PROTEIN PROFILE CHANGES IN ACID ADAPTED LISTERIA MONOCYTOGENES EXHIBITING CROSS-PROTECTION AGAINST AN ACTIVATED LACTOPEROXIDASE SYSTEM IN TRYPTIC SOY BROTH

Foodborne pathogens often tolerate and survive environmental stress conditions including extreme acidity to varying degrees. One possible reason for this survival may be the production of protective stress proteins during acid shock (ASR) and/or tolerance (ATR) responses. The ASR and ATR of Listeria monocytogenes strains V7, V37 and CA in tryptic soy broth without dextrose acidified with lactic acid were studied. Possible cross-protection of acid adapted cells against an activated lactoperoxidase system was also determined. The strains were either directly challenged at pH 4.0 and 3.5 to study their ASR or initially adapted at pH 5.5 for the equivalent of 1 generation before challenging at pH 4.0 and 3.5 to study their ATR. Adapted and nonadapted cells were challenged at pH 4.5 with or without an activated lactoperoxidase system. In all cases viability was determined by enumeration over a period of 24 or 48 h after challenge and the production of stress proteins analyzed by 2-dimensional gel electrophoresis. While there were some differences in the survival responses for each strain, the acid adapted cells of each strain survived to a greater degree than nonadapted cells at both pH 4.0 (at least 10 fold at 24 h) and pH 3.5 (at least 1000 fold at 6 h) but not at pH 4.5. The acid adapted cells exposed to the lactoperoxidase system survived better (at least 5-fold) than their nonadapted counterparts for all 3 strains at 24 and 48 h. The 2-dimensional gel analysis for all 3 strains showed that the adapted and nonadapted cells underwent a change in their physiology, (at pH 4.0 compared to the control at pH 7.0; at pH 4.5 with the addition of lactoperoxidase system components) in that there was induction as well as repression of several proteins.     

Stress response of Listeria monocytogenes isolated from cheese and other foods

The responses to pH and sodium chloride of four strains of Listeria monocytogenes isolated from Portuguese cheese, with a sodium chloride concentration of about 2% (w/v) and a pH value from 5.1 to 6.2, were studied. Two isolates from meat and two clinical isolates related to food-borne listeriosis, in which the implicated food product had about 2-3.5% (w/v) sodium chloride, also were studied. The effect of temperature on pH and sodium chloride sensitivity was also determined. The results show that natural isolates vary in response to these stresses and the data were often at variance with previously published data. Strains varied in sensitivity to low pH and to high sodium chloride concentration but the cheese isolates tended to be more resistant. A lower temperature was associated with a decrease in resistance to low pH and to sodium chloride. All strains showed an acid tolerance response induction when grown at pH 5.5 and although the time required for maximum induction of the response varied between strains, 2 h of acid adaptation, at least, was necessary which is longer than previously reported. Some strains showed an osmotolerance response after incubation in 3.5% (w/v) sodium chloride. Osmoadaptation, in addition to inducing an osmotolerance response, also induced cross-protection against acid shock conditions (pH 3.5). The acid tolerance response also induced a cross-protection against osmotic shock conditions (20% (w/v) sodium chloride). In some cases there was a relationship between the degree of resistance and adaptation, but usually the behaviour of a particular strain was independent of the conditions from which it was isolated.     

Acid adaptation of Listeria monocytogenes strains does not offer cross-protection against an activated lactoperoxidase system.

Listeria monocytogenes has been implicated in foodborne illness outbreaks involving several types of cheeses made from acidified milk. Acid shock response (ASR) and acid tolerance response (ATR) could be possible reasons for its survival. The ASR and ATR of three strains of L. monocytogenes (V7, V37, and CA) in skim milk acidified to pH 4.0 and 3.5 with lactic acid and held at 32 degrees C were studied. Studies were also done to determine if acid adaptation of the organism enhanced survival in the presence of an activated lactoperoxidase system. The cells were directly shocked at pH 4.0 and 3.5 in skim milk to study the ASR. To study the ATR, cells were initially adapted in skim milk at a mild pH of 5.5 for the equivalent of one generation before being shocked at pH 4.0 and 3.5 in skim milk. Cells adapted at pH 5.5 in tryptic soy broth without dextrose and nonadapted cells were challenged at pH 4.5 in skim milk with or without an activated lactoperoxidase system. In all cases, viability and pH were measured 24 or 48 h after challenge. In pH 4.0 skim milk, for all three strains, the adapted cell population survived better (0.5 to 1.0 log higher) than that of nonadapted cells for 24 h. In pH 3.5 skim milk, the acid-adapted populations of all three strains were 3 to 4 logs greater than those of nonadapted cells at 6 h. The acid adapted cells of all three strains had survival rates similar to those of the nonadapted cells at pH 4.5 both in the presence and absence of an activated lactoperoxidase system. It was also evident that these strains do not exhibit an adaptive ATR at pH 4.5, although they do at lower pH levels (pH 4.0 and 3.5). Survival due to the ATR was better seen at pH 3.5 than at pH 4.0.     

