High voltage atmospheric cold plasma inactivation of Listeria monocytogenes in fresh Queso Fresco cheese.

Listeria (L.) monocytogenes is a significant pathogen found in ready-to-eat meat and dairy products. Soft cheeses, such as Queso Fresco cheese (QFC), are particularly sensitive to Listeria contamination, and occasionally serve as a source of food-borne illness outbreaks. In the present study, clinical and cheese isolates of L. monocytogenes were assayed for phenotypic characteristics following sub-lethal high voltage atmospheric cold plasma (HVACP) treatment. Reductions in biofilm formation, swimming motility, and growth dynamics were observed following HVACP treatment. Microbial enumeration of 1-, 10-, and 100-g fresh QFC following 0, 1, 2, or 3 min of HVACP demonstrated significant reductions in L. monocytogenes after 1 min (P-value <0.05), with increasing efficacy with prolonged exposure. A mass-dependent effect was observed between treatments of 1-, 10-, and 100-g QFC in regard to treatment efficacy. This result indicates that greater L. monocytogenes reduction on a larger QFC mass requires greater exposure of the L. monocytogenes to the reactive gas species. Optical absorption spectroscopy confirmed a reduction in reactive gas species for each log increase in QFC mass, however, an equivalent volume of inert foam resulted in increased reactive gas generation compared to QFC. In conclusion, we demonstrate both the application and limitations of HVACP treatments of QFC in the currently defined experimental parameters.

Detection and genotyping of Listeria monocytogenes in artisanal soft cheeses from Ecuador.

Listeria monocytogenes is one of the most important bacteria associated with foodborne diseases, soft cheese being an important L. monocytogenes vehicle. In Ecuador, soft cheese is consumed in 84.3% of urban households. We determined the prevalence of L. monocytogenes and serogroups in 260 fresh artisanal soft cheese samples collected in 18 of 24 Ecuadorian provinces. Listeria spp. detection was carried out by culture-dependent and independent methods; 14.23% of samples were positive for L. monocytogenes. Serogroup IVb was found in 83.78% of the food isolates. Serogroups IIb and IIa were present in 8.11% of our isolates. To our knowledge, this is the first report of L. monocytogenes serogroups associated with food in Ecuador; we also found serogroup similarities between cheese isolates and clinical isolates.

A model to predict the fate of Listeria monocytogenes in different cheese types - A major role for undissociated lactic acid in addition to pH, water activity, and temperature.

Undissociated lactic acid has been shown to play a major role in complete growth inhibition of Listeria monocytogenes in Gouda cheese. In addition, low water activity conditions may contribute to growth inhibition. In the current study, it was assessed whether the major factors that inhibit growth of L. monocytogenes in Gouda cheese are the factors that determine growth in other types of ready-to-eat cheese as well. Various types of cheeses were selected, some of which had been associated with listeriosis, while others had not. Based on the composition of the different cheese types, the concentrations of undissociated lactic acid were calculated for each type. The ability to support growth of L. monocytogenes was predicted using the Gamma model, based on literature data on total lactic acid content, moisture content, fat content, pH, Aw, and temperature, and optimal growth rates in milk at 30-37 °C. In addition, the actual specific growth rates of L. monocytogenes in the various cheeses were calculated based on available experimental growth data. In 9 out of the 10 RTE cheeses reviewed, the undissociated lactic acid concentrations and aw determined growth/no growth of L. monocytogenes. No growth was correctly predicted for feta, Cheddar and Gouda, and growth was correctly predicted for ricotta, queso fresco, Camembert, high-moisture mozzarella, cottage and blue cheese. Growth of L. monocytogenes was not observed in practice upon inoculation of Emmental, whereas growth in this cheese type was predicted when including the above mentioned factors in the models. Other factors, presumably acetic and propionic acid, are thought to be important to inhibit growth of the pathogen in Emmental. The results from our study show that for cheeses in which lactic acid is a main acid, our model based on undissociated lactic acid, temperature, pH and aw gives a good prediction of potential outgrowth of L. monocytogenes. Implications for L. monocytogenes legislation are discussed per type of RTE cheese reviewed.

Evaluation of Lactose Oxidase as an Enzyme-Based Antimicrobial for Control of L. monocytogenes in Fresh Cheese.

