A Turkey survey of hygiene indicator bacteria and Yersinia enterocolitica in raw milk and cheese samples

In this research, a total of 200 dairy (raw milk, cheese) samples obtained from Ankara, were examined for the presence of Yersinia spp., total coliform and Escherichia coli. As expected, raw milk 55% (55/100) were significantly contaminated with Yersinia spp., than cheese samples 14% (14/100). Y. enterocolitica was the most commonly isolated species, and was recovered from 47.3% in raw milk 35.7% in cheese samples. The other Yersinia spp. were identified as Y. frederiksenii (31.0%, 21.4%), Y. kristensenii (12.7%), Y. intermedia (7.2%, 7.1%) and atypical Yersinia spp. (1.8%, 35.7%) in raw milk and cheese samples, respectively. All the samples of cheese examined were negative for Y. kristensenii. All Y. enterocolitica strains tested gave negative results in the autoagglutination tests and crystal violet binding test.Furthermore total coliform bacteria and E. coli were investigated in these samples. According to the analysis results, most of the samples containing Yersinia spp.were found high number of viable count cell total coliform than E. coli. Total coliform bacteria and E. coli, were detected (3.0 × 10⁴ cfu/ml and 1.5 × 10⁴ cfu/ml) in four milk samples containing Y. enterocolitica while, they were detected (2.5 × 10⁴ cfu/ml and 1.0 × 10⁴ cfu/ml) in eight milk samples containing Y. enterocolitica. In the present study, total coliform bacteria and E. coli were detected >1.1 × 10⁵ cfu/g in six cheese samples containing Y. enterocolitica and atypical Yersinia spp. The results indicate that Yersinia spp. is more likely to be isolated from foods with a high level of coliforms than from foods with low coliform counts.     

Evaluation of the factors affecting the activity of sakacin C2 against E. coli in milk

Sakacin C2 is a novel, broad spectrum bacteriocin secreted by Lactobacillus sake C2 isolated from Chinese traditional fermented cabbage. Effects of milk fat, emulsifiers, preservatives and homogenization on the activity of sakacin C2 against Escherichia coli ATCC 25922 in milk were evaluated by determined the changes of viable cell counts of E. coli ATCC 25922 after inoculation (10⁵ CFU/ml) then storage at 4 °C. Milk fat in pasteurized and homogenized milk products (low fat milk and whole milk) decreased the activity of sakacin C2 against E. coli ATCC 25922. Addition of 5 μl/ml of Tween 80 decreased the effect of sakacin C2 against E. coli ATCC 25922, but 5 mg/ml of lecithin increased the effect against E. coli ATCC 25922 in pasteurized and homogenized whole milk. Nisin and ε-Polylysine significantly increased the effect of sakacin C2 against E. coli ATCC 25922 in pasteurized and homogenized whole milk. Homogenization interfered with the activity of sakacin C2 against E. coli ATCC 25922 in pasteurized skim milk and whole milk. This study might lay the groundwork for the application of sakacin C2 as bio-preservative in milk and dairy products.     

Prevalence of Yersinia enterocolitica from food and pigs in selected states of Malaysia

The aim of this study was to determine the prevalence of Yersinia enterocolitica and its bioserotypes from food and pigs in Malaysia. Fifty-eight raw porcine (raw pork meat, internal organs and other parts) and 48 non-porcine food (raw beef, poultry products, seafood, vegetables, tofu, and pasteurised milk) from wet markets located in Kuala Lumpur, Selangor, Perak, and Pahang were examined for the presence of Y. enterocolitica. Specimens (nasal, oral and rectal swabs) from 165 pigs (from nine farms) located at central and northern parts of Malaysia were also collected for Y. enterocolitica detection. Presumptive isolates were characterised biochemically and further confirmed by PCR. Out of 58 raw porcine food, Y. enterocolitica was detected in 7 (12.1%) samples in which raw pork meat (whole meat) had the highest prevalence 5/21 (23.8%), followed by raw pork liver 1/5 (20.0%) and raw pork intestine 1/8 (12.5%). No Y. enterocolitica was isolated from the 48 non-porcine foods. Overall, two pathogenic (bioserotypes 3 variant/O:3 and 1B/O:8) and one non-pathogenic (bioserotype 1A/O:5) Y. enterocolitica strains were isolated from food. Out of 165 pigs examined, 3 (1.8%) pigs were carriers for Y. enterocolitica. All 3 pigs were asymptomatic grower pigs from Penang, carried Y. enterocolitica bioserotype 3 variant/O:3. Post-enrichment PCR approach gave a higher prevalence, 60.3%, 41.7% and 27.9% for porcine food, non-porcine food and pigs, respectively. Both pathogenic and non-pathogenic Y. enterocolitica were present in our domestic pigs and food. Improper food handling and processing may cause cross contamination of this pathogen to humans, affirms a potential risk for public health.     