Comparative analysis of acid resistance between susceptible and multi-antimicrobial-resistant Salmonella strains cultured under stationary-phase acid tolerance-inducing and noninducing conditions

This study compared acid resistance levels among five antimicrobial-susceptible strains of Salmonella and five strains that were simultaneously resistant to a minimum of six antimicrobial agents. The induction of a stationary-phase acid tolerance response (ATR) was attempted by both transient low-pH acid shock and acid adaptation. For acid shock induction, strains were grown for 18 h in minimal E medium containing 0.4% glucose (EG medium) and exposed to sublethal acid stress (pH 4.3) for 2 h, and subsequently, both shocked and nonshocked cultures were acid challenged (pH 3.0) for 4 h. Acid adaptation was achieved by growing strains for 18 h in tryptic soy broth containing 1.0% glucose (TSB+G), while nonadapted cultures were grown for 18 h in glucose-free tryptic soy broth (TSB-G). Acid-adapted and nonadapted inocula were acid challenged (pH 2.3) for 4 h. Initial (0 h) mean populations of nonchallenged Salmonella were 8.5 to 8.7, 8.4 to 8.8, and 8.2 to 8.3 log CFU/ml for strains grown in EG medium, TSB-G, and TSB+G, respectively. After 4 h of acid challenge, mean populations were 3.0 to 4.8 and 2.5 to 3.7 log CFU/ml for previously acid-shocked susceptible and resistant strains, respectively, while corresponding counts for nonshocked strains were 4.3 to 5.5 log CFU/ml and 3.9 to 4.9 log CFU/ml. Following 4 h of acid exposure, acid-adapted cultures of susceptible and resistant strains had mean populations of 6.1 to 6.4 log CFU/ml and 6.4 to 6.6 log CFU/ml, respectively, while corresponding counts for nonadapted cultures were 1.9 to 2.1 log CFU/ml and 1.8 to 2.0 log CFU/ml, respectively. A low-pH-inducible ATR was not achieved through transient acid shock, while an ATR was evident following acid adaptation, as adapted populations were 4.2 to 4.8 log units larger than nonadapted populations following acid exposure. Although some strain-dependent variations in acid resistance were observed, results from this study suggest no association between susceptibility to antimicrobial agents and the ability of the Salmonella strains evaluated to survive low-pH stress.     

Influence of acid adaptation on the tolerance of Escherichia coli O157:H7 to some subsequent stresses

Three stains of Escherichia coli O157:H7, including ATCC 43889, ATCC 43895, and 933, were first subjected to acid adaptation at a pH of 5.0 for 4 h. Thermal tolerance at 52 degrees C and survival of the acid-adapted as well as the nonadapted cells of E. coli O157:H7 in the presence of 10% sodium chloride, 0.85% bile salt, or 15.0% ethanol were investigated. Results showed that the effect of acid adaptation on the survival of E. coli O157:H7 varied with the strains and types of subsequent stress. Acid adaptation caused an increase in the thermal tolerance of E. coli O157:H7 ATCC 43889 and ATCC 43895, but no significant difference in the thermal tolerance was noted between acid-adapted and nonadapted cells of E. coli O157:H7 933. Although the magnitude of increase varied with strains of test organisms, acid adaptation generally led to an increase in the tolerance of E. coli O157:H7 to sodium chloride. On the other hand, the susceptibility of acid-adapted cells of the three strains of E. coli O157:H7 tested did not show a significant difference from that of their nonadapted counterparts when stressed with bile salt. The acid-adapted cells of E. coli O157:H7 ATCC 43889 and ATCC 43895 were less tolerant than the nonadapted cells to ethanol, whereas the tolerance of adapted and nonadapted cells of E. coli O157:H7 933 showed no significant differences.     