Listeria monocytogenes is a ubiquitous pathogen that can cause morbidity and mortality in the elderly, immune compromised, and the fetuses of pregnant women. The intrinsic properties of fresh cheese-high water activity (aW), low salt content, and near-neutral pH-make it susceptible to L. monocytogenes contamination and growth at various points in the production process. The aim of this study was to investigate the ability of lactose oxidase (LO), a naturally derived enzyme, to inhibit the growth of L. monocytogenes in fresh cheese during various points of the production process. Lab-scale queso fresco was produced and inoculated with L. monocytogenes at final concentrations of 1 log CFU/mL and 1 CFU/100 mL. LO and LO sodium thiocyanate (TCN) combinations were incorporated into the milk or topically applied to the finished cheese product in varying concentration levels. A positive control and negative control were included for all experiments. When L. monocytogenes was inoculated into the milk used for the cheese-making process, by day 28, the positive control grew to above 7 log CFU/g, while the 0.6 g/L treatment (LO and LO + TCN) fell below the limit of detection (LOD) of 1.3 log CFU/g. In the lower inoculum, the positive control grew to above 7 log CFU/g, and the treatment groups fell below the LOD by day 21 and continued through day 28 of storage. For surface application, outgrowth occurred with the treatments in the higher inoculum, but some inhibition was observed. In the lower inoculum, the higher LO and LO-TCN concentrations (0.6 g/L) reduced L. monocytogenes counts to below the LOD, while the control grew out to above 7 log CFU/g, which is a >5 log difference between the control and the treatment. These results suggest that LO could be leveraged as an effective control for L. monocytogenes in a fresh cheese.

Efficacy of nisin derivatives with improved biochemical characteristics, alone and in combination with endolysin PlyP100 to control Listeria monocytogenes in laboratory-scale Queso Fresco.

Nisin is an antimicrobial peptide that is commonly used as a food preservative and capable of inhibiting the pathogen Listeria monocytogenes. However, nisin is ineffective in controlling L. monocytogenes in Queso Fresco (QF). To address the challenge, in this work, we used synthetic biology strategies to create a series of nisin A derivatives by substituting residues 27, 30, 31 and 32 with positively charged amino acids (H, K and R). Our results showed that nisin derivatives exhibited reduced antilisterial activity in vitro compared to nisin A; however, they were all more stable under QF-like experimental conditions (pH 7 + 22% milk fat), notably H27/31K. Compared to nisin A, the derivatives H31K and V32K exhibited slight antilisterial improvement in QF and H27/31K was able to reduce the initial population of L. monocytogenes by up to 1.5 Log CFU/g. L. monocytogenes isolates exhibited similar susceptibility to nisin A or H27/31K after 7 or 14 days of nisin exposure in QF. Notably, when combined with endolysin PlyP100, the application of H27/31K resulted in non-enumerable levels of L. monocytogenes after 14 days of cold storage. Our results highlight the potential of bioengineered nisin derivatives for stabilized and enhanced control of L. monocytogenes in QF.

Evaluation of combinations of nisin, lauric arginate, and ε-polylysine to control Listeria monocytogenes in queso fresco.

Listeria monocytogenes contamination is of great concern in queso fresco (QF), and listeriosis outbreaks linked to consumption of QF continue to happen. Hurdle approaches such as combining antimicrobials provide an alternative to improve QF safety. In this work, the efficacy of antimicrobial combinations of nisin (NIS), lauric arginate (LAE), and ε-polylysine (EPL) to inhibit L. monocytogenes growth in QF was evaluated. First, antimicrobials were screened for potential synergy in vitro. Later, antimicrobial treatments were challenged in QF inoculated with ∼4 log10 cfu/g of L. monocytogenes 5-strain cocktail and stored for 28 d at 4°C. Our results showed that combinations of NIS-LAE and EPL-LAE were synergistic in vitro. Limited antilisterial control was observed in QF treated with NIS, LAE, and NIS-LAE; however, EPL and EPL-LAE exhibited listeristatic effect in QF for up to 14 and 28 d of cold storage, respectively. Additionally, L. monocytogenes QF isolates had similar susceptibility to EPL or LAE. A consumer panel was able to distinguish between QF added with EPL (250 µg/g) + LAE (66.66 µg/g) and control QF, most likely associated with manufacturing and storage rather than antimicrobials' taste. Our results support the use of EPL-LAE combination to control L. monocytogenes growth in QF.

Evaluation of the efficacy of commercial protective cultures against mold and yeast in queso fresco.

