Antimicrobial peptides derived from hen egg lysozyme with inhibitory effect against Bacillus species

In food industry, Bacillus species are encountered in deteriorating many food products thus shortening their shelf-life. Moreover, Bacillus cereus and the subtilis group (Bacillus subtilis, Bacillus licheniformis, and Bacillus pumilus) have been recognized as food poisoning agents. Lysozyme peptides preparation (LzP) is a commercially available as a natural food preservative. Although, LzP derived from lysozyme yet it showed only 11% of the lysozyme lytic activity. LzP at a concentration of 100 μg ml⁻¹ completely inhibited B. subtilis, B. licheniformis, B. megaterium, B. mycoides, B. pumilus, B. coagulans, B. amyloliquefaciens, B. polymexa and B. macerans. However, B. cereus and B. stearothermophilus showed a slightly higher resistance. Interestingly, LzP at concentration ⩾10 μg ml⁻¹ showed inhibitory effect on both vegetative and spore forms of B. subtilis. Moreover, LzP was stable at 95 °C for 30 min and at different pH values (4.5–7). In conclusion, LzP may be useful to control growth of Bacillus spoilage organisms.     

Characterization and application of an anti-Listeria bacteriocin produced by Pediococcus pentosaceus 05-10 isolated from Sichuan Pickle,a traditionally fermented vegetable product from China

Pediococcus pentosaceus 05-10, isolated from a traditionally fermented Sichuan Pickle, produced a bacteriocin (Pediocin 05-10) active against Listeria, Lactobacillus, Streptococcus, Enterococcus, Pediococcus and Leuconostoc. Pediocin 05-10 was sensitive to proteolytic enzymes, but stable between pH 2–10 and heat resistant (15 min at 121 °C). It did not adhere to the surface of the producer cells. However, adsorption to both resistant and sensitive cells was observed. Production of the bacteriocin started at the early exponential phase and reached its maximum at the early stationary phase. This result suggested that Pediocin 05-10 was produced in a growth-associated manner. Its mode of action was bactericidal, as determined against Listeria monocytogenes 54002. Pediocin 05-10 was estimated below 6.5 kDa by tricine–SDS–PAGE. The application experiment showed that Pediocin 05-10 could significantly reduce the counts of L. monocytogenes 54002 in pork ham during storage at 4 °C for 10 days. Thus, Pediocin 05-10 has potential for application in food preservation, especially in the meat products industry.     

Heat inactivation of Listeria innocua in broth and food products under non-isothermal conditions

The objective of this work was to study the effect of three linear temperature profiles (heating rates of 1.5, 1.8 and 2.6 °C/min, from 20 to 65 °C) on Listeria innocua inactivation in liquid medium. The inactivation was also analyzed in artificially contaminated parsley (heating rate of 1.8 °C/min) and throughout a frying process, using a pre-cooked frozen food as case study. Inactivation showed a sigmoidal behaviour and all data was fitted with a Gompertz-inspired model. Results demonstrated that, in liquid media, Listeria inactivation is influenced by the temperature profile used. As heating rate increases, the shoulder decreases and the tail effect disappears. If Listeria was in parsley, its heat resistance increased (for identical experimental conditions in broth). Besides model adequacy was proven in all studied situations, the heating rate affected parameters’ precision.     

Efficiency of high pressure treatment on inactivation of pathogenic microorganisms and enzymes in apple,orange, apricot and sour cherry juices

The purpose of this study was to investigate the effect of high hydrostatic pressure with a mild heat treatment on Staphylococcus aureus 485, Escherichia coli O157:H7 933 and Salmonella Enteritidis FDA in apple, orange, apricot and sour cherry juices. The effectiveness of the treatment on polyphenol oxidase activity in apple juice and pectinesterase activity in orange juice were also determined. An inoculum of microorganisms was completely inactivated at 350 MPa and 40 °C in 5 min. The residual polyphenol oxidase activity in apple juice after treatment at 450 MPa and 50 °C for 60 min was obtained as 9 ± 2.2%. The residual pectinesterase activity in orange juice after treatment at 450 MPa and 50 °C for 30 min was determined as approximately 7 ± 1.6%. It compares with 12 ± 0.2% at a treatment of 40 °C and 450 MPa for 60 min. Pressure resistant isoenzymes were thought to be responsible for the final residual activity. The inactivation is irreversible and the enzyme is not reactivated upon storage. High pressure processing constitutes an effective technology to inactivate the enzymes in fruit juices. Pressures higher than 400 MPa can be combined with mild heat (<50 °C) to accelerate enzyme inactivation.     