Influence of the sigB gene on the cold stress survival and subsequent recovery of two Listeria monocytogenes serotypes

The influence of serotype and the role of the sigB gene of Listeria monocytogenes during the survival and recovery on different substrates were determined. Wild-type and sigB mutants of two serotypes of L. monocytogenes were inoculated into buffer and onto beef steaks, and incubated at 4 degrees C for 8 weeks during which samples were removed and Listeria numbers determined. Growth kinetics of stationary phase wild-type and sigB mutant cells were compared, without prechilling and after prechilling at 4 degrees C. The two serotypes had similar survival capabilities under the conditions examined, and the sigB gene was influential in survival of chill stress, but was dependent upon additional nutritional factors. Prechilling cells prior to growth extended the lag phase of both strains, and this lag phase extension was compounded by the absence of a functional sigB gene. In conclusion, the sigB gene is involved in the survival and recovery from chill stress by the two serotypes tested. Additional factors such as previous growth conditions, nutritional requirements and serotype susceptibility are also contributory. This study adds relevant information regarding the influence of the sigB gene, in conjunction with the historical growth conditions and serotype differences. Understanding the significance of these factors may be useful in creating improved recovery systems for the detection of L. monocytogenes from at-risk foods.     

Survival and growth of alkali-stressed Listeria monocytogenes on beef frankfurters and thermotolerance in frankfurter exudates

Cells of Listeria monocytogenes exposed at 4 degrees C to 1% solutions of two alkaline cleaners or alkali-adapted in tryptose phosphate broth (pH 10.0) at 37 degrees C for 45 min, followed by 4 degrees C for 48 h, were inoculated onto beef frankfurters containing high fat (16 g) and high sodium (550 mg) or low fat (8 g) and low sodium (250 mg) per 57-g serving. Frankfurters were surface inoculated (2.0 log10 CFU/g), vacuum packaged, stored at -20, 4, or 12 degrees C, and analyzed for populations of L. monocytogenes at 2-day to 2-week intervals. Populations did not change significantly on frankfurters stored at -20 degrees C for up to 12 weeks. After storage at 4 degrees C for 6 weeks (I week before the end of shelf life), populations of control cells and cells exposed to alkaline cleaners were ca. 6.0 log10 CFU/g of low fat, low sodium (LFLS) frankfurters and ca. 3.5 log10 CFU/g of high fat, high sodium (HFHS) frankfurters. Growth of alkali-adapted cells on both types of frankfurters was retarded at 4 degrees C. Growth of L. monocytogenes on frankfurters stored at 12 degrees C was more rapid than at 4 degrees C, but a delay in growth of alkali-adapted cells on HFHS and LFLS frankfurters was evident during the first 9 and 6 days, respectively. Alkali-adapted cells had a significantly (P < or = 0.05) lower logistic D59 degrees C-value (decimal reduction time) than alkaline cleaner-exposed cells, but the D59 degrees C-value was not different from that of control cells. Cells exposed to a nonbutyl alkaline cleaner, and then heated in LFLS frankfurter exudates, had a significantly lower D62 degrees C-value than cells that had been exposed to some of the other treatments. Growth characteristics of L. monocytogenes inoculated onto the surface of frankfurters may be altered by previous exposure to alkaline environments. Differences in growth characteristics of L. monocytogenes on HFHS versus LFLS beef frankfurters stored at refrigeration temperatures indicate that composition influences the behavior of both alkaline-stressed and control cells.     

EFFECT OF ACID ADAPTATION AND DIFFERENT SALT CONCENTRATIONS ON SURVIVAL OF LISTERIA MONOCYTOGENES IN TURKISH WHITE CHEESE

ABSTRACT

Survival of acid‐adapted and nonacid adapted Listeria monocytogenes in Turkish white cheese with different salt concentrations (3.8 and 6.7% NaCl) was investigated. Cheese blocks were inoculated with L. monocytogenes and stored at 4C for 15 days. Samples were taken on different days and analyzed for the numbers of the pathogen, and the samples were challenged with simulated gastric fluid (SGF) for determining acid tolerance. There was no significant differences between the numbers of acid‐adapted and nonadapted L. monocytogenes cells regardless of the salt concentration on sampling days (P > 0.05). Challenge in SGF revealed that there was no significant difference between acid‐adapted and nonadapted bacteria in cheese with 3.8% NaCl throughout the storage period, while significant (P < 0.05) differences between those cells in cheese with 6.7% NaCl were found after day 5. These results suggest that acid‐adapted L. monocytogenes exposed to high salt concentration such as 6.7% may have an advantage in SGF.

