Recent developments in low-moisture foods: microbial validation studies of thermal pasteurization processes.

Outbreaks associated with low-moisture foods (e.g., wheat flour, nuts, and cereals) have urged the development of novel technologies and re-validation of legacy pasteurization process. For various thermal pasteurization processes, they share same scientific facts (e.g., bacterial heat resistance increased at reduced water activity) and guidelines. However, they also face specific challenges because of their different heat transfer mechanisms, processing conditions, or associated low-moisture foods' formulations. In this article, we first introduced the general structural for validating a thermal process and the shared basic information that would support our understanding of the key elements of each thermal process. Then, we reviewed the current progress of validation studies of 7 individual heating technologies (drying roasting, radiofrequency-assisted pasteurization, superheated steam, etc.) and the combined treatments (e.g., infrared and hot air). Last, we discussed knowledge gaps that require more scientific data in the future studies. We aimed to provide a process-centric view point of thermal pasteurization studies of low-moisture foods. The information could provide detailed protocol for process developers, operators, and managers to enhance low-moisture foods safety.

Inhibition of TNBS-induced intestinal inflammation in crucian carp (Carassius carassius) by oral administration of bioactive Bioactive food derived peptides.

Intestinal enteritis is a main issue in crucian carp production which results in massive economic loss. Traditional antibiotics used for disease prevention of crucian carp (Carassius carassius) have been banned, thus an alternative approach needs to be identified. In this study, the bioactive peptide was evaluated as a diet supplement for preventing intestinal inflammation in crucian carp. Intestinal inflammation was induced by intrarectal administration of a 2,4,6-trinitrobenzene sulfonic acid (TNBS) solution. The fish samples were fed with different diets for 14 days. The disease activity index (DAI), which included, fish swimming, food intake, anal inflammation, body surface, and ascites was determined daily. Intestine segments were stained with haematoxylin and eosin (H.E.) for histopathological analysis. The expression of cytokines, including interleukin-1ß (IL-1ß), interleukin-8 (IL-8), tumor necrosis factor α (TNF-α), and myeloperoxidase (MPO) in crucian carp were determined. In TNBS-induced groups, the DAI scores were dramatically increased compared to the control group. The histopathological analysis showed that the damage of the fish intestine after the injection of TNBS. The relative expression levels of pro-inflammation cytokines (TNF-α, IL-1ß, IL-8, MPO) were significantly increased compared to the control group on day 1. In the TNBS-induced group feed with a diet supplemented with bioactive peptide, the symptoms of intestinal inflammation were relieved on day 3 and the mRNA expression levels of pro-inflammation cytokines (TNF-α, IL-1ß, IL-8, MPO) were reduced compared to day 1. On day 7, the fish samples enrofloxacin group and bioactive peptide group were recovered from TNBS-induced intestinal inflammation. This study showed that the fish diet supplemented with bioactive peptide could help to prevent and recover from intestinal inflammation. Thus, the bioactive peptide can be used as a replacement for antibiotics to prevent disease in aquaculture production.

Thermal inactivation kinetics of Salmonella enterica and Enterococcus faecium NRRL B-2354 as a function of temperature and water activity in fine ground black pepper.

Fine ground black pepper generally consumed as a seasoning without any further processing has been associated with Salmonella enterica outbreaks. Thermal inactivation kinetics data is necessary to develop a pasteurization process for fine ground black pepper. This study investigates the influence of temperature and water activity on thermal inactivation kinetics of Salmonella in fine ground black pepper. It also assesses the suitability of Enterococcus faecium as a surrogate for Salmonella. Fine ground black pepper of varying water activities, aw (0.40, 0.55, 0.70) was subjected to isothermal treatments at different temperatures (65-80 °C) for five equidistant time points with intervals ranging from 18 s to 250 min. The survival data were used to fit two primary models (log-linear and Weibull) and two secondary models (response surface and Modified Bigelow). Results indicated that among the two primary models, the Weibull model explained the thermal inactivation kinetics better with lower RMSE (0.24 - 0.56 log CFU/g) and AICc values at all aw and temperatures. Water activity and treatment temperature significantly enhanced the thermal inactivation of Salmonella. E. faecium NRRL B-2354 was found to be a suitable surrogate for Salmonella in fine ground black pepper at all tested treatment conditions. The developed modified Bigelow model based on the Weibull model could be applied to predict the inactivation kinetics of Salmonella in black pepper and would benefit the spice industry in identifying process parameters for thermal pasteurization of fine ground black pepper.

