria, and it describes the key factors on the inactivation of E. coli cells by citral. SIGNIFICANCE AND IMPACT OF THE STUDY: Knowledge about the mechanism of microbial inactivation by citral helps establish successful combined preservation treatments.

Thermobacteriological characterization of Enterobacter sakazakii.

In the present study the influence of various environmental and physiological factors on the heat resistance of Enterobacter sakazakii (Cronobacter) have been investigated. Our results demonstrated that the heat resistance of E. sakazakii depended on the strain studied, the growth conditions - phase and temperature - the characteristics of treatment medium and the recovery conditions. The strain STCC 858 (ATCC type strain 29544) showed maximum heat resistance among the strains tested and it was selected for the further study. Stationary-phase cells grown between 20 and 37 degrees C (mean D(60)=0.9 min) resulted to be more resistant than cells grown at 10 degrees C (D(60)=0.2 min). Resistance decreased when the treatment medium pH was lower than pH 6.0, and it increased with decreasing water activity of the treatment medium, with a 32-fold increase in resistance when lowering water activity to 0.96. z value at pH 4.0 (z=4.79 degrees C) was significantly higher than at pH 7.0 (z=4.06 degrees C), although E. sakazakii cells were approximately 10 times more heat resistant at pH 7.0 than at pH 4.0 within the range of temperatures tested. Contrary to pH, the magnitude of the influence of a(w) on heat resistance did not significantly change with treatment temperature. The proportion of sublethally damaged cells was similar regardless of the treatment medium pH, but it decreased when lowering the water activity. Nevertheless, increasing treatment temperature would not result in a decreased proportion of sublethally injured E. sakazakii cells within the surviving population. Thus, the design of a theoretical combined process that could take advantage of the occurrence of sublethally injured cells would be similarly effective at low and high temperatures. E. sakazakii proved to be more heat resistant in four different liquid food matrixes than in buffers at the same pH, and this disagreement was especially higher in orange juice, which resulted to be the product that induced a greater protective effect in E. sakazakii cells against heat.

Part-time work among older workers with disabilities in Europe.

OBJECTIVE: To analyse the use of part-time work among older workers with disabilities compared with their non-disabled counterparts within a European context. STUDY DESIGN: Cross-sectional. METHODS: Data were drawn from the 2004 Survey of Health, Ageing and Retirement in Europe. The key advantage of this dataset is that it provides a harmonized cross-national dimension, and contains information for European individuals aged 50 years or over on a wide range of health indicators, disability, socio-economic situation, social relations, etc. RESULTS: Older people with disabilities (aged 50-64 years) are more likely to have a part-time job compared with their non-disabled counterparts. Although there is an important employment gap between the two groups, many older workers with disabilities use part-time work to achieve a better balance between their health status and working life. The econometric analysis corroborated that being disabled has a positive effect on the probability of working on a part-time basis, although this effect varies by country. CONCLUSIONS: Policy makers must encourage part-time employment as a means of increasing employment opportunities for older workers with disabilities, and support gradual retirement opportunities with flexible and reduced working hours. It is crucial to change attitudes towards older people with disabilities in order to increase their labour participation and reduce their levels of poverty and marginalization.

Role of the alternative sigma factor sigma on Staphylococcus aureus resistance to stresses of relevance to food preservation.

AIMS: To examine the role of the alternative general stress sigma factor sigma(B) on the resistance of Staphylococcus aureus to stresses of relevance to food preservation, with special emphasis on emerging technologies such as pulsed electric fields (PEF) and high hydrostatic pressure (HHP). METHODS AND RESULTS: S. aureus strain Newman and its isogenic DeltasigB mutant were grown to exponential and stationary growth phases and its resistance to various stresses was tested. The absence of the sigma(B) factor caused a decrease in the resistance to heat, PEF, HHP, alkali, acid and hydrogen peroxide. In the case of heat, the influence of the sigma(B) factor was particularly important, and decreases in decimal reduction time values of ninefold were observed as a result of its deficiency. The increased thermotolerance of the parental strain as compared with the sigB mutant could be attributed to a better capacity to sustain and repair sublethal damages caused by heat. CONCLUSIONS: sigma(B) factor provides S. aureus cells with resistance to multiple stresses, increasing survival to heat, PEF and HHP treatments. SIGNIFICANCE AND IMPACT OF THE STUDY: Results obtained in this work help in understanding the physiological mechanisms behind cell survival and death in food-processing environments.

Recovery of Saccharomyces cerevisiae sublethally injured cells after Pulsed Electric Fields.

The objective was to investigate the influence of the recovery liquid medium on the repair of Saccharomyces cerevisiae sublethally injured cells after Pulsed Electric Fields (PEF) in media of different pH. Sublethal injury was detected in the yeast S. cerevisiae after 50 pulses at 12.0 kV cm(-1) in both pH 4.0 and 7.0, by using a selective medium plating technique. PEF treatments cause a repairable sublethal injury in S. cerevisiae. Injured cells showed their maximum repair capacity when suspended in Sabouraud Broth compared to Peptone Water or citrate-phosphate buffer of pH 4.0. The extent to which cells repair their injuries depended on the treatment medium pH, and on the nature of the storage medium. No repair was detected when the recovery liquid medium was citrate-phosphate buffer of pH 7.0. Acid conditions favour repair and survival of PEF-treated S. cerevisiae cells. This work contributes to the description of the mechanisms of PEF injury and inactivation, which would be useful in defining adequate PEF treatments, alone or in combination with additional hurdles, to assure food stability.