PRACTICAL APPLICATIONS

Evaluation of potential stress responses of pathogens in food is important to understand the behavior of pathogens that can survive under stresses. As an environmental contaminant, Listeria monocytogenes may acquire adaptation to low acid conditions in cheese plants. Our results suggested that although there was no significant difference between acid‐adapted and nonadapted cells both in industrial (3.8% salt) and traditional (6.7% salt) white cheese, salt concentration significantly affected the survival of acid‐adapted cells in simulated gastric fluid (pH 2.3). These findings may indicate that L. monocytogenes, which is adapted to acid before contaminating high salt cheeses, may survive in stomach and cause infection. It can be said that increased salt concentration in cheese may not provide a barrier against acid‐adapted L. monocytogenes.

     

西藏牦牛奶渣中优势乳酸菌产胞外多糖及其耐受性

从分离自西藏牦牛奶渣的113株乳酸菌中筛选出4株优势菌株(GY-L003、GY-L004、GY-L055、GYL112),对其产胞外多糖(exopolysaccharide,EPS)能力、耐酸能力、耐胆酸盐能力、耐渗透压能力及抑菌能力进行了比较研究。结果表明:4株乳酸菌产EPS能力不尽相同,其中性能最优的菌株为GY-L003,为379.94 mg/L;4株菌株表现出不同的耐受能力,其中GY-L003和GY-L004在pH 1.5条件下仍能存活;当胆酸盐质量分数高于0.4%时,GY-L003和GY-L004亦能生存,GY-L055和GY-L112则受到抑制;在耐渗透压方面,GY-L003和GY-L004性能较优,9%Na Cl溶液中仍能生长;4株菌均具较好的抑菌特性,对金黄色葡萄球菌抑制最强的为GY-L004,抑菌圈可达(15.94±0.31)mm;GY-L003对大肠杆菌和沙门氏菌的抑制效能最强,抑菌圈分别为(18.21±0.65)、(16.27±0.50)mm;GY-L112对单增李斯特菌抑菌效果佳,抑菌圈为(15.24±0.35)mm。综上,GY-L003和GYL004产EPS、耐受性、抑菌能力等性能俱佳,具有较好的应用潜力,为工业化利用提供了参考依据。     

酿酒酵母工业菌株胁迫条件耐受性分析

对酿酒酵母（Saccharomyces cerevisiae）工业菌株胁迫条件，包括高浓度酒精、高渗透压、高温、营养饥饿、氧化胁迫、糠醛毒性的耐受性进行了分析，同时测定了抗生素G418对这些菌株的最低抑菌浓度。结果表明，所测定的酵母菌株对这些逆境条件的耐受性有明显差别，表现出良好耐受性的是6508和安琪酵母菌株，同时多倍性的酿酒酵母工业菌株的耐受性均比单倍性实验室菌株高。     

The Acetic Acid Tolerance Response induces cross-protection to salt stress in Salmonella typhimurium

Salmonella typhimurium induces an Acid Tolerance Response (ATR) upon exposure to mildly acidic conditions in order to protect itself against severe acid shock. This response can also induce cross-protection to other stresses such as heat and salt. We investigated whether both the acetic acid induced and lactic acid induced ATR in S. typhimurium provided cross-protection to a salt stress at 20 degrees C. Acid-adapted cells were challenged with both a sodium chloride (NaCl) and potassium chloride (KCl) shock and their ability to survive ascertained. Acetic acid adaptation provided cells with protection against both NaCl and KCl stress. However, lactic acid adaptation did not protect against either osmotic stressor and rendered cells hypersensitive to NaCl. These results have implications for the food industry where hurdle technology means multiple sub-lethal stresses such as mild pH and low salt are commonly used in the preservation of products.     