In this study, we evaluated the efficacy of 3 commercial protective cultures designated PC1 (Lactobacillus spp.), PC2 (Lactobacillus rhamnosus), and PC3 (Lactobacillus rhamnosus) as biopreservatives in queso fresco (QF) against 9 yeast strains (Candida zeylanoides, Clavispora lusitaniae, Debaryomyces hansenii, Debaryomyces prosopidis, Kluyveromyces marxianus, Meyerozyma guilliermondii, Pichia fermentans, Rhodotorula mucilaginosa, and Torulaspora delbrueckii) and 11 mold strains (Aspergillus cibarius, Aureobasidium pullulans, Penicillium chrysogenum, Penicillium citrinum, Penicillium commune, Penicillium decumbens, Penicillium roqueforti, Mucor genevensis, Mucor racemosus, Phoma dimorpha, and Trichoderma amazonicum). All fungal spoilage strains were previously isolated from dairy processing environments. A positive control (C) with no protective culture was included. Fungal spoilage organisms were inoculated on cheese surfaces at an inoculum level of 20 cfu/g, and cheeses were stored at 6 ± 2°C throughout the study. For yeast enumeration, cheeses were sampled on d 0, 7, 14, and 21 postinoculation. Significant inhibition was detected for each yeast strain by comparing yeast counts for each cheese treated with protective culture against the control cheese using one-way ANOVA with Bonferroni correction performed individually at d 7, 14, and 21 postinoculation. Mold growth was visually observed and imaged weekly through 70 d postinoculation. Whereas PC3 inhibited Cl. lusitaniae, Mey. guilliermondii, and Ph. dimorpha, PC2 inhibited the outgrowth of Cl. lusitaniae, D. hansenii, and Ph. dimorpha. Protective culture 1 had the broadest spectrum of efficacy across yeast and molds, delaying spoilage caused by 4 distinct yeast strains (Cl. lusitaniae, D. hansenii, D. prosopidis, and Mey. guilliermondii), and inhibiting visible growth of 2 mold strains (P. chrysogenum and Ph. dimorpha). Results demonstrated that commercial protective cultures vary in performance, as indicated by the breadth of mold and yeast inhibition at both the genus and species level. This study suggests that manufacturers looking into using protective cultures should investigate their efficacy against specific fungal strains of concern.

Evaluation of single and combined antimicrobial treatments to inhibit Salmonella in queso fresco.

Hispanic style soft non-fermented cheeses, such as queso fresco (QF) have been linked to outbreaks and recalls. Salmonella is one of the main causes of these incidents. Due to lack of ripening or post-processing antimicrobial treatments, incorporating GRAS antimicrobials to production process may be a suitable approach to minimize microbial risk in QF. The aim of this study was to evaluate the efficiency of nisin (N), caprylic acid (CA) and trans-cinnamaldehyde (CN) as single or combined treatments to reduce Salmonella populations in QF during storage. Batches of QF were inoculated after curding with approx. 4 Log CFU/g of 5-strain cocktails of Salmonella and stored at 8 °C for 20 days. The final Salmonella counts in control samples ranged from 6.96 to 7.14 Log CFU/g. Application of CN at 0.6 g/kg inhibited Salmonella growth during storage, resulting in at least 3 Log CFU/g difference with the untreated controls (p < 0.05). Addition of N (0.5 g/kg) and CA (0.4 g/kg) with CN (0.3 and 0.6 g/kg) further enhanced the antimicrobial activity resulting in complete suppression of growth and even caused a 1 Log CFU/g reduction by the end of the experimental period compared to initial counts. Samples treated with the combined treatment (N, CA, CN) were evaluated in a consumer panel (n = 112). Participants preferred the control and commercial QF to the treated samples. However, treated samples with 0.3 g/kg CN were still within the acceptable range of neutral to like slightly. Results obtained, revealed that combined treatment of N, CA and CN can provide a solution to reduce the count of Salmonella in QF, whether in process or during storage.

Predictive Model of Listeria monocytogenes Growth in Queso Fresco.

Listeria monocytogenes is a hardy psychrotrophic pathogen that has been linked to several cheese-related outbreaks in the United States, including a recent outbreak in which a fresh cheese (queso fresco) was implicated. The purpose of this study was to develop primary, secondary, and tertiary predictive models for the growth of L. monocytogenes in queso fresco and to validate these models using nonisothermal time and temperature profiles. A mixture of five strains of L. monocytogenes was used to inoculate pasteurized whole milk to prepare queso fresco. Ten grams of each fresh cheese sample was vacuum packaged and stored at 4, 10, 15, 20, 25, and 30°C. From samples at each storage temperature, subsamples were removed at various times and diluted in 0.1% peptone water, and bacteria were enumerated on Listeria selective agar. Growth data from each temperature were fitted using the Baranyi model as the primary model and the Ratkowsky model as the secondary model. Models were then validated using nonisothermal conditions. The Baranyi model was fitted to the isothermal growth data with acceptable goodness of fit statistics (R2 = 0.928; root mean square error = 0.317). The Ratkowsky square root model was fitted to the specific growth rates at different temperatures (R2 = 0.975). The tertiary model developed from these models was validated using the growth data with two nonisothermal time and temperature profiles (4 to 20°C for 19 days and 15 to 30°C for 11 days). Data for these two profiles were compared with the model prediction using an acceptable prediction zone analysis; >70% of the growth observations were within the acceptable prediction zone (between -1.0 and 0.5 log CFU/g). The model developed in this study will be useful for estimating the growth of L. monocytogenes in queso fresco. These predictions will help in estimation of the risk of listeriosis from queso fresco under extended storage and temperature abuse conditions.