Prevalence and characterization of Yersinia enterocolitica isolated from retail foods in China

Yersinia enterocolitica is an important food-borne enteropathogen that causes gastrointestinal syndromes. The aims of this study were to identify Y. enterocolitica in food samples in China, and to assess the pathogenic potential and antimicrobial resistance, and to characterize the genotypes of the isolates. From July 2011 to May 2014, a total of 2320 food samples were obtained, and 47 (2.03%) were found positive for Y. enterocolitica, while 706 retail-level ready-to-eat products and 249 vegetable samples were negative. A total of 58 Y. enterocolitica strains were isolated. All isolates belonged to biotype 1A, and the primary serotype was O:8. All strains lacked the ail, virF, ystA, and ystC virulence genes, but harbored the ystB, fepD, ymoA, fes, and sat genes. All 58 strains were sensitive to kanamycin and sulfonamide, but were resistant to two or more antibiotics. Most of the strains expressed the β-lactamase genes; the presence of blaA and blaB was detected in 97% and 100% of isolates, respectively. Many strains were resistant to trimethoprim/sulfamethoxazole (79.3%), ampicillin (91.4%), and cephalothin (91.4%). The 58 strains were grouped into three clusters and one singleton by enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) at a similarity coefficient of 70%, and each cluster was largely organized by geographical region. This study provides a valuable accounting of the prevalence of Y. enterocolitica from a nationwide survey of foods in China, and highlights the seasonal effects of Y. enterocolitica prevalence in foods in China for the first time.     

Evaluation of chlorine,benzalkonium chloride and lactic acid as sanitizers for reducing Escherichia coli O157:H7 and Yersinia enterocolitica on fresh vegetables

The effectiveness in the assurance of fresh vegetable microbiological quality of wash solutions containing 200 ppm free chlorine, 0.1 mg/ml benzalkonium chloride, 0.2% and 1% lactic acid was assessed on Escherichia coli O157:H7 and Yersinia enterocolitica contaminated lettuce and tomatoes. Y. enterocolitica reduction on tomatoes (5.08, 4.77 and 4.21 log after 0.2% lactic acid, 200 ppm chlorine and 0.1 mg/ml benzalkonium chloride treatments, respectively) were significantly higher than those for Y. enterocolitica on lettuce and E. coli O157:H7 on both vegetables. Antimicrobial treatment effects on bacterial counts and product quality after subsequent 7 day storage (4 °C and 22 °C) were determined. No pathogens were found in natural microflora of fresh vegetables.     

Biofilm formation of Yersinia enterocolitica and its persistence following treatment with different sanitation agents

The biofilm formation by Yersinia enterocolitica was studied under conditions simulating a meat processing environment. Y. enterocoltica, especially biotype 4, readily formed biofilm in pork meat juice (MJ: meat juice). Biofilm was detectable after repeated treatment simulating the everyday cleaning routine which implies the potential to survive and persist in a pork processing environment. Sodium hypochlorite was more effective a sanitizer than quaternary ammonium compound (QAC: quaternary ammonium compound) in cleaning biofilm of Y. enterocolitica.     

Growth potential of Listeria monocytogenes in soft,semi-soft and semi-hard artisanal cheeses after post-processing contamination in deli retail establishments