Thermal inactivation of Salmonella on chicken wings cooked in domestic convection and air fryer ovens.

Chicken wings are among the most popular poultry products for home and foodservice consumption. Poultry products must be handled and cooked safely to decrease the risk of foodborne salmonellosis for consumers. This study aims to validate the use of domestic appliances (convection and air fryer ovens) for the thermal inactivation of Salmonella on chicken wings. Wings (n = 3, 46.5 ± 4.3 g) were inoculated with a five-strain cocktail of Salmonella (ca. 8 log10 CFU/wing) and cooked in a convection oven (179.4°C) or an air fryer (176, 190, or 204°C) for 2, 5, 10, 15, 20, 22, or 25 min. Thermocouples recorded temperature profiles of wings and appliances. Salmonella counts were determined on XLD agar for rinsates (100 ml/sample), and rinsates were enriched to recover bacteria below the limit of quantification. The recommended internal cooking temperature (73.8°C) was achieved after a range of 7.5 to 8.5 min in both appliances. Salmonella counts were reduced by 6.5 log10 CFU/wing when this temperature was achieved. Cumulative lethality (F-value) calculations predicted a 9-log reduction after 7.0 to 8.1 min of cooking. However, sample enrichments tested positive for Salmonella for all cooking times below 22 min. Ultimately, cooking at the temperature-time combinations recommended by manufacturers and online recipes helped achieve complete microbial elimination in both appliances. This study contributes to the validation of home cooking methods to ensure consumer safety.

Heating of milk powders at low water activity to 95°C for 15 minutes using hot air-assisted radio frequency processing achieved pasteurization.

Salmonella persistence in milk powders has caused several multistate foodborne disease outbreaks. Therefore, ways to deliver effective thermal treatment need to be identified and validated to ensure the microbial safety of milk powders. In this study, a process of hot air-assisted radio frequency (HARF) followed by holding at high temperatures in a convective oven was developed for pasteurization of milk powders. Heating times were compared between HARF and a convection oven for heating milk powders to a pasteurization temperature, and HARF has been shown to considerably reduce the come-up time. Whole milk powder (WMP) and nonfat dry milk (NFDM) were inoculated with a 5-serotype Salmonella cocktail and equilibrated to a water activity of 0.10 to simulate the worst case for the microbial challenge study. After heating the sample to 95°C using HARF, followed by 10 and 15 min of holding in the oven, more than 5 log reduction of Salmonella was achieved in WMP and NFDM. This study validated a HARF-assisted thermal process for pasteurization of milk powder based on previously collected microbial inactivation kinetics data and provides valuable insights to process developers to ensure microbial safety of milk powder. This HARF process may be implemented in the dairy industry to enhance the microbial safety of milk powders.

Evaluation of Enterococcus faecium NRRL B-2354 as a surrogate for Salmonella in ground black pepper at different water activities.