Análisis fisicoquímico y sensorial de queso fresco con reemplazo de grasa por lípidos de aguacate (Persea americana Mill V. Hass) / Physico-chemical and sensory analysis of fresh cheese with replacement of milk fat by avocado lipids (Persea americana Mill V. Hass)

RESUMEN Una tendencia actual en la elaboración de quesos es sustituir la grasa láctea por su alto contenido de ácidos grasos saturados (AGS), con lípidos de origen vegetal y el aguacate por su alto contenido de ácidos grasos insaturados(AGI), principalmente, el ácido oleico y fitosteroles (campesterol, β- Sitosterol), ha sido valorado al promover beneficios saludables al consumidor. El objetivo de este estudio fue evaluar, con jueces, el agrado o desagrado, mediante prueba hedónica, los quesos elaborados con sustitución de grasa y analizar el perfil de textura (TPA). Se analizaron los quesos elaborados con tres niveles de pulpa, mediante prueba hedónica de 5 puntos, con 109 jueces y el análisis de perfil de textura con texturometro, modelo TA-XT2i. El T4, con el 18% de pulpa adicionada, fue el que más agradó, de acuerdo con la prueba DMS, al determinar la aceptación de los quesos, aunque no se encontró diferencias entre el T3 y el T4 y, el T2, el que menos gustó. En el análisis de perfil de textura, se encontró diferencias significativas respecto al control; la pulpa de aguacate, como sustituto de grasa en los quesos, generó bajos valores en los parámetros texturales; los quesos retienen alta humedad y bajo contenido de grasa, obteniendo un producto desmoronable, debido a la modificación de la matriz de la caseína, que es la que da rigidez a los quesos.

ABSTRACT A current trend in cheese making is to substitute milk fat for its high content of saturated fatty acids (SFA), with lipids of vegetable origin and avocado for its high content of unsaturated fatty acids (AGI) mainly oleic acid and phytosterols. (campesterol, β-Sitosterol), has been valued by promoting healthy benefits to the consumer. The objective of this study was to evaluate with judges the liking or dislike of the cheeses made with fat replacement and to analyze the texture profile (TPA). The cheeses elaborated with three levels of avocado pulp were analyzed by means of a 5point hedonic test with 109 judges and the analysis of texture profile with texturometer model TA-XT2i. The T4 with the 18% added pulp was the one liked most according to the DMS test when determining the acceptance of the cheeses, although no differences were found between the T3 and the T4; the T2 was the least liked product. In the texture profile analysis, significant differences were found with respect to the control; the avocado pulp as a substitute for fat in the cheeses generated low values in the textural parameters, the cheeses retain high humidity and low fat content, obtaining a crumbly product, due to the modification of the casein matrix which is the one that gives rigidity to the cheeses.

Comparación de tres métodos de recuento de Staphylococcusaureus en queso fresco de expendio en los mercados populares de la ciudad de La Paz-Bolivia / Comparison of three methods of counting Staphylococcus aureus in fresh cheese for sale in the central markets of the city of La Paz-Bolivia

Staphylococcus aureus puede contaminar una gran gama de alimentos, constituyéndose el queso fresco en un buen medio diferencial y selectivo para el desarrollo de este microorganismo. La intoxicación estafilocóccica trasmitida por alimentos, resulta de la ingesta de enterotoxina termoestable preformada en el alimento que fue generada por una cepa toxigénica de Staphylococcus aureus llegando a constituirse en un riesgo para la salud de los consumidores. El presente estudio tiene el objetivo de comparar tres métodos: i) ISO 6888-1:2003 ii) NB 32004:2004 y iii) Placas secas rehidratables (Placas Petrifilm) para el recuento de Staphylococcus aureus en queso fresco de expendio en los mercados populares de la ciudad de La Paz. La comparación entre el método de cultivo convencional empleando medio Agar Baird Parker y el método alternativo por placa seca rehidratable fue hecha por medio del muestreo al azar de 30 muestras de queso fresco, el análisis fue hecho por los tres métodos métodos, donde por el método de placa seca rehidratable 10 muestras se encontraban dentro de norma y 20 se encontraban fuera de norma; por medio del método de cultivo convencional 22 muestras se encontraban en norma y 8 fuera de norma. Concluyendo que el método de placa seca rehidratable es más sensible y fácil de aplicar en comparación con el método convencional según norma Bolivia y norma ISO.

Staphylococcus aureus can contaminate a wide range of foods, constituting fresh cheese in a good differential and selective medium for the development of this microorganism. Staphylococcal foodborne poisoning results from the intake of thermostable enterotoxin preformed in the food that was generated by a toxigenic strain of Staphylococcus aureus, becoming a risk to the health of consumers. The present study has the objective of comparing three methods: i) ISO 6888-1: 2003 ii) NB 32004: 2004 and iii) Dry rehydratable plates (Petrifilm plates) for the recount of Staphylococcus aureus in fresh cheese for sale in popular markets from the city of La Paz. The comparison between the conventional culture method using Baird Parker Agar medium and the alternative method by rehydratable dry plate was made by random sampling of 30 samples of fresh cheese, the analysis was done by the three methods methods, where by the method dry rehydratable plate 10 samples were within norm and 20 were out of norm; By means of the conventional cultivation method, 22 samples were in standard and 8 out of norm. Concluding that the rehydratable dry plate method is more sensitive and easier to apply compared to the conventional method according to the Bolivia standard and ISO standard.

