Reduced Susceptibility of Salmonella Typhimurium Strains to Oregano Essential Oil and Enrofloxacin: An In Vitro Assay.

Several European animal nutrition companies have incorporated essential oils (EOs) into animal feed as a result of the prohibition of antibiotics to promote animal growth. Previous studies of EOs have highlighted the absence of bacterial resistance for these substances, although most of the published works focus on studying their tolerance to subinhibitory doses. For this study, oregano essential oil (OEO) was chosen for its proven inhibitory and bactericidal activity. This study is an in vitro assay of the possible induction of Salmonella enterica serovar Typhimurium strains with reduced susceptibility to OEO by mutation, seeking to calculate the mutant prevention concentration (MPC) since this is an important measurement for the control Salmonella's resistance to fluoroquinolones such as enrofloxacin (ENR), the treatment of choice for this infection. To establish the MPC, we used a bacterial inoculum ≥109 colony-forming unit (CFU)/mL and examined the bases for points of resistance to ENR and mutations of target genes of the quinolone resistance determining region (QRDR). The three strains of Salmonella Typhimurium used in this study showed an MPC of four times the minimum inhibitory concentration (MIC) for ENR. In all cases, strains with reduced susceptibility to ENR were obtained, although none reached the point of resistance. The QRDR characterization region was in all cases of wild type (wt). Two of the strains tested with OEO grew at a concentration of 1 × MIC, which could be strains with reduced susceptibility, associated with mutation or not. In this case, the MPC was 2 × MIC. Once isolated and identified as Salmonella Typhimurium, the MIC against OEO of all strains obtained in the induction test indicated a possible reduction in susceptibility. However, the result obtained for both strains coincided with MIC of the original strains, rejecting a priori such a reduced susceptibility of Salmonella Typhimurium to OEO.

Transcriptional profiling of Salmonella enterica serovar Enteritidis exposed to ethanolic extract of organic cranberry pomace.

Non-typhoidal Salmonella enterica serovars continue to be an important food safety issue worldwide. Cranberry (Vaccinium macrocarpon Ait) fruits possess antimicrobial properties due to their various acids and phenolic compounds; however, the underlying mechanism of actions is poorly understood. We evaluated the effects of cranberry extracts on the growth rate of Salmonella enterica serovars Typhimurium, Enteritidis and Heidelberg and on the transcriptomic profile of Salmonella Enteritidis to gain insight into phenotypic and transcriptional changes induced by cranberry extracts on this pathogen. An ethanolic extract from cranberry pomaces (KCOH) and two of its sub-fractions, anthocyanins (CRFa20) and non-anthocyanin polyphenols (CRFp85), were used. The minimum inhibitory (MICs) and bactericidal (MBCs) concentrations of these fractions against tested pathogens were obtained using the broth micro-dilution method according to the Clinical Laboratory Standard Institute's guidelines. Transcriptional profiles of S. Enteritidis grown in cation-adjusted Mueller-Hinton broth supplemented with or without 2 or 4 mg/ml of KCOH were compared by RNASeq to reveal gene modulations serving as markers for biological activity. The MIC and MBC values of KCOH were 8 and 16 mg/mL, respectively, against all tested S. enterica isolates. The MIC value was 4 mg/mL for both CRFa20 and CRFp85 sub-fractions, and a reduced MBC value was obtained for CRFp85 (4 mg/ml). Treatment of S. Enteritidis with KCOH revealed a concentration-dependent transcriptional signature. Compared to the control, 2 mg/ml of KCOH exposure resulted in 89 differentially expressed genes (DEGs), of which 53 and 36 were downregulated and upregulated, respectively. The upregulated genes included those involved in citrate metabolism, enterobactin synthesis and transport, and virulence. Exposure to 4 mg/ml KCOH led to the modulated expression of 376 genes, of which 233 were downregulated and 143 upregulated, which is 4.2 times more DEGs than from exposure to 2 mg/ml KCOH. The downregulated genes were related to flagellar motility, Salmonella Pathogenicity Island-1 (SPI-1), cell wall/membrane biogenesis, and transcription. Moreover, genes involved in energy production and conversion, carbohydrate transport and metabolism, and coenzyme transport and metabolism were upregulated during exposure to 4 mg/ml KCOH. Overall, 57 genes were differentially expressed (48 downregulated and 9 upregulated) in response to both concentrations. Both concentrations of KCOH downregulated expression of hilA, which is a major SPI-1 transcriptional regulator. This study provides information on the response of Salmonella exposed to cranberry extracts, which could be used in the control of this important foodborne pathogen.

