Influence of food soiling matrix on cleaning and disinfection efficiency on surface attached Listeria monocytogenes

Cleaning and disinfection are essential steps in preventing contamination of foods with pathogenic and spoilage bacteria. The efficacy of cleaning and disinfection products differ depending on target bacteria and type of soiling. We evaluated the effectiveness of cleaning and disinfecting products against Listeria monocytogenes attached on food soiled inert surfaces in a laboratory model. The number of bacteria on surfaces before and after treatment was quantified using indirect conductometric measurements. L. monocytogenes attached to stainless steel surfaces in fish broth systems reached approx. 10⁴ CFU/cm². However, all cleaning and disinfection products were equally effective in this system since all bacteria were removed or killed. When the steel disks were immersed in a fish or meat emulsions, a level of approx. 10⁵–10⁶ L. monocytogenes per cm² was reached after 2–3 days at 20 °C. In this case, 2–3 log were removed or killed by alkaline cleaning products (MC103 or FC140). Direct use of a peracetic acid based disinfectant (Oxivit Active Plus) killed all bacteria when attached in salmon emulsions, whereas only 1–2 log were removed or killed in the meat emulsion system. Thus, the efficiency of cleaning and disinfection products against L. monocytogenes is strongly influenced by the food matrix.     

Disinfectant action of Cymbopogon sp. essential oils in different phases of biofilm formation by Listeria monocytogenes on stainless steel surface

Disinfectant solutions based on the essential oils of Cymbopogon citratus (D.C.) Stapf. and Cymbopogon nardus (L.) Rendle, applied alone or in combination, and a control disinfectant solution were tested in two phases (3 and 240 h) of biofilm formation by Listeria monocytogenes ATCC 19117 on AISI 304 (#4) stainless steel surface. Disinfectant solutions based on essential oils have effectively reduced the number of surface-adhered cells, especially after 60 min of contact. The disinfectant solutions based on a combination of essential oils were capable of reducing 100% (5.64 Log CFU cm⁻²) the number of surface-adhered cells after 60 min of contact, at 240 h of biofilm formation. Essential oils of C. citratus and C. nardus, alone or in combination, are new alternatives for disinfection of industrial stainless steel surfaces contaminated by L. monocytogenes.     

Production of biofilm by Listeria monocytogenes in different materials and temperatures

Listeria monocytogenes, considered as one of the most important foodborne pathogens, is easily found on surfaces, particularly in the form of a biofilm. Biofilms are aggregates of cells that facilitate the persistence of these pathogens in food processing environments conferring resistance to the processes of cleaning and may cause contamination of food during processing, thus, representing a danger to public health. Little is known about the dynamics of the formation and regulation of biofilm production in L. monocytogenes, but several authors reported that the luxS gene may be a precursor in this process. In addition, the product of the inlA gene is responsible for facilitating the entry of the microorganism into epithelial cells that express the receptor E-cadherin, also participates in surface attachment. Thus, 32 strains of L. monocytogenes isolated from different foods (milk and vegetables) and from food processing environments were analyzed for the presence of these genes and their ability to form biofilms on three different surfaces often used in the food industry and retail (polystyrene, glass and stainless steel) at different temperatures (4, 20 and 30 °C). All strains had the ilnA gene and 25 out of 32 strains (78.1%) were positive for the presence of the luxS gene, but all strains produced biofilm in at least one of the temperatures and materials tested. This suggests that genes in addition to luxS may participate in this process, but were not the decisive factors for biofilm formation. The bacteria adhered better to hydrophilic surfaces (stainless steel and glass) than to hydrophobic ones (polystyrene), since at 20 °C for 24 h, 30 (93.8%) and 26 (81.3%) produced biofilm in stainless steel and glass, respectively, and just 2 (6.2%) in polystyrene. The incubation time seemed to be an important factor in the process of biofilm formation, mainly at 35 °C for 48 h, because the results showed a decrease from 30 (93.8%) to 20 (62.5%) and from 27 (84.4%) to 12 (37.5%), on stainless steel and glass, respectively, although this was not significant (p = 0.3847). We conclude that L. monocytogenes is capable of forming biofilm on different surfaces independent of temperature, but the surface composition may be important factor for a faster development of biofilm.     