Comparison of aluminum thermal-death-time disks with a pilot-scale pasteurizer on the thermal inactivation of Escherichia coli K12 in apple cider

This study was conducted to compare thermal inactivation kinetics obtained using a pilot-scale pasteurizer and a bench-scale processing system. Pilot-scale pasteurizers are useful for product development, but comparisons on thermal inactivation kinetics with smaller scale systems are lacking. Using an Armfield pilot-scale pasteurizer and aluminum thermal-death-time (TDT) disks, the D-values and z-values of Escherichia coli K12 in apple cider were determined in the temperature range of 54–62 °C. Come-up times to 58 °C were also measured and were 35 and 61 s for the TDT disks and pasteurizer, respectively. The D-values from the TDT disks were 9.66, 4.01, 1.44 and 0.44 min at temperatures of 54, 56, 58, and 60 °C, respectively. The D-values from the pasteurizer were 3.48, 1.22, 0.10 and 0.05 min at temperatures of 56, 58, 60, and 62 °C, respectively. The z-values from the TDT disks and the pasteurizer were 4.68 and 3.60 °C, respectively. There was no significant (P > 0.05) difference in the D-values of the TDT disks and pasteurizer at 56 and 58 °C, while there was a significant (P < 0.05) difference in the D-value at 60 °C and in the z-value. This study revealed that the thermal inactivation kinetics obtained using bench scale TDT disks and an Armfield pilot-scale pasteurizer under certain conditions are similar. However, based on ease of use and other factors, TDT disks are preferable for acquiring thermal inactivation kinetics.     

Antimicrobial activity and partial characterization of bacteriocin-like inhibitory substances (BLIS) produced by Bifidobacterium infantis BCRC 14602

Bifidobacterium infantis BCRC 14602 was found to produce a bacteriocin-like inhibitory substance (BLIS) with inhibitory activities against a wide range of Gram- positive and Gram-negative bacteria. An activity level of 400 AU/ml in the middle of the exponential phase (i.e. 6 h) and maximum activity (1600 AU/ml) at the beginning of the stationary phase (i.e. 16 h) was recorded in MRS broth at 37 °C. BLIS was partially purified by a two-step purification protocol resulting in a specific activity of 31,605 AU/mg and a purification fold of 120. Based on Tricine–SDS–PAGE, the BLIS is approximately 3.0 kDa in size. Complete inactivation of BLIS activity was observed after treatment with proteolytic enzymes, but not with catalase, α-amylase and lipase. The adsorption of the BLIS to the producer cells was strongly affected by the pH of the broth culture of which 100% adsorption to the killed cells occured between pH 6.0 and 7.0, whereas at pH values below 6.0 and above 7.0, the adsorption ratio decreased to 43 and 60%, respectively. BLIS showed high temperature stability up to 121 °C for 15 min with no loss in its activity, and had pH stability in the range of 4–10. The temperature and heat stability of BLIS makes it useful for applications in food processing technologies and food safety control applications.     

Non-thermal inactivation of Escherichia coli K12 in buffered peptone water using a pilot-plant scale supercritical carbon dioxide system with a gas–liquid porous metal contactor

This study evaluated the effectiveness of a supercritical carbon dioxide (SCCO₂) system, with a gas–liquid porous metal contactor, for reducing Escherichia coli K12 in diluted buffered peptone water. 0.1% (w/v) buffered peptone water inoculated with E. coli K12 was processed using the SCCO₂ system at CO₂ concentrations of 3.1–9.5 wt%, outlet temperatures of 34, 38, and 42 °C, a system pressure of 7.6 MPa, and a flow rate of 1 L/min. Increased CO₂ concentrations and temperatures significantly (P < 0.05) enhanced microbial reduction. A maximum reduction of 5.8-log was obtained at 8.2% CO₂ and 42 °C. To achieve a 5-log reduction of E. coli K12 in 0.1% buffered peptone water, minimum CO₂ concentrations of 9.5%, 5.5%, and 5.3% were needed at 34, 38, and 42 °C, respectively. Further reductions of cells were observed after storage for 7 days at 4 °C. But storage at 25 °C increased the number of viable cells to 8-log cfu/mL after 7 days. This study showed the potential of the pilot scale SCCO₂ system with a gas–liquid porous metal contactor for microbial inactivation in liquid food.     