Efficacy of plant-derived antimicrobials for controlling Salmonella Schwarzengrund on dry pet food.

Salmonella enterica is a major human pathogen that is responsible for 23,000 hospitalizations annually in the United States. Contact with contaminated pet food and infected companion animals can transmit salmonellosis to humans. Recent multistate human outbreaks of salmonellosis linked to commercial contaminated dry dog foods underscore the need for controlling the pathogen in pet foods for protecting pet and public health. In this study, the efficacy of five Generally Recognized as Safe (GRAS) status, plant-derived antimicrobials (PDAs), namely trans-cinnamaldehyde (TC), carvacrol (CR), thymol (TY), eugenol (EG), and caprylic acid (CA) applied as a vegetable oil or chitosan based antimicrobial spray on dry pet food for reducing Salmonella Schwarzengrund was investigated. Three hundred gram portions of a commercial dry dog food were inoculated with a two-strain mixture of nalidixic acid (NA) resistant S. Schwarzengrund (~6 log CFU/g), followed by a spray treatment with 0%, 0.5%, 1% or 2% of TC, CR, TY, EG or CA in combination with 5% vegetable oil or 1% chitosan as a carrier. The control and treated dog food samples were stored at 25 °C for 28 days. On days 0, 1, 3, 5, 7, 14, 21, and 28, Salmonella on pet food was enumerated by serial dilution and plating on xylose lysine desoxycholate (XLD) agar. All PDAs at 1% and 2% applied in vegetable oil or chitosan reduced S. Schwarzengrund by at least ~2 log CFU/g on day 3 of storage when compared to control (P < 0.05). No significant reductions in Salmonella were observed on feed sprayed with only vegetable oil or chitosan (P > 0.05). Overall, 2% TC in vegetable oil or chitosan was the most effective treatment, where at least 3 to 3.5 log CFU/g reduction in bacterial populations was observed during storage (P < 0.05). Results suggest that the aforementioned PDAs could potentially be used as an antimicrobial spray to reduce S. Schwarzengrund on dry dog food. However, further studies on the acceptance of PDA-treated dry food by dogs are needed.

Tolerance to benzalkonium chloride and antimicrobial activity of Butia odorata Barb. Rodr. extract in Salmonella spp. isolates from food and food environments.

Salmonellosis, caused by the consumption of contaminated foods, is a major health problem worldwide. The aims of this study were to assess the susceptibility of Salmonella spp. isolates to benzalkonium chloride (BC) disinfectant and the antimicrobial activity of Butia odorata Barb. Rodr. extract against the same isolates from food and food environments. Moreover, phenotypic and genotypic resistance profiles, the presence of virulence genes and biofilm forming ability were determined. The minimum inhibitory concentration (MIC) of B. odorata extract against Salmonella spp. ranged from 10 to >19 mg.mL-1. Resistance to ampicillin, streptomycin, nalidixic acid, sulfonamide, trimethoprim/sulfamethoxazole, trimethoprim, tetracycline, and chloramphenicol was observed. In addition, multidrug resistance was observed in seven isolates (26.92%). The MIC of BC ranged from 32 to 64 mg.L-1, higher concentrations in comparison with wild-type MICs, and therefore were considered tolerant. Several resistance genes were detected, of which the most common were aadA, qacEΔ1, blaTEM, int1, sul1, and tetA. All isolates carried at least one virulence gene and produced biofilms on stainless steel surfaces at 10 and 22 °C. On the other hand, the B. odorata extract showed activity against Salmonella spp., and it has the potential to be used as a natural antimicrobial to control this important foodborne pathogen, despite its virulence potential and antimicrobial resistance profile.

In-package atmospheric cold plasma treatment of bulk grape tomatoes for microbiological safety and preservation.