Lactobacillus sakei 1 and its bacteriocin influence adhesion of Listeria monocytogenes on stainless steel surface

Listeria monocytogenes is of particular concern for the food industry due to its psychrotolerant and ubiquitous nature. In this work, the ability of L. monocytogenes culturable cells to adhere to stainless steel coupons was studied in co-culture with the bacteriocin-producing food isolate Lactobacillus sakei 1 as well as in the presence of the cell-free neutralized supernatant of L. sakei 1 (CFSN-S1) containing sakacin 1. Results were compared with counts obtained using a non bacteriocin-producing strain (L. sakei ATCC 15521) and its bacteriocin free supernatant (CFSN-SA). Culturable adherent L. monocytogenes and lactobacilli cells were enumerated respectively on PALCAM and MRS agars at 3-h intervals for up to 12 h and after 24 and 48 h of incubation. Bacteriocin activity was evaluated by critical dilution method. After 6 h of incubation, the number of adhered L. monocytogenes cells in pure culture increased from 3.8 to 5.3 log CFU/cm² (48h). Co-culture with L. sakei 1 decreased the number of adhered L. monocytogenes cells (P < 0.001) during all sampling times with counts lower than 3.0 log CFU/cm². The CFNS-S1 also led to a significant and similar reduction in culturable adhered L. monocytogenes counts for up to 24 h of incubation, however after 48 h of incubation, re-growth of L. monocytogenes number of adhered cells was observed, likely due to lack of competition for nutrients. L. sakei ATCC 15521 or its supernatant (CFNS-SA) did not reduce the number of adhered L. monocytogenes cells on stainless steel surface and from 6 h of incubation, listerial counts were between 4.3 and 4.5 log CFU/cm². These results indicate that L. sakei 1 and its bacteriocin sakacin 1 may be useful to inhibit early stages of L. monocytogenes adherence to abiotic surface.     

Surveillance of listeriosis and its causative pathogen,Listeria monocytogenes

To manage the problem of foodborne listeriosis requires an understanding of the burden of the disease on a worldwide scale as foods that are prone to contamination are eaten widely domestically and many are traded globally. Surveillance of the disease, caused by Listeria monocytogenes, is typically restricted to developed countries, but many of these do not consider listeriosis as a notifiable disease and estimate the numbers by other means. Incidence rates range from 0.3 to 1.3 per 100,000, but most are in the 0.3–0.5 range, irrespective of the regulatory system and industry control programmes that have been in place. Ready-to-eat foods are the vehicle for transmission of the Listeria through contamination somewhere in the food chain. Meat, poultry and dairy products have been most frequently implicated, but other foods including produce may also have been vehicles of transmission. Large outbreaks are usually linked to errors in food processing plants, such as contaminated slicing machines, followed by opportunities for growth of the pathogen. Less is known about home-generated illnesses but incorrect use of refrigerators can allow cross-contamination and growth of the pathogen to levels that can cause infections. In the U.S., door-to-door salesmen have sold contaminated Hispanic soft cheeses that have led to outbreaks and stillbirths. In addition to outbreak investigation, case-control studies, and the use of experts, risk assessments, and food attribution studies can help focus on areas of greatest risk for prevention and control measures throughout the food chain.     

Temperature distribution in Spanish domestic refrigerators and its effect on Listeria monocytogenes growth in sliced ready-to-eat ham

In order to evaluate the increase of L. monocytogenes concentration during home storage, a low level of the pathogen (<10 CFU/g) was inoculated in sliced ham. Two storage temperatures (5 °C and 9 °C) were selected from a previous study in 33 home refrigerators. Three days of storage or less were necessary to reach the regulated limited concentration (100 CFU/g), at both temperatures. When storage time was extended to 5 days, the pathogen achieved risky values (>10³ CFU/g).     