Thermal inactivation kinetics of Salmonella in low moisture foods are necessary for developing proper thermal processing parameters for pasteurization. The effect of water activity on thermal inactivation kinetics of Salmonella and Enterococcus faecium NRRL B-2354 in ground black pepper has not been studied previously. Identification of a suitable surrogate assists in conducting in-plant process validations. Ground black pepper was inoculated with a 5-serotype Salmonella cocktail or E. faecium NRRL B-2354, equilibrated to water activities of 0.25, 0.45 or 0.65 in a humidity-controlled chamber, and isothermally treated at different temperatures. The survivor data were used for fitting the log-linear models to obtain the D and z-values of Salmonella and E. faecium in ground black pepper. Modified Bigelow models were developed to evaluate the effects of temperature and water activity on the thermal inactivation kinetics of Salmonella and E. faecium. Water activity and temperature showed significant negative effects on the thermal resistance of Salmonella and E. faecium in ground black pepper. For example, significantly higher D values of Salmonella were observed at water activity of 0.45 (D70°C = 20.5 min and D75°C = 7.8 min) compared to water activity of 0.65 (D70°C = 3.9 min and D75°C = 2.0 min). D-values of E. faecium were significantly higher than those of Salmonella at all three water activities, indicating that E. faecium is a suitable surrogate for Salmonella in thermal processing validation.

Inactivation of Salmonella enterica and Enterococcus faecium NRRL B2354 on cumin seeds using gaseous ethylene oxide.

The objectives of this study were to investigate the effects of processing parameters (relative humidity (RH), temperature, and exposure time) on the ethylene oxide (EtO) microbial inactivation of Salmonella spp. and to evaluate Enterococcus faecium NRRL B2354 as a suitable surrogate for Salmonella inactivation on cumin seeds. Five grams of cumin seeds inoculated with either Salmonella or E. faecium were treated with EtO at different temperatures (46, 53, and 60 °C) and RH (30, 40, and 50%) levels for different exposure time to investigate the effects of process parameters on the microbial inactivation. The Weibull model fit the survival data of both bacteria with a shape parameter p < 1, which showed a tailing effect with concave shape indicating that the sensitive cells were inactivated first, and the sturdy ones survived at low RH treatment conditions. In general, the log reductions of both bacteria on cumin seeds increased with the increasing RH and temperature for EtO treatment. RH is a critical factor for successful EtO inactivation treatment. RH must be higher than 40% to implement a successful and efficient EtO decontamination of cumin seeds. E. faecium consistently showed lower log reductions than those of Salmonella under all EtO treatment conditions investigated in this study, demonstrating that E. faecium is a suitable surrogate for Salmonella. Twenty minutes of EtO treatment at 50% RH achieved ~5 log reductions of both bacteria at all three temperatures. A response surface model was developed to predict the log reductions of both bacteria under different treatment conditions and the contour plots representing log reductions were created. Inactivation is positively correlated to temperature and RH. Therefore, a higher temperature is required to achieve the desired log reduction at lower RH and vice versa. The developed response surface model is a valuable tool for the spice industry in identifying the possible combinations of EtO process parameters (temperature, RH, and exposure time) required to achieve a desired microbial reduction of Salmonella for ensuring microbial food safety of spices.

A Comparison of Three Methods for Determining Thermal Inactivation Kinetics: A Case Study on Salmonella enterica in Whole Milk Powder.

ABSTRACT: Different methods for determining the thermal inactivation kinetics of microorganisms can result in discrepancies in thermal resistance values. In this study, thermal resistance of Salmonella in whole milk powder was determined with three methods: thermal death time (TDT) disk in water bath, pouches in water bath, and the TDT Sandwich system. Samples from three production lots of whole milk powder were inoculated with a five-strain Salmonella cocktail and equilibrated to a water activity of 0.20. The samples were then subjected to three isothermal treatments at 75, 80, or 85°C. Samples were removed at six time points and cultures were enumerated for survivors. The inactivation data were fitted to two consolidated models: two primary models (log linear and Weibull) and one secondary model (Bigelow). Normality testing indicated that all the model parameters were normally distributed. None of the model parameters for both consolidated models were significantly different (α = 0.05). The amount of inactivation during the come-up time was also not significantly different among the methods (α = 0.05). However, the TDT Sandwich resulted in less inactivation during the come-up time and overall less variation in model parameters. The survivor data from all three methods were combined and fitted to both consolidated models. The Weibull had a lower root mean square error and a better fit, according to the corrected Akaike's information criterion. The three thermal treatment methods produced results that were not significantly different; thus, the methods are interchangeable, at least for Salmonella in whole milk powder. Comparisons with more methods, other microorganisms, and larger varieties of food products using the same framework presented in this study could provide guidance for standardizing thermal inactivation kinetics studies for microorganisms in foods.