Effects of dielectric barrier discharge atmospheric cold plasma (DACP) treatment on the inactivation of Salmonella and the storability of grape tomato were investigated. Grape tomatoes, with or without inoculation with Salmonella, were packaged in a polyethylene terephthalate (PET) commercial clamshell container and cold plasma-treated at 35 kV at 1.1 A for 3 min using a DACP system equipped with a pin-type high-voltage electrode. DACP treatment inactivated Salmonella (p < 0.05) without altering the color or firmness of the grape tomatoes (p > 0.05). DACP treatment inactivated Salmonella uniformly in both layers of the double-layer configuration of the grape tomatoes regardless of the position of the tomatoes in each layer. Salmonella was most efficiently inactivated when the headspace to tomato volume ratio of the container was highest. Integration of rolling of tomatoes during treatment significantly increased the Salmonella reduction rates from 0.9 ±â€¯0.2 log CFU/tomato to 3.3 ±â€¯0.5 log CFU/tomato in the double-layer configuration of the tomato samples. Rolling-integrated DACP also initially reduced the number of total mesophilic aerobic bacteria and yeast and molds in the double-layer configuration of tomato samples by 1.3 ±â€¯0.3 and 1.5 ±â€¯0.2 log CFU/tomato, respectively. DACP treatment effectively reduced the growth of Salmonella and indigenous microorganisms at 10 and 25 °C, and did not influence the surface color, firmness, weight loss, lycopene concentration and residual ascorbic acid of grape tomatoes during storage at 10 and 25 °C. DACP treatment holds promise as a post-packaging process for improving microbial safety against Salmonella and storability of fresh grape tomatoes.

Citrus fruit extracts with carvacrol and thymol eliminated 7-log acid-adapted Escherichia coli O157:H7, Salmonella typhimurium, and Listeria monocytogenes: A potential of effective natural antibacterial agents.

Despite the widespread belief that citrus fruit extracts (CFEs) are microbiologically safe due to their acidity, limited bactericidal effect results in low applicability as antibacterial agent and outbreaks occurred by acid-adapted pathogens. Here, we examined the antibacterial effects of CFEs [lime (Citrus medica), lemon (Citrus limon), calamansi (Citrus microcarpa)] combined with essential oil components (EOCs; carvacrol and thymol) against non-acid-adapted/acid-adapted Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes under 22 °C for 5 min. CFEs (<20%) alone or small amounts of EOCs (2.0 mM; 0.032%) alone could not inactivate the target bacteria effectively. However, combined treatments exhibited marked synergy: CFE + EOCs eliminated all the bacteria (>6.9 log CFU/ml). Among the CFEs tested, the highest synergism was shown by calamansi, an exotic citrus fruit previously unrecognized as an antibacterial agent. Although acid-adaptation improved bacterial survival, calamansi (<20%) + EOCs (<0.032%) completely inactivated even the most resistant pathogen (E. coli O157:H7). Validation test also showed that all tested commercial juice products also eliminated acid-adapted pathogens when used with EOCs. Physicochemical analysis of tested CFEs (pH measurement and HPLC analysis of components) revealed that low pH and flavanone (hesperidin) did not contribute to the synergistic bactericidal effects. Rather, the high citric acid content is likely to contribute to the strong synergistic effect with EOCs by damaging susceptible bacterial membranes. Sensory scores for CFEs were not altered by addition of EOCs at concentrations up to 1.5 mM. This study provides new insight into the utility of CFEs with EOCs to improve not only the microbiological safety of food products containing CFEs but also their applicability as natural antibacterial complex.

Effect of overhead spray and brush roller treatment on the survival of Pectobacterium and Salmonella on tomato surfaces.

Overhead spray and brush roller (OSBR) treatment has been shown to remove significantly more Salmonella from tomato surfaces than flume treatment. However, OSBR is not widely used in tomato packing facilities compared with other commodities, and little is known about whether brushing causes microabrasions or other physical damage. Bacteria such as Pectobacterium, a soft rot-producing plant pathogen, and Salmonella, a human pathogen, show increased survival and growth on damaged tomato surfaces. This study evaluated whether OSBR treatment had a negative effect on the safety and/or marketability of tomatoes by examining its effect on Pectobacterium and Salmonella survival. Pectobacterium survival was evaluated on inoculated tomatoes that were OSBR treated with water or sanitizer (100 ppm of NaOCl, 5 ppm of ClO2, or 80 ppm of peracetic acid). A 15-s OSBR treatment using water or sanitizer achieved a 3-log CFU/ml reduction in Pectobacterium levels. Survival of Pectobacterium and Salmonella on OSBR-treated, untreated, and puncture-wounded tomatoes stored at 25°C and 75 to 85 % relative humidity for 7 days was also assessed. Both Pectobacterium and Salmonella populations declined rapidly on OSBR-treated and untreated tomatoes, indicating that brushing does not damage tomato fruit to the extent of promoting better pathogen survival. In contrast, the survival of both organisms was significantly (P ≤ 0.05) higher on artificially wounded fruit. These results indicate that OSBR treatment does not increase the survival and growth of Pectobacterium or Salmonella on tomato surfaces and that it is effective in reducing Pectobacterium levels on the surface of inoculated tomatoes. These results suggest that, if used properly, an OSBR system in packinghouses is effective in removing surface contamination and does not affect tomato quality or safety.