Effect of modified atmosphere packaging on the growth of spoilage microorganisms and Listeria monocytogenes on fresh cheese.

Queso Fresco has a limited shelf life and has been shown to support the rapid growth of Listeria monocytogenes during refrigerated storage. In addition to improving quality and extending shelf life, modified atmosphere packaging (MAP) has been used to control the growth of pathogenic microorganisms in foods. The objectives of this study were to determine the effects of MAP conditions on the survival and growth of spoilage microorganisms and L. monocytogenes during storage of Queso Fresco manufactured without starter cultures. For L. monocytogenes experiments, cheeses were surface inoculated at ∼4 log10 cfu/g before packaging. Inoculated and uninoculated (shelf life experiments) cheeses were placed in 75-µm high-barrier pouches, packaged under 1 of 7 conditions including air, vacuum, or combinations of N2 and CO2 [100% N2 (MAP1), 30% CO2:70% N2 (MAP2), 50% CO2:50% N2 (MAP3), or 70% CO2:30% N2 (MAP4), 100% CO2 (MAP5)], and stored at 7°C. Samples were removed weekly through 35 d of storage. Listeria monocytogenes counts were determined for inoculated samples. Uninoculated samples were assayed for mesophilic and psychrotolerant counts, lactic acid bacteria, coliforms, and yeast and mold. In general, cheeses packaged under conditions consisting of higher contents of CO2 had lower pH levels during storage compared with those stored in conditions with lower levels or no CO2 at all. Similarly, the antimicrobial efficacy of MAP in controlling spoilage microorganisms increased with increasing CO2 content, whereas conditions consisting of 100% N2, vacuum, or air were less effective. Mean L. monocytogenes counts remained near inoculation levels for all treatments at d 1 but increased ∼2 log10 cfu/g on cheeses packaged in air, vacuum, and 100% N2 (MAP1) conditions at d 7 and an additional ∼1.5 log10 cfu/g at d 14 where they remained through 35 d. In contrast, treatments consisting of 70% CO2 (MAP4) and 100% CO2 (MAP5) limited increases in mean L. monocytogenes counts to <1 log10 cfu/g through 14 d and ∼1.5 log10 cfu/g by d 21. Mean L. monocytogenes counts increased to levels significantly higher than inoculation (d 0) on cheeses stored in MAP2 and MAP3 on d 21, on d 28 for MAP4, and on d 35 for cheeses stored under MAP5 conditions. Overall, significant treatment × time interactions were observed between air, vacuum, and MAP1 when each was compared with MAP2, MAP3, MAP4, and MAP5. These data demonstrate that packaging fresh cheese under modified atmospheres containing CO2 may be a promising approach to extend shelf life while limiting L. monocytogenes growth during cold storage.

Microbial analysis of commercially available US Queso Fresco.

Queso Fresco (QF), a fresh Hispanic-style cheese, is often associated with Listeria monocytogenes outbreaks and recalls. Queso Fresco's susceptibility to bacterial contamination is partially due to its high pH and moisture content as well as Listeria's tolerance for the salt content typical for QF. Nine different brands of US QF, 2 packages from 4 different lots (to account for temporal variability), were sampled. The pH, salt content, and moisture content were analyzed in addition to microbial testing including yeasts and molds, coliforms, lactic acid bacteria enumeration, and L. monocytogenes counts. The cheeses were also inoculated with a cocktail of 5 food and human isolates of food-borne outbreak-associated Listeria monocytogenes strains to evaluate how the differences between brands influenced Listeria growth. Three of the cheeses underwent additional genus-level microbial analysis using extracted 16S rDNA, allowing for phylogenetic analysis between bacterial taxa including diversity and relative abundance. We found little variation between the sampled QF pH (range = 6.62-6.86), salt content (1.53-2.01%), and moisture content (43.90-54.50%). Yeasts and molds were below the detection limit of enumeration in all of the cheeses and coliforms were below the detection limit across the first 3 lots, but were detected at varying levels in the fourth lot (>3.0 most probable number/g) for 3 of the brands. Listeria monocytogenes was not isolated after enrichment in any of the samples. All cheeses tested positive for the presence of lactic acid bacteria, with only 1 of the cheeses being labeled as produced with added cultures having substantial counts. Fourteen days after inoculation with L. monocytogenes, at least 2.5 log10 cfu/g of growth was found for all QF brands stored at 4°C. Microbial genus analysis showed that, among the 3 brands, the microbial community was more similar within brand than when compared with the other 2 brands. Thermus, Anoxybacillus, and Streptococcus accounted for the dominant genera of brands A, B, and C, respectively. These variations within the microbial community may account for sensory differences and help manufacturers determine quality control consistency more readily than culture-based methods.