Opportunities for post-processing contamination of cheese may occur in deli retail establishments, either during the further cheese ripening (at maximum 14 °C), during storage and display in the refrigeration cabinet (at maximum 7 °C) or during slicing. A L. monocytogenes post-processing contamination was simulated by inoculation either on the cheese slicing surface or the cheese rind of three soft cheeses (one white-molded raw cow's milk cheese, one pasteurized cow's milk cheese with spicy herbs, one washed rind pasteurized cow and sheep's milk cheese) and two semi-hard cheeses (one smear-ripened raw cow's milk cheese and one natural-ripened raw cow's milk cheese). L. monocytogenes challenge testing was performed on 3 batches of each cheese to assess the growth potential of L. monocytogenes after 14 days storage at either 7 or 14 °C. Substantial growth of L. monocytogenes (>0.5 log CFU/g) was obtained in 79.2% of all individual challenge tests (n = 178) that were performed although huge variation in growth potential was noted among the different cheese types and storage conditions. The growth potential on soft cheeses stored at 7 °C ranged from 1.8 to 4.0 log units and from 3.6 to 5.5 log units upon storage at 14 °C, whilst on semi-hard cheese, this was in general lower, and ranged from 0.1 to 1.4 log units at 7 °C and from 0.0 to 3.0 log units at 14 °C. Overall, increased outgrowth of L. monocytogenes was noted when inoculation was performed on the cheese slicing surface compared to the cheese rind. Thus if occasional post-processing contamination takes place during storage or handling of the cheese, L. monocytogenes has the potential to grow to elevated numbers throughout a reasonably expected storage period of up to 14 days notwithstanding the presence of high numbers of indigenous lactic acid bacteria in these cheeses. Also for a defined cheese type both a considerable inter-batch and intra-batch variability was sometimes noted from the replicate testing, indicating no consistent behavior of L. monocytogenes in these fermented dairy products. As such it is recommended that appropriate hygienic measures are taken to prevent post-processing contamination. Noting the growth potential, absence of L. monocytogenes in 25 g of cheese using a multiple sample subunit approach (n = 5) at the time of production is important to ensure compliance to EU legislation 2073/2005.     

Incidence of pathogenic psychrotrophs in ice creams sold in some retail outlets in Mumbai,India

With a view to determine the microbial quality of ice creams, 30 samples of commercial brands of three flavours sold in the `open' and `packaged, (cone, cup)' forms were analyzed for their total bacterial counts (TBC), yeast and mold counts (YMC), coliforms and pathogenic psychrotrophs; Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Yersinia enterocolitica and Salmonella spp. In general in both the types of ice creams bacterial load (2.3 × 10⁴–8.5 × 10⁶ cfu/ml) was higher; particularly coliform levels were 10–100 fold higher (3.0 × 10²–5.8 × 10⁴ cfu/ml) than the safety limits prescribed by Indian Standards Institute (ISI). Staph. aureus was detected in both the types of ice creams but occurrence of B. cereus was more frequent in open samples (40%) than in packed ones (26.6%). Salmonella was not detected in any of the 30 samples tested. While 53% of the packed and 100% of the open ice creams exhibited Listeria contamination, Yersinia were detected in 33% of packed and 40% of open ones. L. monocytogenes and/or Y. enterocolitica was detected only in one of the open ice cream samples. Growth profile of Y. enterocolitica 5692 and L. monocytogenes 036 at simulated temperature abuse conditions during commercial frozen storage showed that after 10 days L. monocytogenes could grow to >1 log and 1 log cycle at 8–10°C and 2–4°C, respectively and Y. enterocolitica grew 2 log cycles at both the temperatures. Results are discussed in the context of present microbiological specifications and the need for its implementation by regulatory agencies to ward off possible health hazards arising from pathogens.     

A small study of Yersinia enterocolitica in pigs from birth to carcass and characterisation of porcine and human strains

The objectives of this study were to investigate the present of Yersinia enterocolitica at the different stages of production from birth to chilled carcasses; to characterise the isolates in terms of bioserotype, virulence factors (ail, ystA, ystB, inv, yadA and virF) and antibiotic resistance profiles and to examine strains causing diarrheal illness in Ireland. Rectal, throat, floor, partition, wall and/or carcass swabs and water samples were testing for Y. enterocolitica using the bacteriological analytical method. Presumptive positive isolates were confirmed using API 20E strips. The isolates were then combined with 10 clinical strains, biotyped, serotyped, their antibiotic resistance phenotypes determined by disc diffusion and tested for the presence of ail, ystA, ystB, inv, yadA and virF genes using multiplex PCR assays. Y. enterocolitica had an overall prevalence rate of 0.55% (4/726). Y. enterocolitica 2/O:9 and 1A/O:9 were detected on the farm and/or in the abattoir. The clinical isolates were 1A/O:5, 1A/O:8, 1A/O:9, 2/O:9 or 4/O:9. All biotype 1A strains were ail, ystA, yadA and virF negative but carried the ystB gene. All biotype 2 and 4 strains were ail, ystA, yadA and virF positive. Resistant to cephalothin, ampicillin and amoxicillin/clavulanic acid was common and one 2/O:9 strain was resistant to compound sulphonamides and tetracycline. Intermediate resistance to neomycin and streptomycin was also observed. The data presented may explain the low incidence of yersiniosis in Ireland and suggests that biotype 1A may present a public health risk. Neomycin and tetracycline resistance have rarely been reported in Y. enterocolitica and should be monitored in future surveillance studies.     