Effect of white mustard essential oil on inoculated Salmonella sp. in a sauce with particulates.

White mustard essential oil (WMEO), from white mustard seed (Sinapis alba L.), is obtained by solvent extraction of defatted and wetted ground mustard; endogenous myrosinase catalyzes the hydrolysis of the glucosinolate sinalbin to yield 4-hydroxybenzyl isothiocyanate (4-HBITC), the antimicrobial component of WMEO. Sauce with particulates was made by mixing sauce, which served as the carrier for WMEO, with frozen vegetable and chicken particulates inoculated with Salmonella sp. WMEO (at 250 to 750 ppm of 4-HBITC) was able to reduce inoculated Salmonella counts by 0.8 to 2.7 log (CFU/g) in a frozen sauce with particulates in a dose-dependent manner, starting from the point of formulating the sauce through the microwave cooking step. High-pressure liquid chromatography-based analytical data confirmed that 4-HBITC was present in all of the samples in the expected concentrations and was completely hydrolyzed after the recommended cooking time in microwave ovens. In another experiment simulating unintentional abuse conditions, where the WMEO containing sauce with particulates was kept at room temperature for 5 h, WMEO (at 250 to 750 ppm of 4-HBITC) was able to reduce inoculated Salmonella counts from the point of first contact and up to 5 h by 0.7 to 2.4 log (CFU/g). Despite the known hydrolytic instability of the active component 4-HBITC, particularly at close to neutral pH values, WMEO was effective in controlling deliberately inoculated Salmonella sp. in a frozen sauce with particulates.

Impact of storage time and temperature on thermal inactivation of Salmonella Enteritidis PT 30 on oil-roasted almonds.

UNLABELLED: Whole Nonpareil variety almonds were inoculated with Salmonella Enteritidis PT 30 and stored at 4 or 23 °C for up to 48 wk. At 1, 12, 24, 37, and 48 wk of storage, almonds were heated by immersion in 121 °C oil. After heating for 0.5 to 2.5 min, almonds were drained, transferred to tryptic soy broth, and mixed with a stomacher prior to plating onto tryptic soy and bismuth sulfite agars. Over the 48 wk of storage, Salmonella declined by 0.5 and 2.1 log CFU/g at 4 and 23 °C, respectively. The survivor inactivation curves were upwardly concave with rapid initial reductions in the levels of Salmonella. For up to 24 wk of storage, the mean counts of the survivors after treatment were not significantly different. The Weibull model predicted 4- and 5-log reductions of Salmonella in 0.85 ± 0.16 and 1.8 ± 0.43 min, respectively, for almonds stored at 4 °C, and in 1.6 ± 0.53 and 3.2 ± 1.0 min, respectively, for almonds stored at 23 °C. Refrigerated storage had little impact on heat resistance of Salmonella that were inoculated on almonds. PRACTICAL APPLICATION: This research provides information of value in performing or evaluating validation studies for thermally processed almonds. The sensitivity of Salmonella to oil roasting is demonstrated during typical commercial almond storage times and temperatures.

Serovariedades y patrones de susceptibilidad a los antimicrobianos de cepas de Salmonella aisladas de alimentos en Cuba / Serotypes and antimicrobial susceptibility patterns of Salmonella strains isolated from food in Cuba

The serotypes of 178 isolates of Salmonella enterica taken from food in different regions of Cuba between January 2008 and December 2009 were identified, and the antimicrobial susceptibility pattern of 100 selected isolates was determined by stratasampling. A total of 20 Salmonella serotypes were identified, with a predominance of S. Enteritidis (23%), S. Agona (13.5%), and S. London (11.2%). Of all the strains, 75%were resistant or presented intermediate resistance to at least one of the drugs tested, in the following order: tetracycline (70.7%), ampicillin (22.7%), and nalidixic acid (14.7%). Ten different resistance patterns were identified. The most frequent patternscorresponded to strains that were either drug-resistant or had intermediate resistance (89.3%). Three strains (identified as S. Infantis, S. Derby, and S. Enteritidis) were multiresistant, and one of them, S. Enteritidis, was not sensitive to either nalidixic acid or ciprofloxacin. To control salmonellosis, the importance of maximizing integrated health surveillance is emphasized.