Evaluation of Enterococcus faecium NRRL B-2354 as a surrogate for Salmonella enterica in milk powders at different storage times and temperatures.

While the increase in thermal resistance of microorganisms at reduced water activity is demonstrated for low-moisture food products, the effect of storage time on the thermal resistance of microorganisms in low-moisture foods is not well established. As low-moisture foods are stored for long periods and are used as ingredients, cross-contamination can occur at any time period before the lethality step. Therefore, this study was designed to investigate the effect of storage time (30, 60, and 90 d) on the thermal resistance of Salmonella and Enterococcus faecium NRRL B-2354 in milk powders at a low water activity of 0.10 (conservative level). In this study, 2 milk powders, whole milk powder (WMP) and nonfat dry milk (NFDM), were inoculated with a 5-serotype Salmonella cocktail or E. faecium and equilibrated to a water activity of 0.10. The thermal resistance of Salmonella and E. faecium in WMP and NFDM were determined at different storage times (30, 60, and 90 d) at 85°C. The storage time had no effect on the thermal inactivation kinetics of Salmonella within 90 d of storage at 85°C. In the second part of this study, isothermal treatments were also conducted at higher temperatures (90 and 95°C) to evaluate the suitability of E. faecium as a surrogate for Salmonella in milk powders. The D-values of Salmonella in WMP with 30 d of storage at 85, 90, and 95°C were 7.98, 3.35, and 1.68 min. The corresponding values for E. faecium were 16.96, 7.90, and 4.16 min. Higher D-values of E. faecium indicates that it is a conservative surrogate. Similar results were found for NFDM. In general, D-values of both microorganisms are slightly higher in NFDM than WMP. Two primary models (log-linear and Weibull) were compared for their goodness-of-fit. The Weibull model was found to be more appropriate than the log-linear model. This study provides valuable information for establishing process validation for the pasteurization of milk powders.

Effect of water activity on the thermal inactivation kinetics of Salmonella in milk powders.

Persistence of Salmonella in milk powders has caused several foodborne outbreaks. The determination of proper pasteurization processing conditions requires an understanding of the thermal inactivation kinetics of Salmonella in milk powders. However, there is a lack of knowledge related to the effects of water activity (aw) and fat content on Salmonella inactivation in milk powder during thermal processing. Two types of milk powders, nonfat dry milk and whole milk powder, with different fat contents (0.62 and 29.46% wt/wt, respectively) were inoculated with a 5-strain cocktail of Salmonella and equilibrated to 3 aw levels (0.10, 0.20, and 0.30) for isothermal treatments at 75, 80, and 85°C to obtain D-values (the time required to achieve a 10-fold reduction of the bacteria at the isothermal treatment temperature) and z-values (the increase in temperature required to achieve a 90% reduction of the decimal reduction time D). Stability tests showed that the inoculation method used in this study provided a high and stable population of Salmonella for thermal inactivation studies. A moisture sorption isotherm was measured to understand the relationship between aw and moisture content of milk powders. The thermal resistance of Salmonella was found to significantly increase as aw decreased, which suggested that a higher temperature or longer processing time would be required at low aw to achieve the desired inactivation of Salmonella. The microbial inactivation kinetics were not significantly different for the 2 milk powders; therefore, data were combined to develop a universal model. A response surface model was compared with a modified Bigelow model. The modified Bigelow model performed well to predict D-values [root mean square error (RMSE) = 1.47 min] and log reductions (RMSE = 0.48 log cfu/g). The modified Bigelow model developed here could be used to estimate D-value as a function of water activity and temperature to design a thermal pasteurization system for milk powders.