Assessment of antimicrobial activity of Nα -lauroyl arginate ethylester (LAE®) against Yersinia enterocolitica and Lactobacillus plantarum by flow cytometry and transmission electron microscopy

N^α -lauroyl arginate ethylester, LAE^®, which was approved as GRAS by the US Food and Drug Administration (FDA) and by the European Food Safety Authority (EFSA) in 2007, is a surfactant that exhibits antimicrobial activity. To assess its antimicrobial effect, treated cell suspensions of Yersinia enterocolitica and Lactobacillus plantarum were analysed for reduction of cell viability. Membrane dysfunction was determined by staining with bis-oxonol to detect the loss of membrane potential, and with propidium iodide to detect permeabilized membranes by flow cytometry. LAE^® treatment for 30 min induced a 4 log₁₀ reduction in cell viability in both bacteria; different subpopulations with variable degrees of cellular damage were observed by flow cytometry. Permeabilized membranes suggested the leakage of cellular material; this was also indicated by the loss of potassium ion(s), which was higher in L. plantarum than in Y. enterocolitica. Structural changes involving collapse of the cytosol and alterations of the cellular envelopes, mainly in Y. enterocolitica, and the formation of mesosomes in L. plantarum were observed by transmission electron microscopy.     

Effectiveness of several chemical decontamination treatments against Gram-negative bacteria on poultry during storage under different simulated cold chain disruptions

Five chemical compounds – 12% trisodium phosphate (TSP), 1200 ppm acidified sodium chlorite (ASC), 2% citric acid (CA), 220 ppm peroxyacids (PA) and 50 ppm chlorine dioxide (CD) – were analyzed to determine their effectiveness against Gram-negative bacteria (Salmonella enterica serotype Enteritidis, Yersinia enterocolitica, Escherichia coli and Pseudomonas fluorescens) artificially inoculated on skinless chicken legs. Samples were stored for 120 h under three different simulated cold chain disruptions: T1 (12 h at 1 ± 1 °C, followed by 6 h at 15 ± 1 °C and then 102 h at 4 ± 1 °C), T2 (18 h at 1 ± 1 °C, 6 h at 15 ± 1 °C and 96 h at 10 ± 1 °C) or T3 (18 h at 4 ± 1 °C, 6 h at 20 ± 1 °C and 96 h at 7 ± 1 °C). Microbiological analyses and pH determinations were carried out at 0, 24, 72 and 120 h of storage. TSP, ASC and CA significantly (P < 0.05) reduced microbial populations, relative to control (untreated) samples. TSP and ASC caused higher (P < 0.05) reductions of S. enterica than CA under moderate temperature abuse conditions (T2 and T3). TSP and ASC (T2) or TSP and CA (T1 and T3) were the most effective compounds against E. coli. TSP and ASC showed the highest antimicrobial effect (P < 0.05) at mild temperature abuse conditions (T1) for Y. enterocolitica, while ASC was the most active compound against this pathogen under T2 and T3 conditions. TSP caused the largest average reductions of Ps. fluorescens at T1. PA and DC originated scant microbial reductions under all of the conditions proposed. These findings extend current knowledge about these chemical decontaminants, and may help the Regulatory Authorities in their decisions about the poultry decontamination authorization process within the European Union.     

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.     

Occurrence and characterization of Aeromonas hydrophila and Yersinia enterocolitica in minimally processed fresh vegetable salads

A range of commercially available minimally processed ready to eat salads was examined for the presence of Aeromonas and Yersinia, to provide information about their occurrence and characterize them by some phenotypic criteria. The SDS–PAGE of whole-cell proteins was also applied as a taxonomic tool for the rapid and effective identification of Aeromonas hydrophila and Yersinia enterocolitica found among a number of Aeromonas and Yersinia isolates. Aeromonas isolates were obtained from 61.5% of the samples and more than 80% of them were characterized as A. hydrophila. Two isolates were classified by both phenotypic criteria and the SDS–PAGE of whole-cell proteins as Y. enterocolitica. These results therefore suggest the prevalence of A. hydrophila isolates and the low occurrence of Y. enterocolitica in the minimally processed salads.     