Se describen los serotipos de 178 cepas de Salmonella enterica aisladas de alimentos en diferentes regiones de Cuba entre enero de 2008 y diciembre de 2009, y el patrón de susceptibilidada los antimicrobianos de 100 aislados seleccionados mediante muestreo por estratos. Se identificaron 20 serovariedades de Salmonella entre las que predominaron S. Enteritidis (23%);S. Agona (13,5%) y S. London (11,2%). Del total, 75% de las cepas fueron resistentes o presentaronresistencia intermedia a al menos uno de los fármacos probados, en el siguiente orden, según su frecuencia: tetraciclina (70,7%); ampicilina (22,7%) y ácido nalidíxico (14,7%). Seidentificaron 10 patrones de resistencia diferentes y predominaron las cepas resistentes o con resistencia intermedia a un fármaco (89,3%). Tres cepas (S. Infantis, S. Derby y S. Enteritidis)fueron multirresistentes y una, de S. Enteritidis, dio un resultado no sensible al ácido nalidíxico y la ciprofloxacina. Se destaca la necesidad de extremar la vigilancia sanitaria integrada en el país para el control de la salmonelosis.

Effect of irradiation on Salmonella survival and quality of 2 varieties of whole green onions.

Two varieties of green onions, Banner and Baja Verde, were inoculated with a cocktail of 3 Salmonella strains using dip and spot inoculation and irradiated at 0, 0.3, 0.6, 0.9, and 1.2 kGy using electron beam. Salmonella survivors were enumerated using a XLD underlay/TSAYE overlay plating method. The D values were in the range of 0.26 to 0.32 kGy depending on variety but not on the method of inoculation. This indicated that a 5-log reduction of Salmonella can be achieved at a dose of 1.6 kGy. For the quality study, both varieties of green onions were irradiated at 0, 1.5, 2.0, and 2.5 kGy and evaluated for changes in microbial counts, color, texture, and visual quality during storage at 4 °C. Irradiation reduced total plate counts and psychrotrophs by 3 logs. Although the counts increased during storage, they did not exceed the initial counts of control. No significant difference was observed in color and texture between irradiated samples and control. The control maintained good visual quality for about 13 d as compared to 15 d for 1.5 and 2.5 kGy samples. The 2.0 kGy samples maintained good visual quality for 17 d suggesting that irradiation can increase shelf life by reducing spoilage microorganisms but higher doses can be detrimental to quality. At the dose levels required to achieve a 5-log reduction in Salmonella, the shelf life of whole green onion can be extended. This study shows that irradiation can be used to enhance safety without adverse effects on quality.

[Serotypes and antimicrobial susceptibility patterns of Salmonella strains isolated from food in Cuba]. / Serovariedades y patrones de susceptibilidad a los antimicrobianos de cepas de Salmonella aisladas de alimentos en Cuba.

The serotypes of 178 isolates of Salmonella enterica taken from food in different regions of Cuba between January 2008 and December 2009 were identified, and the antimicrobial susceptibility pattern of 100 selected isolates was determined by strata sampling. A total of 20 Salmonella serotypes were identified, with a predominance of S. Enteritidis (23%), S. Agona (13.5%), and S. London (11.2%). Of all the strains, 75% were resistant or presented intermediate resistance to at least one of the drugs tested, in the following order: tetracycline (70.7%), ampicillin (22.7%), and nalidixic acid (14.7%). Ten different resistance patterns were identified. The most frequent patterns corresponded to strains that were either drug-resistant or had intermediate resistance (89.3%). Three strains (identified as S. Infantis, S. Derby, and S. Enteritidis) were multiresistant, and one of them, S. Enteritidis, was not sensitive to either nalidixic acid or ciprofloxacin. To control salmonellosis, the importance of maximizing integrated health surveillance is emphasized.

Minimal effects of high-pressure treatment on Salmonella enterica serovar Typhimurium inoculated into peanut butter and peanut products.