NaClO胁迫下单核细胞增生李斯特菌WaX12及其sigB基因缺失突变株抗氧化胁迫相关功能基因表达比较

目的:探究单核细胞增生李斯特菌sigB基因对次氯酸钠(NaClO)胁迫的调控作用。方法:在半致死浓度NaClO(3 mmol/L)胁迫下,比较该菌野生株WaX12与sigB基因敲除突变株的耐受程度及菌内的活性氧水平,并通过实时定量聚合酶链式反应技术(real-time quantitative polymerase chain reaction,RT-PCR)进一步观察lmo1433、lmo0906、lmo2344、ohrR、lmo2770基因随胁迫时间的转录水平变化。结果:野生株和sigB突变株的表型测定结果显示,野生株比缺失株更耐NaClO胁迫,而且此现象在稳定期更加明显;当胁迫时间达到30 min时,突变株内的活性氧水平增长率比野生株高出19.97%,进一步说明sigB基因对NaClO胁迫有一定的调控作用。RT-PCR结果表明,基因lmo1433、lmo2344和ohrR在野生株的表达水平显著高于突变株(P0.01),而基因lmo0906、lmo2770的表达水平在野生株和缺失株中的差异并不明显。结论:研究表明,单核细胞增生李斯特菌sigB基因对NaClO胁迫有一定的影响作用,且部分作用可能由激活部分调节机体氧化还原平衡的基因得以实现。     

Acid-adapted Listeria monocytogenes displays enhanced tolerance against the lantibiotics nisin and lacticin 3147.

Log-phase Listeria monocytogenes cells become tolerant to a variety of environmental stresses following acid adaptation at pH 5.5. We demonstrated that adapted cells also exhibit increased tolerance to nisin and, to a lesser extent, lacticin 3147. At nisin concentrations of 100 and 200 IU/ml the survival of acid-adapted cells was approximately 10-fold greater than nonadapted cells. However, acid adaptation had only a moderate effect on the tolerance of L. monocytogenes to lacticin 3147, a phenomenon that possibly reflects the distinct mode of action of this bacteriocin. Analysis of the fatty acid composition of the bacterial membrane indicated that straight-chain fatty acids C14:0 and C16:0 were significantly increased in acid-adapted cells while levels of C18:0 decreased. The results indicate that stress mechanisms that are induced in mildly acidic conditions provide protection against the antimicrobial action of bacteriocins. This increased resistance of acid-adapted L. monocytogenes could cause increased survival of this pathogen in food products in which nisin or other bacteriocins are used as preservatives.     

Heat stress adaptation induces cross-protection against lethal acid stress conditions in Arcobacter butzleri but not in Campylobacter jejuni

The ability of many bacteria to adapt to stressful conditions may later protect them against the same type of stress (specific adaptive response) or different types of stresses (multiple adaptive response, also termed cross-protection). Arcobacter butzleri and Campylobacter jejuni are close phylogenetic relatives that occur in many foods of animal origin and have been linked with human illness (mainly diarrhoea). In the present study, sublethal stress adaptation temperatures (48 °C and 10 °C) and mild and lethal acid conditions (pH 5.0 and pH 4.0) were determined for A. butzleri and C. jejuni. In addition, it was evaluated whether these sublethal stress adaptations cause specific adaptive responses or cross-protection against subsequent mild or lethal acid stresses in these bacteria. The studies were conducted in broth adjusted to the different conditions and the results were determined by the dilution series plating method. It was shown that heat stress adapted A. butzleri (incubated for 2 h at 48 °C) were significantly more resistant to subsequent lethal acid stress (pH 4.0) than non-adapted cells at the 1 h time-point (p < 0.01 in Wilcoxon rank sum test). No specific adaptive responses against the stresses in A. butzleri or C. jejuni and no cross-protection in C. jejuni were found. The ability of heat stressed A. butzleri to tolerate later lethal acid conditions should be taken into account when designing new food decontamination and processing strategies.     

Survival of osmotic and acid stress by Listeria monocytogenes strains of clinical or meat origin

The ability of 30 Listeria monocytogenes strains, 15 of meat origin and 15 of clinical origin, to use carnitine as an osmoprotectant and to resist acid stress was determined. All strains examined were able to use carnitine as an osmoprotectant, indicating the importance of this characteristic to the survival of L. monocytogenes in natural environments. Clinical and meat strains, however, differed with respect to this characteristic. Specifically, 73% of meat strains reached a lower maximum cell density in the presence of carnitine with osmotic stress than in its absence with no stress. Only 33% of clinical strains displayed the same feature whereas the remaining clinical strains reached a higher maximum cell density in the presence of carnitine with osmotic stress than in its absence with no stress. The physiological reasons and advantage of this difference are unclear. When exposed to conditions of severe acid stress (pH 2.5) for 2 h, only two L. monocytogenes strains (L66 and L78), both of meat origin, displayed significant reductions (P < 0.05) in number (3.51 and 2.79 log cfu, respectively). Acid-sensitive strains were not found among the clinical isolates examined, highlighting the importance of acid stress resistance in the infection process.     