Effects of temperatures and storage time on resting populations of Escherichia coli O157:H7 and Pseudomonas fluorescens in vitro

Assessment of microbial interactions is crucial for documenting bacterial growth in pure and mixed cultures and their potential for biological applications. Pseudomonas fluorescens (non-plant pathogenic and non-pectinolytic) has been used as a biocontrol microbe for plant pathogens and food-borne bacteria. We determined the growth of Escherichia coli O157:H7 (Ec) and P. fluorescens(Pf) in monocultures and co-cultures in sterile distilled water (SDW), buffered peptone water (BPW) and trypticase soy broth (TSB). The effects of temperatures (5, 10, 15, 20, 25, 35, and 37 °C) and storage time (0, 2, 4, 6, 24, and 48 h) on bacteria populations were assessed. Bacteria counts in monocultures in SDW ranged from 2.14 to 3.03 and 2.54 to 3.31 Log CFU/ml for Ec and Pf; respectively. In BPW, mean bacteria counts (monocultures) ranged from 3.15 to 6.14 and 2.54 to 6.41 Log CFU/ml for Ec and Pf, respectively. Ec populations in co-culture varied with storage temperatures and time. After 48 h, Ec 43894 monocultures in TSB ranged from 2.17 to 8.75 and 2.31 to 8.85 Log CFU/ml at 20 and 35 °C; respectively. In co-cultures with Pf 2-79, Ec 43894 counts ranged from 1.71 to 5.83 (20 °C) and 1.90 to 9.03 Log CFU/ml (35 °C) in TSB. The reductions of Ec by Pf 2-79 varied among strains and generally ranged from 0.20 to 0.90, 0.63 to 1.18, and 0 to 0.56 Log CFU/ml in BPW (10 °C). Substrate availability, storage temperatures, and time significantly (P < 0.05) impacted Ec populations in co-culture. The liquid substrate experiments indicated suppressive conditions of Ec by Pf, however; the reduction of produce contamination by E. coli O157:H7 during transitory temperature abuse conditions such as the transportation of produce from fields needs further investigation.     

How safe is European Internet cheese? A purchase and microbiological investigation

The suitability for consumers of a variety of raw milk cheeses purchased over the Internet was investigated in terms of packaging, labelling, physicochemical parameters and microbiological safety. 108 purchases from seven European countries were examined. The prevalences of Salmonella spp., Listeria monocytogenes, Escherichia coli and coagulase positive staphylococci (SA) were determined. All 108 samples were described on websites as raw milk cheeses and thereby qualified for this study. However, after delivery it was noted that 4.6% (5/108) of cheeses were labelled to be manufactured from heat-treated or pasteurized milk. Delivery duration ranged from 24 h to six days. Immediately upon receipt cheese temperatures were observed to range between 5 and 23 °C, whereas in 61.5% of all cases the temperature was higher than 15 °C. Cheese labelling was examined in respect of EC Guideline 2000/13 and Regulation No. 853/2004. Only 17.6% (19/108) of cheeses were properly labelled and fulfilled all European guideline requirements. In 50.9%, 38.8%, 46.3% and 39.8% of all cases (i) specific storage requirements, (ii) name and address of the manufacturer/packer or seller, (iii) net weight and (iv) shelf life (use by date), were missing. Even the labelling information “made from raw milk” was not apparent on 36% of all cheese items delivered. The major foodborne pathogen L. monocytogenes was detected in 1.9% of all samples, one of which had counts of 9.5 × 10³ CFU/g. None of the 108 investigated cheeses showed a pH ≤ 5.0 and a_w value ≤0.94 which are the limiting values for growth of L. monocytogenes. For two samples (0.9%) and 11 samples (10.2%) the pH and the a_w value was ≤4.4 or ≤0.92, respectively at least at one of three stipulated time points (receipt, mid-shelf-life and at expiry). Salmonella spp. could not be detected in any of the samples. E. coli and SA could be detected in a total of 29.6% (≥10 CFU/g; 32/108) and 8.3% (≥100 CFU/g; 9/108) of samples, respectively, indicating poor conditions of hygiene. Results reveal that labelling and hygiene concerns about the safety of Internet purchased cheeses in Europe are justified.     