Evaluación de los indicadores de desempeño de 3M Petrifilm Staph Express (STX) frente a la norma ISO 6888-1: 2003 en el recuento de Staphylococcus aureus en quesos frescos por contaminación artificial / Evaluation of the performance indicators of 3M Petrifilm Staph Express (STX) against the ISO 6888-1: 2003 standard in the count of Staphylococcus aureus in fresh cheeses by artificial contamination

Staphylococcus aureus puede contaminar una gran gama de alimentos, siendo los quesos frescos un medio diferencial y selectivo para el desarrollo de este microorganismo, llegando a la producción de enterotoxina termoestable ocasionando una intoxicación estafilocóccica trasmitida por alimentos. El empleo del método 3M Petrifilm Staph Express (STX) supone considerables ventajas frente al método convencional o de referencia, facilitando y favoreciendo muchos aspectos técnicos y económicos, dentro de ellos tenemos la optimización de tiempo en cuanto se refiere a la preparación de medios de cultivo, necesidad de equipamiento, requerimiento de infraestructura, menor requerimiento de personal y la disminuye los costos del uso de consumo eléctrico, disminuye la generación de residuos sólidos, entre otros. La comparación entre el método alternativo frente al método convencional fue hecha por medio de la contaminación artificial de una matriz libre de S. aureus y un recuento bajo de microbiota acompañante, donde se establecieron diferentes niveles de contaminación, evaluándose los indicadores de desempeño (exactitud relativa, precisión relativa, linealidad, curva de calibración y límite de cuantificación). En indicador de exactitud relativa se obtuvo un 95,6% de recuperación para el nivel más bajo (aproximadamente 8 UFC/g) y 98,7% de recuperación para el nivel de contaminación más alto (aproximadamente 5300 UFC/g), teniendo un rango de 70 a 120% de aceptación. Para la precisión relativa se calcularon la RSD de todos los niveles ensayados, obteniéndose datos que aceptan la precisión relativa del método de placa seca rehidratable porque los valores fueron menores al RSD teórico calculado. El límite de detección fue ensayado con <10UFC/g obteniéndose un CV de 1,16%. Por los resultados obtenidos podemos concluir que el método 3M Petrifilm Staph Express (STX) cumple con los indicadores de desempeño frente a la norma ISO 6888-1:2003.

Staphylococcus aureus can contaminate a wide range of foods, being fresh cheeses a differential and selective medium for the development of this microorganism, reaching the production of thermostable enterotoxin causing a staphylococcal intoxication transmitted by food. The use of the 3M Petrifilm Staph Express (STX) method has considerable advantages over the conventional or reference method, facilitating and favoring many technical and economic aspects, within them we have the optimization of time as regards the preparation of culture media, need for equipment, infrastructure requirements, lower personnel requirements and decreases the costs of the use of electricity consumption, decreases the generation of solid waste, among others. The comparison between the alternative method and the conventional method was made by means of the artificial contamination of a free matrix of S. aureus and a low accompanying microbiota count, where different levels of contamination were established, evaluating the performance indicators (relative accuracy, relative precision, linearity, calibration curve and limit of quantification). In the relative accuracy indicator, 95.6% recovery was obtained for the lowest level (approximately 8 CFU / g) and 98.7% recovery for the highest level of contamination (approximately 5300 CFU / g), with a range of 70 to 120% acceptance. For the relative precision, the RSD of all the tested levels was calculated, obtaining data that accept the relative accuracy of the dry rehydratable plate method because the values were lower than the calculated theoretical RSD. The limit of detection was tested with <10 CFU /g, obtaining a CV of 1.16%. Based on the results obtained, we can conclude that the 3M Petrifilm Staph Express (STX) method complies with the performance indicators against the ISO 6888-1: 2003 standard.

Antimicrobial behavior of phage endolysin PlyP100 and its synergy with nisin to control Listeria monocytogenes in Queso Fresco.

Hispanic-style fresh cheeses, such as Queso Fresco (QF), have been linked to numerous listeriosis outbreaks in the United States. In this work, we have studied the antilisterial behavior and effectiveness of the Listeria phage endolysin PlyP100 in QF, as well as the potential synergy between PlyP100 and nisin. PlyP100 showed similar bacterial reduction regardless of varying L. monocytogenes inoculum size in QF, and when the inoculation size was 1 Log CFU/g, no pathogen recovery after cheese enrichment was observed. PlyP100 was stable in QF for up to 28 days of cold storage exhibiting similar antilisterial activity regardless of when contamination with L. monocytogenes occurred. PlyP100 alone exhibited a strong listeriostatic effect in QF, on the contrary, nisin alone was not effective to control the pathogen in QF during cold storage. The combination of nisin and PlyP100 showed a strong synergy in QF with non-enumerable levels of L. monocytogenes after 4 weeks of refrigerated storage. Moreover, L. monocytogenes isolates from cheeses treated with nisin, PlyP100, and their combination did not develop resistance to nisin or PlyP100. Our results support the use of PlyP100 combined with nisin as an efficient L. monocytogenes control measure in QF.