About 1.2 billion pounds of peanut butter are consumed annually in the United States. In 2008 to 2009, an outbreak involving Salmonella Typhimurium in peanut butter led to a recall of over 3900 products by over 200 companies. More than 700 people became sick, 100 were hospitalized, and 9 people died from this outbreak. This study examines the efficacy of high-pressure processing (HPP) to decrease S. Typhimurium American Type Culture Collection (ATCC) 53647 inoculated into peanut butter and model systems. The viability of S. Typhimurium in peanut butter stored at room temperature was investigated. A culture of S. Typhimurium (6.88 log CFU/g) was inoculated into peanut butter. Following 28 d at 20 °C there was a 1.23-log reduction. Approximately 10(6) to 10(7) CFU/g S. Typhimurium were inoculated into 4 brands of peanut butter, 3 natural peanut butters and peanut flour slurries at 2, 5, and 10% peanut flour protein in peanut oil and in distilled water. All were treated at 600 MPa for 5 min at 45 °C. While significant differences were found between natural peanut butter and peanut protein mixtures, the reduction was <1.0 log. The peanut flour/oil mixtures had a 1.7, 1.6, and 1.0-log reduction from HPP (2, 5, and 10% protein, respectively) whereas peanut flour/water mixtures had a 6.7-log reduction for all protein levels. Oil had a protective effect indicating HPP may not help the microbial safety of water-in-oil food emulsions including peanut butter. Practical Application: There have been multiple outbreaks of foodborne illness involving peanut butter products. This study looks at the potential use of high-pressure processing to reduce the bacteria that may be in peanut butter.

Arginine and lysine decarboxylases and the acid tolerance response of Salmonella Typhimurium.

Salmonella Typhimurium CECT 443 inactivation at pH 2.5 in Mineral Medium (MM) and MM supplemented with 0.01% (w/v) arginine, lysine or glutamic acid was studied using stationary-phase cells grown in buffered BHI pH 7.0 (non-acid adapted cells) and acidified BHI up to pH 4.5 with acetic, citric, lactic and hydrochloric acids (acid adapted cells). In all cases, acid adapted cells, with D-values ranging from 23.34 to 86.90 min, showed a significantly higher acid resistance than non-acid adapted cells, with D-values between 8.90 and 10.29 min. Whereas the conditions used for acid adaptation did not exert a significant effect on the acid resistance of the S. Typhimurium CECT 443 resulting cells, the inclusion of lysine and arginine in the challenge medium protected them against acid inactivation, reaching D-values of about 2 and 3 times higher, respectively, than those found in MM or MM supplemented with glutamic acid. None of these three amino acids significantly modified the acid resistance of non-acid adapted cells. The relative expression level of adiA (encoding the arginine decarboxylase), adiY (encoding the transcriptional activator of adiA), cadA (encoding the lysine decarboxylase) and cadB (encoding the lysine/cadaverine transport protein) was examined by quantitative PCR. Acid adapted cells showed higher relative expression levels for both systems, arginine decarboxylase and lysine decarboxylase, which demonstrates that the induction of specialized pH-homeostatic systems plays an important role in S. Typhimurium CECT 443 protection against acid stress. However, the increased acid resistance showed by acid adapted cells challenged in MM arginine or lysine free suggests the existence of other microbial survival strategies.

Effects of Maillard reaction products on hilA expression in Salmonella typhimurium.

Salmonella is a foodborne pathogen causing severe gastroenteritis. Three types of Maillard reaction products (MRP) generated by heat sterilization of D-glucose and L-lysine, L-histidine, and L-arginine were studied at 2 different levels of supplementation (0.5% and 1.0%) for their influence on growth and virulence of Salmonella. Two methods, namely, real-time polymerase chain reaction (RT-PCR) and a beta-galactosidase gene fusion assay, were used to determine the expression of hilA, a regulatory gene for Salmonella pathogenicity. Neither the type of MRP nor their quantities up to 1.0% affected the growth rates of S. Typhimurium EE658 (P > 0.05). When determined by beta-galactosidase assay, lysine MRP in both levels of supplementation were not found to have any effect on the hilA expression compared to the control. The addition of histidine and arginine MRP to M9 media (0.5%) increased by 2-fold hilA induction and up to 6-fold at the higher level (1%) supplementation of these compounds. Although somewhat inconsistent, RT-PCR analyses of hilA expression confirmed the greater induction effect of arginine MRP on hilA compared to lysine MRP. In contrast to beta-galactosidase assay results, however, lysine MRP were found to increase hilA expression compared to the control in both supplementation levels in all trials. The potential of MRP serving as a bacterial virulence modulator may be a factor to be considered in food thermal processing when assessing Salmonella risk for causing foodborne disease.