Growth of Escherichia coli and Staphylococcus aureus in individual lasagne layers and evidence for migration of E. coli throughout the product

Growth of Escherichia coli and Staphylococcus aureus within individual layers of lasagne was studied after spiking of ～10⁵–10⁶ CFU/g of each bacterial species into bolognese or béchamel sauces. Both E. coli and S. aureus grew by 3–4 log₁₀ cycles in each meal component. In a second study, alternate layers within a composite lasagne meal were spiked with ～10⁵–10⁶ CFU/g of E. coli and adjoining layers were monitored for possible bacterial migration. Spiked composite meals were subjected to either low temperature storage at 4 °C up to 8 h or to freeze chilling, which involved freezing at ?18 °C for 24 h followed by thawing at 4 °C up to 40 h. Migration of E. coli from inoculated layers to the adjoining layers was indicated by a recovery of substantial populations following both storage treatments. Migration appeared to be more limited for meals which underwent freeze chill treatment. In contrast, migration was evident throughout all product layers in meals stored at 4 °C only. Migration of bacteria throughout a multi-layered food may arise from differing compositional or structural characteristics within the product or from differing storage treatments. Therefore as a result of bacterial migration, it would appear that microbiological safety of multi-layered products such as lasagne relies on ensuring safety of each individual layer.     

Thermal inactivation and growth of Listeria monocytogenes during production and storage of caramel apples

During the fall of 2014, commercially produced pre-packaged caramel apples were linked to 35 cases of listeriosis in 12 states. In response, this study aimed to assess 1) the reduction of different outbreak and non-outbreak strains of Listeria monocytogenes during caramel dipping of apples, and 2) subsequent growth of the apple outbreak strains within caramel apples during storage at 22 and 4 °C. In aim 1, three unwaxed Jonathan apples were dip-inoculated with three different 4-strain L. monocytogenes cocktails (apple outbreak, unrelated outbreak or unrelated environmental) at ∼8 log CFU/apple, dried for 1 h, dipped for 5 s in caramel at 82, 88, 93 or 99 °C, cooled for 1 h at room temperature and assessed for survivors. In aim 2, Jonathan apples were spot-inoculated with the apple outbreak cocktail (∼3 log CFU/apple) at the stem juncture, dried for 1 h, pushed onto wooden sticks, and dipped in caramel at 82 °C. During storage at 4 and 22 °C for 28 and 14 days, respectively, four different apple sections (top, middle, bottom and core) were cut from three apples, homogenized and plated for Listeria. After dipping apples in caramel at 82 and 99 °C, the apple outbreak, unrelated outbreak and environmental Listeria strains decreased 2.0 ± 0.6 and 2.7 ± 0.1, 1.8 ± 0.3 and 2.6 + 0.1, and 1.7 ± 0.1 and 2.9 ± 0.2 logs, respectively, with the environmental cocktail significantly less heat resistant (P < 0.05) at 99 °C compared to the other two cocktails. After 14 days of storage at 22 °C, Listeria populations were significantly higher (P < 0.05) in the core (7.4 ± 0.6 log CFU/g) compared to the other three sections (4.9–5.4 log CFU/g). The same trend was seen for the core (7.7 ± 0.6 log CFU/g) and the other three sections (5.0–5.4 log CFU/g) after 28 days of storage at 4 °C. Since dipping in hot caramel cannot ensure pathogen elimination, producers of caramel apples should implement good agricultural practices, post-harvest preventive controls and refrigeration of the final product to minimize the risks from Listeria.     

Growth and biofilm formation by Listeria monocytogenes in cantaloupe flesh and peel extracts on four food-contact surfaces at 22 °C and 10 °C