Effect of organic acids,hydrogen peroxide and mild heat on inactivation of Escherichia coli O157:H7 on baby spinach

Minimally processed baby spinach contaminated with Escherichia coli O157:H7 has been associated with multiple outbreaks of foodborne illnesses recently. Chlorinated water is widely used to wash vegetables commercially, but this washing procedure has limited efficacy and can lead to the formation of carcinogenic substances. This study was conducted to determine the effects of organic acids and hydrogen peroxide alone and in binary combinations with or without mild heat (40 and 50 °C) on the inactivation of Escherichia coli O157:H7 on baby spinach. Baby spinach leaves were dip-inoculated with E. coli O157:H7 to a level of 6 log CFU/g and stored at 4 °C for 24 h before treatment. Individual washing solutions (1% and 2% lactic acid [LA], citric acid [CA], malic acid [MA], tartaric acid [TA], acetic acid [AA], hydrogen peroxide [H₂O₂] as well as binary combinations of LA, CA, MA and H₂O₂ at final concentrations of 1% were used to decontaminate spinach leaves at 22, 40 or 50 °C for 2–5 min to test their efficacy in reducing E. coli O157:H7. Chlorinated water (200 ppm free chlorine) decreased the population of E. coli O157:H7 on baby spinach by only 1.2–1.6 log CFU/g, which was not significantly different from DI water washing. Washing with 1% LA at 40 °C for 5 min was the most effective treatment achieving a 2.7 log reduction of E. coli O157:H7 which is significantly higher than chlorine washing. Washing with LA + CA or LA + HP at 40 °C for 5 min was equally effective against E. coli O157:H7, resulting in a 2.7 log reduction of E. coli O157:H7. The application of mild heat significantly enhanced the efficacy of washing solutions on the inactivation of E. coli O157:H7. There was, however, no significant difference between treatments at 40 °C for 5 min and 50 °C for 2 min. The results suggested that the use of organic acids in combination with mild heat can be a potential intervention to control E. coli O157:H7 on spinach.     

HPLC determination for prostaglandins from seaweed Gracilaria gigas

Seaweed (Gracilaria gigas) is an edible red alga and occasionally induces food poisoning cases. Prostaglandin E₂ (PGE₂) has been reported to be possible causative agent. In this study, a simple, sensitive, rapid and accurate HPLC method was developed for quantifying prostaglandins in seaweed. The mobile phase was gradient acetonitrile (35–60%) and 0.017 M phosphoric acid at flow rate of 1 mL/min within 30 min. The standard curves of prostaglandins were extremely linear (R² > 0.999) with low correlation coefficients (less than 4.7) in the range of 5–50 μg/mL. To obtain maximum prostaglandins amount, the optimal ratio of seaweed (wet weight) to arachidonic acid was 10 g:2 mg and oxygen was needed in reaction.     

Consumer awareness and perception to food safety hazards in Trinidad,West Indies

Little is known on consumer perception and awareness to food safety in Trinidad, West Indies. A survey was conducted on 121 consumers who handled meat on food safety knowledge and handling practices at homes. Most (P < 0.01; 83.2%) categorised food safety as ‘very’ important. Consumers differed (P < 0.05) in their perception as to the most feared food hazard. Escherichia coli (89.7%) and Salmonella (85.7%) were most known (P < 0.01) of microbial types. Restaurant (55.0%) was regarded to be the most likely place where food poisoning could occur. There was no distinct (P > 0.05) trust in food safety authorities. Gender had (P > 0.05) no influence on responses. The study highlighted gaps in food safety knowledge and critical violations in food handling.     