Efficacy of Antimicrobials Applied Individually and in Combination for Controlling Listeria monocytogenes as Surface Contaminants on Queso Fresco.

Outbreaks of listeriosis are continually attributed to the consumption of Hispanic-style soft cheeses contaminated with Listeria monocytogenes postpasteurization. Once contaminated, L. monocytogenes can grow rapidly in cheeses like Queso Fresco (QF) even when stored at refrigeration temperatures. Several antimicrobials, including acidified calcium sulfate with lactic acid (ACSL), ε-polylysine (EPL), hydrogen peroxide (HP), lauric arginate ethyl ester (LAE), and sodium caprylate (SC), have demonstrated antilisterial activity in food. The objectives of this study were to determine the efficacy of these antimicrobials used individually and in combination to control L. monocytogenes as surface contaminants on QF and to identify additive and synergistic interactions. Cheeses were surface inoculated at ∼4 log CFU/g, dipped in antimicrobial solutions, vacuum packaged, and then stored at 7°C for 35 days. L. monocytogenes counts were determined 24 h after application of the antimicrobials and then weekly throughout storage. Dip treatments in a 5% (v/v) HP solution reduced L. monocytogenes counts to <0.5 log CFU/g within 24 h with no increase in counts through day 35. Dip treatments in LAE at 2 and 5% alone and in combination with EPL at 10% produced initial reductions in pathogen counts (1.5 to 1.8 CFU/g) but did not inhibit pathogen growth compared with the sterile water control. Dip applications of ACSL at 25% also produced an initial ∼1.5-log reduction in L. monocytogenes counts followed by regrowth. Application of SC at 10% alone and in combination with either EPL or LAE inhibited growth to <1 log CFU/g through 21 days of storage. The combination of ACSL+SC worked synergistically to inhibit the growth of L. monocytogenes on QF to <1 log CFU/g through 35 days. These data indicate that HP alone and treatments containing EPL, LAE, or ACSL in combination with SC are promising postlethality treatments and process controls for L. monocytogenes on QF through a 21-day shelf life.

Modelo de quesería artesanal bajo un signo distintivo en el Caribe colombiano: caso Atlántico

Introducción. Se presenta una propuesta de signo distintivo para la producción y comercialización de queso costeño de alto impacto en el departamento del Atlántico y en la región Caribe colombiana. Objetivo. Proponer a los productores de queso costeño, un modelo de producción y comercialización del producto, que siente las bases para convertir la quesería artesanal en un instrumento de desarrollo regional, a partir de la fabricación de un producto inocuo de base biotecnológica. Materiales y métodos. Estudio de tipo descriptivo abordado desde los aspectos socio-cultural y biotecnológico, para la identificación de valores colectivos relacionados con tradición quesera, preferencias de producción, asociatividad, cumplimiento de normas legales, signo distintivo y uso de cultivos lácticos autóctonos en sus procesos productivos para incorporar características del queso típico de la región. Resultados. Mediante el desarrollo de un signo distintivo es posible no solo el posicionamiento del queso costeño en el mercado como producto de alto valor, sino dejar atrás la indiferenciación actual propia de un producto básico, y lograr la reducción de riesgo de contaminación, teniendo en cuenta el valor agregado que representan los inóculos microbianos como innovación biotecnológica en la calidad del producto. Conclusión. El modelo presentado convenció al productor de que bajo el signo distintivo su producto tiene mayor valor agregado y, en consecuencia, podrá obtener un precio mayor al actual.

Introduction. A distinctive sign proposal is presented for high-impact production and marketing of coast cheese in the state of Atlántico and in the Colombian Caribbean region. Objective. To propose coast cheese producers a production and marketing model of the product; a model which lays the foundation to covert craft cheese factories into a regional development instrument from the manufacture of a biotechnological-based innocuous product. Materials and methods. Descriptive research addressed from social-cultural and biotechnological aspects for the identification of collective values associated to cheese-making tradition, production preferences, association capacity, compliance with legal regulations, distinctive sign, and use of native dairy cultures in their production processes in order to incorporate characteristics of the typical cheese of the region. Results. Through the development of a distinctive sign, it is possible not only to position the coast cheese in the market as a high-value product but also to left behind the present non-differentiation typical of a basic product and to achieve the decrease of contamination risks, bearing into account the value added brought by microbial inoculation as biotechnological innovation in the product quality. Conclusion. The model shown has convinced the producers that under the distinctive sign their product has better value added and then they would be able to gain a higher price.