Effect of controlled atmosphere storage, modified atmosphere packaging and gaseous ozone treatment on the survival of Salmonella Enteritidis on cherry tomatoes.

In recent years, outbreaks of infections associated with raw and minimally processed fruits and vegetables have been reported. The objective of this study was to analyse the growth/survival of Salmonella Enteritidis at spot-inoculated or stem-injected cherry tomatoes during passive modified atmosphere packaging (MAP), controlled atmosphere (CA) and to compare the results with those of air storage at 7 and 22 degrees C. During MAP, the gas composition equilibrated to 6% O2/4% CO2. CO2 level was maintained as 5% through the term of CA storage at 7 and 22 degrees C. The results demonstrate that S. Enteritidis can survive and/or grow during the storage of tomatoes depending on the location site of the pathogen on fruit, suspension cell density and storage temperature. During MAP, CA and air storage, S. Enteritidis with initial population of 7.0 log10 cfu/tomato survived on tomato surfaces with an approximate decrease of 4.0-5.0 log10 cfu/tomato in population within the storage period; however, in the case of initial population of 3.0 log10 cfu/tomato, cells died completely on day 4 during MAP storage and on day 6 during both CA and air storage. The death rate of S. Enteritidis on the surfaces of tomatoes that were stored in MAP was faster than that of stored in air and in CA. Storage temperature was effective on the survival of S. Enteritidis for the samples stored at ambient atmosphere; cells died completely on day 6 at 7 degrees C and on day 8 at 22 degrees C. Stem scars provided protective environments for Salmonella; an approximate increase of 1.0 log10 cfu/tomato in stem-scar population was observed during MAP, CA and air storage at 22 degrees C within the period of 20 days. Cells survived with no significant change in number at 7 degrees C. During the research, the effect of ozone treatment (5-30 mg/l ozone gas for 0-20 min) was also considered for surface sanitation before storage. Gaseous ozone treatment has bactericidal effect on S. Enteritidis, inoculated on the surface of the tomatoes and can be used for surface sanitation of S. Enteritidis on tomatoes before storage at different conditions. Ten mg/l ozone gas treatment with different time intervals of 5 and 15 min was found to be effective respectively on low and high dose inoculum levels of S. Enteritidis attached for 1 h. Another variable considered during ozone treatment was the 4 h attachment time.

Suppression of infection-induced endotoxin shock in mice by a citrus flavanone naringin.

The protective effect of the Citrus flavanone naringin was demonstrated in an endotoxin shock model based on Salmonella infection. Intraperitoneal ( i. p.) infection with 10 (8) CFU Salmonella typhimurium aroA caused lethal shock in lipopolysaccharide (LPS) -responder but not LPS-non-responder mice. Administration of 1 mg naringin 3 h before infection resulted in protection from lethal shock, similar to LPS-non-responder mice. The protective effect of naringin was time- and dose-dependent. Treatment with naringin resulted not only in a significant decrease in bacterial numbers in spleens and livers, but also in a decrease in plasma LPS levels. In addition, naringin markedly suppressed TNF-alpha and normalized the activated states of blood coagulation factors such as prothrombin time, fibrinogen concentration and platelet numbers caused by infection. Interestingly, treatment with naringin suppressed high levels of soluble CD14 and high mobility group-1 molecule caused by infection.

Rapid detection of stressed Salmonella spp. in dairy and egg products using immunomagnetic separation and PCR.

The rapid detection of an average of 5.9 stressed Salmonella cells in 25 g of food product using immunomagnetic separation (IMS) and PCR is described. For pasteurised egg yolk, egg yolk powder, ice-cream, whole egg, egg white and cheeses made from pasteurised milk PCR was applied after 16 h of preenrichment in buffered peptone water (BPW) using IMS and alkaline lysis as sample preparation method. For whole egg and egg white the BPW was supplemented with iron. For milk powder, and raw milk cheeses, the 16-h preenrichment in BPW was followed by IMS and a 4-h enrichment in Rappaport-Vassiliadis broth. In the latter case, PCR was applied on the enrichment medium after centrifugation and alkaline lysis. For PCR the primers ST11 and ST15 (Aabo et al., 1993) producing a fragment of 429 bp were used. An internal PCR control, designed to be co-amplified with the target DNA using the same primers but producing a smaller fragment of 240 bp, was used.