Superheated steam reduction of deoxynivalenol in naturally contaminated wheat kernels

Contamination of wheat with the Fusarium mycotoxin deoxynivalenol (DON) is a concern to the ethanol industry as it is stable during most processing operations and will be concentrated in the spent grains, which are potentially a valuable feedstock. Superheated steam at four processing temperatures (110, 135, 160, and 185 °C), three steam velocities (0.65, 1.3, and 1.5 m/s), and processing times of 2–15 min were used to treat wheat kernels naturally contaminated with DON to find the best processing parameters for the reduction of DON. A commercial enzyme-linked immunosorbent assay (ELISA) kit was used to determine DON levels in the wheat samples. Samples became increasingly toasted, displaying a brown color with increasing processing temperatures and times, and became friable after processing at temperatures of 160 and 185 °C. Only samples processed at 185 °C and 1.3 m/s exhibited any starch gelatinization. Significant (P < 0.05) reductions in DON levels were seen at 160 and 185 °C but were not generally seen at 110 and 135 °C and the effect of velocity was not significant (P > 0.05). Reductions of up to 52% were achieved at 185 °C and 6 min processing time and were due only to thermal degradation and not to solubilization and extraction.     

High hydrostatic pressure treatment applied to model cheeses made from cow’s milk inoculated with Staphylococcus aureus

Pasteurized milk was inoculated with two strains of Staphylococcus aureus (CECT4013 or ATCC13565) and used to elaborate soft-curd cheeses with approximately 7.5-log CFU/g of S. aureus. Cheeses were submitted to 10 min high hydrostatic pressure (HHP) treatments of 300, 400 or 500 MPa at 5 °C or 20 °C. Staphylococcus enterotoxin (SE) was evaluated in cheeses containing ATCC13565. Counts of S. aureus were measured after HHP treatment (day 1) and after 2, 15 and 30 days ripening at 8 °C. Inactivation increased with pressure and storage time, but was similar for both treatment temperatures. Maximum S. aureus reductions were achieved after 30 days ripening for samples treated at 500 MPa and 5 °C: 6.0 ± 0.1 and 4.7 ± 0.5-log CFU/g for CECT4013 and ATCC13565, respectively. However, SE was detected in all cheese samples containing ATCC13565 before and after HHP and after 30 days ripening.     

Inactivation of Staphylococcus spp. strains in whole milk and orange juice using ultra high pressure homogenisation at inlet temperatures of 6 and 20 °C

The objective of this work was to evaluate the inactivation induced by ultra high pressure homogenisation (UHPH) of Staphylococcus aureus ATCC 13565 and Staphylococcus carnosus CECT 4491 inoculated into milk and orange juice considering the effect of inlet temperature of the sample (6 and 20 °C) on the lethality values and on the production of sublethal injuries. Samples of UHT whole milk and UHT orange juice were inoculated at a concentration of approximately 7.0 log (CFU/ml) and pressurized with a dual valve UHPH machine at 300 MPa at the primary homogenising valve and at 30 MPa on the secondary valve. Viable and injured bacterial counts were measured 2 h after UHPH treatment and after 3, 6, and 9 days of storage at 4 °C for milk, and after 3, 6, 9, 12, and 15 days of storage at 4 °C for orange juice. The inlet temperature, the food matrix and the kind of strain influenced significantly (P < 0.05) the lethality level, which was higher for S. aureus in whole milk at an inlet temperature of 20 °C. No sublethal injuries were detected after treatments. The change over time of viable counts for both strains showed a very strong decreasing tendency during the storage at 4 °C for orange juice, while the strain S. carnosus showed a low decreasing tendency and greater resistance when inoculated in milk and pressurized at 6 °C.     

Effects of high pressure homogenization on the activity of lysozyme and lactoferrin against Listeria monocytogenes

The effects of high pressure homogenization treatment at 100 MPa (HPH), in comparison to different heat treatments, 70 °C for 30 s, 70 °C for 5 min or 100 °C for 5 min, on the activity of lysozyme and lactoferrin, were studied. The antimicrobial activities of lysozyme and lactoferrin were tested on Listeria monocytogenes inoculated in milk or cultural medium.The results indicated that antimicrobial activities of lactoferrin and lysozyme were enhanced and/or accelerated by HPH treatment. Particularly, the highest immediate inactivation values were recorded when L. monocytogenes cells were added to HPH-treated lactoferrin, processed simultaneously or separately with the target microorganism. Although to a lesser extent than HPH treatment the heat treatments applied also were able to increase the antimicrobial activity of lysozyme.     