The nationwide listeriosis outbreak that occurred in the United States during 2011 highlighted the importance of preventing cantaloupe contamination with Listeria monocytogenes (Lm) within farm and processing environments. The objectives of this study were to determine the effects of strain and temperature on growth and biofilm formation of Lm in cantaloupe flesh and peel extracts on different food-contact surfaces. Growth of Lm strains was markedly greater at high concentration of cantaloupe extracts and temperature in comparison to low concentration and temperature. For 50 mg/ml of cantaloupe extract inoculated with 3 log CFU/ml, the growth of Lm was 8.5 log CFU/ml in 32 h at 22 °C and 6–7 log CFU/ml in 72 h at 10 °C. For 2 mg/ml of cantaloupe extract that was inoculated with Lm, the growth was 7–7.5 log CFU/ml in 72 h at 22 °C and 3.5 log CFU/ml in 72 h at 10 °C. There were no differences (P > 0.05) among Lm strains for biofilm formation in cantaloupe extracts, but biofilm formation was greater at high temperature and high concentration. For 50 mg/ml cantaloupe extracts inoculated with 3 log CFU/ml, the biofilm formation of Lm on stainless steel surface was approximately 7 log CFU/coupon at 22 °C in 4–7 days and 5–6 log CFU/coupon at 10 °C in 7 days. For 2 mg/ml cantaloupe extracts, the biofilm formation of Lm on the stainless-steel surface was approximately 5–6 log CFU/coupon at 22 °C and 4–4.5 log CFU/coupon at 10 °C in 7 days. The biofilm formation by cantaloupe outbreak strain Lm 2011L-2625 in cantaloupe extracts was least on buna-n rubber when compared to stainless steel, polyethylene and polyurethane surfaces (P < 0.05). These findings show that a very low concentration of nutrients from cantaloupe flesh or peel can induce Lm growth and subsequent biofilm formation on different food-contact processing surfaces.     

Survival of microencapsulated Bifidobacterium longum in Cheddar cheese during production and storage

The aim of this study was to evaluate the effect of microencapsulation (ME) in alginate beads on the viability of Bifidobacterium longum 15708 in terms of their tolerance to freezing, storage in a frozen state, cheddar cheese manufacturing and storage as well as to a simulated gastro-intestinal environment. Two ME methods namely i) droplet extrusion method (ADE) and ii) emulsion method, involving two polymers (native (NA) and palmitoylated alginate (PA)) were compared. Results showed that ADE maintained higher viability of B. longum after 24 h freezing at −80 °C with no viability loss as compared to the emulsion process and free cells which lost approximately 0.8 and 1.5 log CFU/mL respectively. However, during a 4 weeks storage period at −80 °C, no significant difference (P > 0.05) was observed in the survival of free and immobilized B. longum, with no loss of viability. Cheddar cheeses supplemented with B. longum culture were prepared and analysed during storage at 4 °C. After 21 days of storage, Cheddar cheese containing encapsulated B. longum in NA and PA polymers produced with the emulsion process showed a good survival with 2 log CFU/mL reduction after 21 days, as compared to ADE-encapsulated B. longum and free cells with 3 and 4 log CFU/mL reductions respectively. The immobilized bacteria in both polymers were also more resistant than free cells to simulated gastric and intestinal environments by a factor of 30.     

Inhibition of Listeria monocytogenes in vitro and in goat milk by liposomal nanovesicles containing bacteriocins produced by Lactobacillus sakei subsp. sakei 2a

Lactobacillus sakei subsp. sakei 2a is a bacteriocinogenic lactic acid bacteria isolated from a Brazilian meat product, capable to inhibit Listeria monocytogenes in vitro and in foods. In this study, bacteriocin produced by this strain were encapsulated in phosphatidylcholine (FC) and 1,2-dioleoyloxy-3-trimethylammonium-propane (DOTAP) liposomes, separately and in combination, were characterized and evaluated for activity against L. monocytogenes in vitro and in experimentally contaminated UHT goat milk, during storage at 7 °C for 14 days and 30 °C for 24 h. The FC and DOTAP/FC (3:1) nanovesicles containing bacteriocins (12.800 AU mL⁻¹) presented zeta potential of −1.54 and + 38.13 mV, Entrapment Efficiency of 80.0% and 94.1%, and diameter of 91.19 and 81.49 nm, respectively. DOTAP/FC nanovesicle presented excellent stability, and maintained the same physicochemical characteristics over 28 days. Both free and encapsulated bacteriocins controlled L. monocytogenes growth in BHI medium and goat milk stored at 30 °C for at least 8 h, in a similar pattern. After 24 h in BHI medium, bacteriocins encapsulated in FC nanovesicles were more effective (p < 0.05) than free bacteriocins. However, in goat milk, no significant differences (p ≥ 0.05) were observed for the two types of nanovesicles. At 7 °C, both free and encapsulated bacteriocins retarded the L. monocytogenes growth, and after 5 days, counts were 5 log lower than in the controls, both in BHI and in goat milk. Encapsulation of bacteriocin in FC and DOTAP/FC nanovesicles did not affect the antimicrobial activity, but the advantages of their application for control of L. monocytogenes in goat milk based dairy products, when compared to free bacteriocins, remain unclear and more studies are needed.