Adaptive response of Listeria monocytogenes to heat and its impact on food safety

Listeria monocytogenes is an important food associated pathogen because of its relatively high heat resistance and ability to multiply in refrigeration temperatures. Its thermotolerance can be increased when its cells are subjected to heat shock. One- to eight-fold increase of D values of L. monocytogenes have been reported, depending on the heat shock duration, the temperature and the heating menstrum. This acquisition of heat tolerance is related to the induction of the synthesis of heat shock proteins (HSPs).The adaptive response of food pathogens has important consequences on the safety of thermally processed foods. It is believed that this is responsible for the frequent occurrence of deviations (tails and shoulders) during heat treatments that are observed in the exponential model of microbial inactivation. These deviations from log-linear kinetic especially encountered under mild heat treatments, mean that prediction of food safety can no longer rely upon D and z values. Adaptive response to heat must be considered when quantifying and modeling microbial inactivation during thermal processing in order to achieve microbiologically safe products without overly conservative heat processes. Therefore a more mechanistic approach is needed for more accurate predictions of thermal inactivation. Prerequisite to this model are thorough studies to understand how L. monocytogenes and other pathogens adapt their cellular physiology to overcome heat and other stresses.     

The challenge of setting risk-based microbiological criteria for Listeria monocytogenes

After more than 20 years of work with discussing the setting of microbiological criteria for Listeria monocytogenes in foods, Codex Alimentarius on Food Hygiene has finalised a proposal that was recently adopted by the Codex Alimentarius Commission. The effort of developing procedures for making the microbiological criteria risk-based to the greatest extent possible has challenged scientists and managers during this long time period. Yet, the establishment of microbiological criteria for L. monocytogenes is still being discussed and several approaches are possible. Setting of microbiological criteria continues to be a risk management decision – even when using mathematical modelling to enhance the scientific level of a risk-based approach.     

The international risk governance council framework and its application to Listeria monocytogenes in soft cheese made from unpasteurised milk

The International Risk Governance Council (IRGC) developed a Risk Governance Framework whose purpose is to help policy makers, regulators and risk managers both understand the concept of risk governance and apply it to their handling of risks. The Framework allows input from the scientific and public health perspective as well as the less-easily measurable side of freedom of choice. For food, the public wants many choices and less regulation, but at the same time safe products to consume. These ideals are at times in conflict. One example of this is the demand for soft cheese made from unpasteurised milk which may be contaminated with L. monocytogenes or other pathogens. Distinguishing between simple, complex, uncertain and ambiguous risks can help to design a risk management strategy. Public health advocates typically see management of soft cheese made from unpasteurised milk as a simple risk (protection of health), whereas for a small but vociferous proportion of the public sees it more of an uncertain or ambiguous one (choice trumps the slight risk of illness). The implications of five management options for the cheese are discussed. The new Codex Guidelines tend to put these cheeses into Option 1, a strict control on the presence of the pathogen, which will be ignored by the cheese aficionados. Other options are worth exploring to give some more choice but at low risk.     

Determination of Listeria monocytogenes growth potential on new fresh salmon preparations

The aim of this work was to evaluate the increase of Listeria monocytogenes on new salmon preparations (salt–sugar–pepper–dill salmon) in comparison with that obtained on cold-smoked salmon. Salmon preparations were inoculated with L. monocytogenes and were analyzed during storage at 4 °C then 8 °C. At 8 °C, the bacteria growth was of 4.53 log CFU g⁻¹ in cold-smoked salmon and of 2.06 log CFU g⁻¹ in salt–sugar–pepper–dill salmon without background microflora. The growth of L. monocytogenes was different in new salmon preparation because the mixture salt–sugar–pepper had an anti-Listeria activity and its presence could inhibitory to the growth. It is difficult to generalize findings observed with cold-smoked salmon to a new salmon preparation.     

A multi-sampling approach to evaluate an infrared surface treatment for reducing Listeria monocytogenes contamination on whole Gorgonzola cheese rinds

The microbial ecology of Gorgonzola cheese rind is the focus of many studies because the surface can be contaminated by pathogenic microorganisms. Among food-borne pathogens, particular attention is focused on the behaviour of Listeria monocytogenes that is able to grow at refrigeration temperatures and it could also grow during ripening. The Consortium for the Protection of Gorgonzola Cheese declares the rind not edible but the pathogen may also be transferred during cutting and portioning. Therefore, the decontamination of rinds is important to increasing cheese safety. To achieve this goal, many different strategies have been proposed. In this study, the application of an infrared surface treatment to decontaminate cheese rinds is proposed. The presence of L. monocytogenes, which was artificially inoculated in cheese rinds together with cheese rind microflora, and the cheese rind microflora were monitored before and after the treatment of 32 samples of Gorgonzola cheese rinds.The infrared surface treatment provided good reduction of the rind microflora, and L. monocytogenes was particularly affected by this. The treatment, applied to cheeses at the end of ripening, does not interfere with the ripening process and offers the advantages of short time exposures and easy installation of the equipment in cheese plants. Moreover, this study demonstrated that the sampling method affects the detection of cheese rind microflora. In fact, a non-destructive sampling method, based on a sponge and often used for surface sampling but never before applied to ready to eat food sampling, was compared with a traditional but destructive method, based on rind scraping. Regarding L. monocytogenes, the sponge method allowed to estimate even only 5.71 ± 0.79 log cfu g⁻¹ of cells reduction after the treatment while the higher reduction when considering the rind scraping method was 4.06 ± 3.38 log cfu g⁻¹. The sponge method, combined with the classic scraping one, besides offering the great advantage of not being destructive, allowed to differentiate the effect that the treatment has on the microflora located on the surface from those in deeper layers.     