Wine is Bactericidal to Foodborne Pathogens

ABSTRACT: Red and white wines without added sulfite were tested for antibacterial activity against stationary-phase grown cells of Escherichia coli O157:H7, Listeria monocytogenes, Salmonella Typhimurium, and Staphylococcus aureus . The wines had bactericidal activity against all strains, with the red wine being most potent. S . Typhimurium was most sensitive, with 6 log reduction after 10 min exposure to wine, whereas S. aureus appeared least sensitive to the wines. Mutants having the gene encoding the alternative sigma factor disrupted were generally more sensitive to wine than their wild-type counterparts. When different combinations of ethanol, organic acids, and acidity were tested against the pathogens, it was found that a composition of 0.15% malic acid, 0.6% tartaric acid, 15% ethanol, and pH 3.0 had a strong bactericidal effect. The compounds in the mixture seemed to act synergistically against the pathogens. The pathogens grew in 25% to 40% white wine diluted in brain hearth infusion broth, with S. aureus being able to grow at the highest concentration of wine. Preincubation of the bacteria in sublethal concentrations of wine and ethanol and pH 4.5 did not increase their tolerance against wine or against the mixture of organic acids and ethanol. In conclusion, wine had an antibacterial effect against the pathogens tested. The synergistic effect of organic acids, ethanol, and low pH seems to be responsible for a major part of the antibacterial effect of wine. The alternative sigma factors seemed to be involved in protection of the bacteria against wine.     

Acid adaptation and temperature effect on the survival of E. coli O157:H7 in acidic fruit juice and lactic fermented milk product.

In this study, two strains of Escherichia coli O157:H7, (ATCC 43889 and ATCC 43895) were acid adapted at pH 5.0 in tryptic soy broth (TSB) for 4 h. Commercial products of mango juice (pH 3.2), asparagus juice (pH 3.6), Yakult--a diluted milk fermented drink (pH 3.6), and low-fat yoghurt (pH 3.9) were inoculated with acid-adapted or nonadapted cells of E. coli O157:H7. Survival of the inoculated E. coli O157:H7 in these commercial food products during storage at 25 or 7 degrees C was examined. It was found that although survival of the acid-adapted and nonadapted E. coli O157:H7 ATCC 43895 in asparagus juice during storage at 7 degrees C did not show marked difference, in general, acid adaptation and low temperature enhanced the survival of E. coli O157:H7 in both the commercial fruit juices tested. On the contrary, acid adaptation reduced the survival of both the strains of the test organism in Yakult and low-fat yoghurt stored at 7 degrees C. Besides, E. coli O157:H7 ATCC 43895 survived longer than ATCC 43889 in all the products examined, regardless of the storage temperature and acid adaptation.     

Combined effects of NaCl, NaOH, and biocides (monolaurin or lauric acid) on inactivation of Listeria monocytogenes and Pseudomonas spp

This study highlighted combinations of chemical stresses that could decrease or eliminate Listeria monocytogenes and Pseudomonas spp. surviving in food processing plants. Strains of L. monocytogenes, Pseudomonas fragi, and Pseudomonas fluorescens isolated from processing environments (meat and milk) were grown at 20 degrees C up to the early stationary phase. The strains were then subjected to 30 min of physicochemical treatments. These treatments included individual or combined acid (acetic acid), alkaline (NaOH), osmotic (NaCl), and biocides (fatty acids) challenges. Survival of the strains was studied after individual or combined acid (acetic acid), alkaline (NaOH), osmotic (NaCl), and biocides (monolaurin, lauric acid) challenges. Individual pH shocks had lower efficiencies than those used in combinations with other parameters. The treatment pH 5.4 followed by pH 10.5 had a low efficiency against L. monocytogenes. The opposite combination, pH 10.5 followed by pH 5.4, led to a 3-log reduction of the L. monocytogenes population. Pseudomonas spp. strains were much more sensitive than L. monocytogenes, and population reductions of 5 and 8 log (total destruction), respectively, were observed after the same treatments. As for L. monocytogenes, the combination pH 10.5 followed by pH 5.4 is more deleterious than the opposite. Whatever the bacterial species, the most efficient treatments were combinations of alkaline, osmotic, and biocide shocks. For instance, the combination pH 10.5 and 10% NaCl plus biocides showed reductions of 5 to 8 log for both bacteria. The origins of the observed lethal effects are discussed.