Introdução. Se apresenta uma proposta de signo distintivo para a produção e comercialização de queijo litorâneo de alto impacto no departamento do Atlântico e na região do Caribe colombiano. Objetivo. Propor aos produtores de queijo litorâneo, um modelo de produção e comercialização do produto, que sente as bases para converter a indústria do queijo artesanal num instrumento de desenvolvimento regional, a partir da fabricação de um produto inócuo de base biotecnológica. Materiais e métodos. Estudo de tipo descritivo abordado desde os aspectos sociocultural e biotecnológico, para a identificação de valores coletivos relacionados com a tradição da indústria do queijo, preferências de produção, associatividade, cumprimento de normas legais, signo distintivo e uso de cultivos lácteos autóctones nos seus processos produtivos para incorporar características do queijo típico da região. Resultados. Mediante o desenvolvimento de um signo distintivo é possível não só o posicionamento do queijo litorâneo no mercado como produto de alto valor, senão deixar atrás a indiferença atual própria de um produto básico, e conseguir a redução do risco de contaminação, tendo em conta o valor agregado que representam os inócuos microbianos como inovação biotecnológica na qualidade do produto. Conclusão. O modelo apresentado convenceu ao produtor de que sob o signo distintivo seu produto tem maior valor agregado e, por consequência, poderá obter um preço maior ao atual.

Antimicrobial treatments to control Listeria monocytogenes in queso fresco.

Queso fresco, is a Hispanic non-fermented cheese highly susceptible to contamination with L. monocytogenes. This research was aimed to determine the effect of GRAS antimicrobial ingredients to control L. monocytogenes. Antimicrobials included caprylic acid (CA), Nisaplin® (N, 2.5% nisin), a mixture of sodium lactate and sodium diacetate (SL/SD), Lactococcus lactis sbp. lactis DPC 3147, monolaurin, and lactic acid (LA). Batches of queso fresco curds were inoculated with 104 CFU/g and stored at 4 °C for three weeks. During storage the count of L. monocytogenes reached 7 to 8 Log CFU/g in control samples. Most individual antimicrobial treatments resulted in less than 1 Log CFU/g reductions in final counts, with the exception of N (0.5 g/kg) and CA (2.9 g/kg) that caused more than 3 and 5 Log CFU/g differences with controls, respectively. Mixtures of ingredients were more effective in inhibiting L. monocytogenes growth, and treatments with N and CA consistently delivered 6 Log CFU/g less counts than controls. Supplementation of 12 g/kg LA to treatments with SL/SD (3%/0.22%) caused differences of more than 4 Log CFU/g in final Listeria populations. Samples treated with the binary mixtures of N and CA (0.5 and 0.7 g/kg, respectively) were evaluated in a consumer panel (n = 67). Panelists slightly preferred control and commercial over treated samples, but all samples were in average rated between "slightly liking" and "moderately liking." These experiments indicated that combined use of antimicrobial ingredients may be an effective way to control the population of Listeria monocytogenes in queso fresco.

Brucelosis: zoonosis de importancia en México / Brucellosis: a zoonosis of importance in Mexico

Introduction: Brucellosis is one of the most frequent zoonosis in most parts of the world. This zoonosis remains a great problem to public health in developing countries, although developed countries have successfully controlled it. Mexico still shows a high annual brucellosis incidence in humans; thus, the country is considered around the world as an endemic brucellosis country. Aim: To describe the connection/association between this zoonosis and the current epidemiological situation in the Mexican population. Methods: Perusal of research reports, epidemiological studies and veterinarian reviews performed in Mexico, using data bases such as PubMed, Thompson Reuters, Mesh research. Conclusion: The risk of infection by Brucella in Mexico is associated with the consumption of unpasteurized dairy products, mainly fresh cheeses.

Introducción: La brucelosis es una de las enfermedades zoonóticas más frecuentes en la mayor parte del mundo. Mientras que en los países desarrollados han logrado con éxito su control, en los países en vías de desarrollo continúa siendo un gran problema de salud pública. México continúa presentando alta incidencia anual de brucelosis en humanos, por lo que es considerado un país endémico de brucelosis. Objetivo: Describir la relación de esta zoonosis con la situación epidemiológica actual en la población de México. Material y Métodos: Consulta de reportes de investigación, estudios epidemiológicos y revisiones veterinarias, realizadas en México, a través de bases de datos como PubMed, Thompson Reuters y Meshresearch. Conclusión: El riesgo del contagio de Brucella spp. en México está asociado al consumo de productos lácteos sin pasteurizar, principalmente quesos frescos.

Use of a miniature laboratory fresh cheese model for investigating antimicrobial activities.

Hispanic-style fresh cheeses, such as queso fresco, have relatively low salt content, high water activity, and near neutral pH, which predisposes them to growth of Listeria monocytogenes. Biosafety constraints limit the incorporation of L. monocytogenes into cheeses manufactured via traditional methods in challenge studies, so few have focused on in situ testing of novel antimicrobials in fresh cheeses. We have developed a modular, miniaturized laboratory-scale queso fresco model for testing the incorporation of novel antilisterials. We have demonstrated the assessment of the antilisterials nisin and ferulic acid, alone and in combination, at various levels. Our results support the inhibitory effects of ferulic acid in cheese, against both L. monocytogenes and its common surrogate Listeria innocua, and we provide preliminary evaluation of its consumer acceptability.