Influence of phenolic compounds from wines on the growth of Listeria monocytogenes

The anti-microbial properties against Listeria monocytogenes of pure flavonoids rutin, catechin and quercetin; non-flavonoids gallic, vanillic, protocatechuic and caffeic acids and total polyphenols of three Argentinean wines, Cabernet Sauvignon, Malbec and Merlot varieties were investigated. The non-flavonoid caffeic acid and the flavonoids rutin and quercetin were the compounds with higher inhibitory activities on L. monocytogenes growth. The knowledge of the anti-listerial effect of different wines varieties could be the basis to demonstrate if the wine consumption with a meal may collaborate in the health protection against some foodborne organisms such as L. monocytogenes.     

Influence of the hydrophobicity and surface roughness of mangoes and tomatoes on the adhesion of Salmonella enterica serovar Typhimurium and evaluation of cleaning procedures using surfactin

Salmonellosis is associated with the consumption of raw vegetables and fruits such as tomatoes, watermelons, alfalfa sprouts, radishes, carrots, lettuce and parsley. The influence of the fruits' roughness on bacterial adhesion was evaluated as measured using a profilometer. The adhesion of Salmonella enterica serovar Typhimurium to mango and tomato surfaces was also evaluated by measuring of the hydrophobicity of the microorganisms and the fruits surfaces. The bacteria adherent on fruit's surface was quantified by plate count and visualize by scanning electron microscopy. In addition, the efficiency of surfactin in removing S. Typhimurium from the fruits' surfaces was analyzed. The average roughness (R_a) of mango (4.54 ± 1.95 μm) was significantly different (p < 0.05) compared to tomato (2.88 ± 2.15 μm). The adhesion of the microorganisms to the fruits' surfaces, as predicted by a determination of the total energy of adhesion (ΔG), was thermodynamically unfavorable. Despite these data, the numbers of bacteria on both fruits' surfaces were similar (p > 0.05), reaching 5.95 ± 0.36 log CFU cm⁻² and 5.81 ± 0.39 log CFU cm⁻² on mango and tomato, respectively. Therefore, these results suggest that the adhesion observed in this experiment is a multifactorial process. Surfactin removed 94.3% and 92.2% of the S. Typhimurium adhered to the surfaces of the mangoes and tomatoes, respectively. Our research showed that the roughness and hydrophobicity of the fruits' surface did not affect the efficiency of each sanitation treatments on removing of S. Typhimurium. It was observed that the chlorine was more efficient treatment (p < 0.05) for tomato surface. For surface of mangoes, chorine and surfactin were better than water treatment for bacteria control.     

Effect of potassium sorbate washing on the growth of Listeria monocytogenes on fresh poultry

This work evaluated the effect of potassium sorbate washing on the growth of Listeria monocytogenes on poultry legs stored at 4 °C for 7 days. Fresh inoculated chicken legs were dipped into either a 2.5% (w/v) or 5% potassium sorbate solution or distilled water (control). Changes in mesophiles, pychrotrophic counts and sensorial characteristics (odor, color, texture and overall appearance) were also evaluated.The shelf life of the samples washed with potassium sorbate was extended by at least 2 days over the control samples washed with distilled water. Legs washed with 5% potassium sorbate showed a significant (p < 0.05) inhibitory effect on L. monocytogenes compared to control legs, with a decrease of about 1.3 log units after 7 days of storage. Sensory quality was not adversely affected by potassium sorbate.     