Decontamination of fresh produce by the use of slightly acidic hypochlorous water following pretreatment with sucrose fatty acid ester under microbubble generation

Treatment by slightly acidic hypochlorous water (SAHW) in combination of pretreatment with sucrose fatty acid ester under microbubble generation was effective for decontamination of lettuce. Sufficient contact time of SAHW containing 30 mg/L of available chlorine on reduction of viable counts of lettuce was determined to be 5 min. For 5 min at 18–20 °C, treatment with 30 mg/L of available chlorine in SAHW appeared more effective in the reduction of bacteria on lettuce compared with 15 mg/L. The treatment of lettuce at 50 °C with SAHW at 30 mg/L of available chlorine showed a 2 log reduction of bacterial counts without injury in the tissue. The treatment at 50 °C, SAHW also delayed browning on cut lettuce for the first 5–6 days of subsequent storage at 6 °C. Among two sucrose fatty acid esters tested, sucrose monopalmitate at 100 mg/L had a higher efficacy for pretreatment under microbubble generation. After pretreatment for 5 min with 100 mg/L sucrose monopalmitate under microbubble generation and subsequent treatment with SAHW at 50 °C for 5 min, viable counts of lettuce were decreased by about 3–4 logs. After the same treatment, Pseudomonas sp. predominant on lettuce decreased drastically. These results indicate the effectiveness of the combined treatments of sucrose fatty acid ester under microbubble generation and SAHW at 50 °C for decontamination of fresh produce.     

Effect of combined physico-chemical treatments based on enterocin AS-48 on the control of Listeria monocytogenes and Staphylococcus aureus in a model cooked ham

Enterocin AS-48 was tested alone or in combination with chemical preservatives and/or heat against Listeria monocytogenes and Staphylococcus aureus in a cooked ham model system. AS-48 (20, 40 and 60 μg g⁻¹) alone was active against L. monocytogenes at 5 and 15 °C, but it was not sufficient to avoid regrowth of Listeria during the 60 days storage. Combination of AS-48 (40 μg g⁻¹) with nitrite/nitrate, pentasodium tripolyphosphate, sodium benzoate or potassium sorbate improved the anti-listeria effect during storage at 5 °C. The most effective combination was AS-48-nitrite/nitrate (0.007%) that reduced listeria below detection level from the beginning to end of storage. Although much more resistant, S. aureus was also inhibited by AS-48 alone at 5 °C, and especially in combinations with nitrite/nitrate, pentasodium tripolyphosphate, sodium lactate and sodium acetate. Best results against both pathogens were obtained when sodium pyrophosphate was applied in combination with 60 μg g⁻¹ AS-48. Sub-lethal heat (60 °C, 2 min) clearly increased AS-48 activity against both Listeria and Staphylococcus.     

Microbial profile of buffalo sausage during processing and storage

A study was made on the microbial levels of buffalo sausage during preparation and storage at 4 ± 1 °C. Microbial counts in raw minced meat were, total plate count (TPC) (log cfu/g) 5.41 ± 0.25; coliforms (MPN/g) 23.2; Staphylococcus aureus (log cfu/g) 1.57 ± 0.11; yeasts and molds (log cfu/g) 2.29 ± 0.07 and lactic acid bacteria (LAB) (log cfu/g) 0.60 ± 0.20. Sausage emulsion showed similar trend in microbial counts with minimal microbial contamination during the preparation of emulsion. Cooked buffalo sausage gave the following microbial counts: TPC (log cfu/g) 3.75 ± 0.31; coliforms (MPN/g) 0.2; LAB (log cfu/g) 0.07 ± 0.01; yeast and molds (log cfu/g) 0.72 ± 0.07. S. aureus, Clostridium perfringens and Bacillus cereus were not detected in cooked sausages. These results indicate that steam cooking for 45 min followed in the study was effective in reducing the microbial counts substantially. The investigation revealed that shelf life of cooked buffalo sausage was 31 days in either vacuum or CO₂ at 4 ± 1 °C. The results indicated that spoilage of vacuum packed cooked buffalo sausage was likely due to LAB while microflora other than LAB may be responsible for spoilage of CO₂ packed cooked buffalo sausage. The study suggests that measures such as low initial microbial counts, hygienic precautions during preparation of sausage, steam cooking for 45 min, vacuum or CO₂ packing and storage at 4 ± 1 °C would control the microbial growth and provide wholesomeness and safety to the buffalo sausage.     