A microbial challenge study for validating continuous radio-frequency assisted thermal processing pasteurization of egg white powder.

Spray dried egg white powder (EWP) is traditionally processed by hot room treatment for a prolonged period of time (67 °C for 15 days) to enhance its functionality (foaming and gelling) and to improve microbial safety of EWP. Our prior research demonstrated that radio-frequency (RF) assisted thermal processing can considerably reduce the processing time, without compromising the functional properties of EWP. In this study, continuous RF processing was evaluated for pasteurization of EWP. EWP samples were inoculated with a 5-strain Salmonella cocktail or Enterococcus faecium NRRL B-2354 for the microbial challenge studies. To evaluate the inoculation method, stability and homogeneity tests were conducted for both Salmonella and E. faecium in EWP. Continuous RF heating of EWP was conducted in a 6-kW, 27.12 MHz pilot-scale parallel-plate RF heating system. RF-assisted thermal processing of EWP at 80 °C for 2 h provided >6.69 log reduction for Salmonella. E. faecium was found to be a suitable surrogate for Salmonella due to its higher resistance and similar inactivation kinetics during RF heating of EWP. The validated RF-assisted thermal process can be scaled up for use in the egg industry.

Radiofrequency pasteurization process for inactivation of Salmonella spp. and Enterococcus faecium NRRL B-2354 on ground black pepper.

Salmonella persistence in ground black pepper has caused several foodborne outbreaks and created public concern about the safety of low water activity (aw) foods. In this study, radiofrequency (RF) processing was evaluated for pasteurization of ground black pepper. Stability and homogeneity tests were done for both Salmonella spp. and E. faecium during moisture equilibration before RF heating to evaluate the inoculation method. Moisture content of samples were conditioned such that the final moisture content after RF heating reached the optimal storage moisture. RF heating was shown to provide more than 5.98 log CFU/g reduction for Salmonella spp. and the reduction of 3.89 log CFU/g for E. faecium with a 130 s of treatment time. The higher thermal resistance of E. faecium indicated its suitability as surrogate for Salmonella spp. during RF heating of ground black pepper. Piperine, total phenolics, volatile compounds, and antioxidant activity were assessed as quality parameters for ground black pepper. The results demonstrated that the RF processing provided effective inactivation of Salmonella spp. with insignificant (p > 0.05) quality deterioration.

Radio-Frequency Processing for Inactivation of Salmonella enterica and Enterococcus faecium NRRL B-2354 in Black Peppercorn.

Several Salmonella outbreaks linked to black pepper call for effective inactivation processes, because current decontamination methods result in quality deterioration. Radio-frequency (RF) heating provides a rapid heating rate and volumetric heating, resulting in a shorter come-up time. This allows for choosing a high-temperature and short-time combination to achieve the desired inactivation with minimal quality deterioration. The objectives of this study were to evaluate RF heating for inactivation of Salmonella enterica and Enterococcus faecium in black peppercorn and evaluate quality changes of RF-treated black peppercorn. Black peppercorns were inoculated with a five-strain cocktail of Salmonella or E. faecium to attain initial population levels of 6.8 and 7.3 log CFU/g, respectively, and were then adjusted to a moisture content of 12.7% (wet basis) and a water activity of 0.60 at room temperature. A stability test was performed to quantify the microbial reduction during inoculation and equilibration before RF heating inactivation. During RF heating, the cold spot was determined to be at the center on the top surface of the treated sample. In addition to inoculating the entire sample, an inoculated packed sample was placed at the cold spot of the tray. An RF heating time of 2.5 min provided a 5.31- and 5.26-log CFU/g reduction in the entire sample contained in the tray for Salmonella and E. faecium, respectively. Color parameters (L*, a*, b*), piperine content, total phenolics, scavenging activity, and most of the volatile compounds of 2.5-min RF-treated samples were not significantly different from those of the control samples. These data suggest that RF heating is a promising thermal inactivation treatment for Salmonella without significant quality deterioration, and E. faecium seems to be a suitable surrogate for Salmonella to validate the efficacy of RF heating of black peppercorn.