Occurrence of Listeria spp. in Brazilian fresh sausage and control of Listeria monocytogenes using bacteriophage P100

Since the 1980s, an increase in outbreaks of human listeriosis linked to contaminated food has been a concern of health authorities. Intensively manipulated foods, such as Brazilian fresh sausage, are frequently responsible for food-borne diseases. In this work the occurrence of Listeria monocytogenes and the efficacy of bacteriophage P100 (LISTEX?) to control the microorganism was evaluated in Brazilian fresh sausage. Eighty samples were analyzed, 40 each of swine and chicken Brazilian fresh sausage. Listeria spp. were isolated from 12 samples (15%), of which three (3.75%) were positive for L. monocytogenes. L. monocytogenes strains isolated belonged to serotype 1/2a. L. monocytogenes 1/2a was inoculated in Brazilian fresh sausage (2.1 × 10⁴ cfu/g) with the bacteriophage added thereafter (3.0 × 10⁷ pfu/g). Samples were analysed immediately (day zero) and then stored at 4 °C for 10 days. The bacteriophage P100 reduced L. monocytogenes counts by 2.5 log units at both 0 and 10 days compared to controls without bacteriophage. In spite of this, the populations of L. moncytogenes increased over the 10 day storage. Our data demonstrate that in one of the samples the use of the bacteriophage dropped the bacteria count below the level of direct detection. This study demonstrates a new alternative for pathogen control in the food industry, especially in the processes used to produce Brazilian fresh sausage.     

The effect of pulsed UV light on Escherichia coli O157:H7, Listeria monocytogenes,Salmonella Typhimurium,Staphylococcus aureus and staphylococcal enterotoxin A on sliced fermented salami and its chemical quality

Pulsed UV light (PL) applied at a fluence of 3 J/cm² was effective to reduce Listeria monocytogenes, Escherichia coli 0157:H7, Salmonella Typhimurium and Staphylococcus aureus for 2.24, 2.29, 2.25 and 2.12 log CFU/g on the surface of dry fermented salami. Further increase in the fluence of PL treatment did not increase levels of microbial inactivation. However, the time interval between the contamination and PL treatment was found to have a significant impact on the efficacy of PL treatment and should be kept as short as possible. After initial PL treatment slices of fermented salami were packed in vacuum or in 80%CO₂/20%N₂ modified atmosphere and stored at 4 °C to investigate the effect of PL treatment on protein and lipid oxidation as the shelf life of fermented salami is not usually limited by microbial deterioration, but by chemical and sensory alterations. In this study observed lipid oxidation values for PL treated vacuum and modified atmosphere packed fermented salami slices fall within the acceptable threshold for the rancid odor, except for the sample treated with the highest fluence tested (15 J/cm²), packed in modified atmosphere and kept in cold storage for 9 weeks (1.23 mg MDA/kg). All values were below the threshold for rancid flavor, too. The significant rise in protein oxidation of PL treated fermented salami slices, perceived as 28% increase of carbonyl content compared to untreated samples, was observed only after 9 weeks of cold storage in both vacuum and modified atmosphere packed samples. The results of chemical analysis are in agreement with previously published results of sensory analysis. Current results show the applicability of PL to improve microbial safety of sliced fermented salami that are prone to cross-contamination without affecting quality attributes by lipid and protein oxidation.     

A review of minimal and defined media for growth of Listeria monocytogenes

Listeria monocytogenes is known to be an important foodborne pathogen and is the causative agent of one of the deadliest foodborne illnesses. The organism has a wide range of environmental conditions under which it will survive and grow, and often contaminates processing plants and retail environments in which ready-to-eat foods are manufactured, prepared and served. Although L. monocytogenes is not a fastidious organism and can grow in a variety of rich media, defined and minimal media are necessary to elucidate the minimal environmental nutrients that are required for the survival and growth of this organism. In addition, many of the virulence factors required for L. monocytogenes to invade and multiply in mammalian host cells are not produced in rich media and thus induction of these factors is best studied in a minimal medium. This review covers the historical development of minimal media for Listeria spp. and explores the various factors required for survival and growth of this organism. To the best of our knowledge, this is the first time that these media have been compared side by side. In order to better compare studies using different chemical substrates such as a different carbon source, all concentrations of components in each medium have been converted to molar concentrations.     

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.