Sensory quality and food safety of boneless chicken breast portions thawed rapidly by submersion in hot water

Boneless chicken breast portions were thawed by submersion in hot water (60 °C) and compared to refrigerator thawing. Thawing in hot water was significantly quicker (2–8.5 min) than refrigerator thawing (10–15.5 h). Thawing time in hot water increased with an increase in meat thickness. Sensory panelists could not distinguish a difference between hot water versus refrigerator thawed and subsequently grilled chicken breast portions. A model for Salmonella growth predicts that thawing chicken breast at the slowest rate in this study (0.5 °C/min) would result in a lower increase in the Salmonella concentration than that expected for room temperature storage for 4 h.     

Formation and diffusion mechanism of histamine in the muscle of tuna fish

This study investigates the formation and accumulation of histamine in the fresh meat of tuna fish. Under sterilized conditions, histamine was not detected in the muscles of Thunnus obesus (T. obesus) stored at 20 °C and 25 °C for 72 h (data not shown). And histamine formation and the diffusion mechanism was studied in T. obesus meat inoculated with two histamine-forming bacteria, Morganella morganii NBRC 3168 (M. morganii NBRC 3168) and Photobacterium phosphoreum NBRC 13896 (P. phosphoreum NBRC 13896) and stored at 20 °C and 25 °C for 3 days. The histamine level of the inoculated A1 point accumulated at a level above 4000 mg/kg in the T. obesus sample inoculated with M. morganii NBRC 3168 for 48 h. And the most level of the histamine in the remote B point was 2000 mg/kg at the time the sample was inoculated with M. morganii NBRC 3168 and stored at 25 °C. For the P. phosphoreum NBRC 13896 however, the level of histamine at the inoculated point A was 1800 mg/kg for 48 h when it was stored at 25 °C, while the highest level of histamine distant from the inoculum site B point was found to be approximately 1800 mg/kg. In contrast, when stored at 20 °C, histamine level was higher for the sample inoculated with P. phosphoreum NBRC 13896 than with M. morganii NBRC 3168. While histamine was diffused from the inoculation point in the M. morganii NBRC 3168 sample, it was diffused not only from the inoculated point, but also the remote area in the P. phosphoreum NBRC 13896 sample.     

Investigation of the effects of food constituents on Bacillus subtilis reduction during high pressure and moderate temperature

Our published results and our studies for optimization of process conditions to inactivate Bacillus subtilis by high hydrostatic pressure and mild heat using response surface methodology indicated that the optimum process parameters for a six-log-cycle reduction of B. subtilis were obtained as temperature, 46 °C; pressure, 479 MPa; and pressure holding time, 14 min. Based on the results, response surface methodology (RSM) was employed in the present investigation, the effects of food constituents like soybean protein, soybean oil, sucrose, and pH of food matrix on the B. subtilis reduction during high pressure and moderate heat was studied, and a quadratic polynomial predictive model for the effects of food constituents and pH of food matrix on B. subtilis reduction during high pressure and moderate heat was built with RSM accurately. The experimental results showed that the efficiencies of B. subtilis reduction in milk buffer and food matrix designed in the present work, under the condition of high pressure treatment process parameters described above, had some differences. The soybean protein (P < 0.0001), sucrose (P < 0.0001), and pH (P = 00006) significantly affected reduction of B. subtilis. The effect of soybean oil on reduction of B. subtilis was not significant (P = 0.8363). The adequacy of the predictive model equation for predicting B. subtilis reduction in food matrix by high pressure and moderate heat was verified effectively using experimental test date that was not used in the development of the model.