Response Surface Methodology for Salmonella Inactivation during Extrusion Processing of Oat Flour.

An increase in the number of foodborne outbreaks and recalls due to Salmonella in low-moisture foods has resulted in the need for the development and validation of process controls to ensure their microbiological safety. Furthermore, the Food Safety Modernization Act Preventive Controls for Human Food final rule requires food processors to validate their process controls to ensure food safety. The objective of this study was to develop a response surface model to predict Salmonella inactivation in oat flour, as affected by moisture, fat content, screw speed, and temperature. Oat flour was adjusted to different moisture (14 to 26% wet basis) and fat (5 to 15% [w/w]) contents and was then inoculated with a five-strain cocktail of Salmonella. Inoculated material was extruded through a single-screw extruder running at different screw speeds (75 to 225 rpm) and temperatures (65 to 85°C), without a die. Once steady-state conditions were attained, extruded samples were collected, cooled, and stored under refrigeration, and Salmonella survivors were enumerated. A split-plot central composite second-order response surface design was used, with the central point replicated six times. Temperature showed a significant ( P < 0.0005) positive effect on microbial reduction. Moisture content showed significant linear ( P = 0.0014) and quadratic ( P = 0.0005) effects, whereas higher fat content showed a significant ( P < 0.0001) protective effect on Salmonella destruction. The screw speed did not play a major role in inactivating Salmonella, but it had a significant ( P = 0.0004) interactive effect with temperature. Results indicated that a >5.5-log reduction was achieved in oat flour extruded at a temperature above 85°C at all moisture and fat contents evaluated at a screw speed of 150 rpm. The developed response surface model can be used to identify the extrusion process conditions to achieve a desired reduction of Salmonella based on the moisture and fat contents of the product.

Evaluation of Enterococcus faecium NRRL B-2354 as a Surrogate for Salmonella During Extrusion of Low-Moisture Food.

Salmonella in low-moisture foods is an emerging challenge due to numerous food product recalls and foodborne illness outbreaks. Identification of suitable surrogate is critical for process validation at industry level due to implementation of new Food Safety Modernization Act of 2011. The objective of this study was to evaluate Enterococcus faecium NRRL B-2354 as a surrogate for Salmonella during the extrusion of low-moisture food. Oat flour, a low-moisture food, was adjusted to different moisture (14% to 26% wet basis) and fat (5% to 15% w/w) contents and was inoculated with E. faecium NRRL B-2354. Inoculated material was then extruded in a lab-scale single-screw extruder running at different screw speeds (75 to 225 rpm) and different temperatures (75, 85, and 95 °C). A split-plot central composite 2nd order response surface design was used, with the central point replicated six times. The data from the selective media (m-Enterococcus agar) was used to build the response surface model for inactivation of E. faecium NRRL B-2354. Results indicated that E. faecium NRRL B-2354 always had higher heat resistance compared to Salmonella at all conditions evaluated in this study. However, the patterns of contour plots showing the effect of various product and process parameters on inactivation of E. faecium NRRL B-2354 was different from that of Salmonella. Although E. faecium NRRL B-2354 may be an acceptable surrogate for extrusion of low-moisture products due to higher resistance than Salmonella, another surrogate with similar inactivation behavior may be preferred and needs to be identified. PRACTICAL APPLICATION: Food Safety Modernization Act requires the food industry to validate processing interventions. This study validated extrusion processing and demonstrated that E. faecium NRRL B-2354 is an acceptable surrogate for extrusion of low-moisture products. The developed response surface model allows the industry to identify process conditions to achieve a desired lethality for their products based on composition.