Listericidal Novel Processing Technological Approaches for the Safety of Milk and Dairy Products: A Systematic Review

Abstract

Listeria monocytogenes is a major public health concern in milk and ready-to-eat dairy products. To meet consumer demand for fresher, minimally processed foods with high nutritional and sensory quality, several non-thermal technologies are being explored as alternatives to conventional heat treatments. This systematic review (2020–2025), following PRISMA guidelines, examines recent applications of selected non-thermal technologies to control Listeria in milk and dairy matrices. Peer-reviewed studies available in full-text, in English or Spanish, focusing on applications at laboratory or pilot plant scales, with milk or dairy produced onsite or purchased, containing Listeria sp., were included. Studies with applications to plant-based or non-dairy products or those not inoculated with Listeria, were excluded. Conference abstracts, corrections, editorials, letters, news, and scientific opinions were excluded as well. The databases searched were Web of Science, Scopus, and ProQuest, which were last consulted in April 2025. Given the naturality of the review, the risk of bias was assessed through independent screening by two of the researchers, focusing on clear objectives, analytical validity, statistical analysis, and methodology. The results are presented in tabulated format. Of the 157 records identified, 22 were included in this review. Seven of the records reported hurdle technologies, while fifteen reported single technology applications, with high-pressure processing being the most frequent. Limitations observed are primarily the use of unreported strains, a lack of information regarding the initial load of inoculum, and expected log reductions. The equipment used is mostly at the laboratory scale, except for HPP. Non-thermal technologies present a promising option for the control of Listeria in dairy products. The basic principles of GMP, HACCP, and cold-chain control in dairy processing are of special importance in safety assurance. This research was funded by Vicerrectoría de Investigación, Universidad de Costa Rica, grant number 735-C3-460.

1. Introduction

Milk and dairy derivatives may be responsible for a wide variety of foodborne outbreaks due to their favorable physicochemical and nutritional features, which facilitate pathogen growth [1]. During the last decade, several listeriosis outbreaks have been associated with the consumption of contaminated dairy products, particularly cheese and ready-to-eat foods [2]. Listeria monocytogenes represents a significant public health concern in this food group, given its survival capacity in a broad range of temperatures and environmental conditions, including refrigerated storage, due to its psychrophilic nature, and a wide pH range between 4.6 and 9.5 [1,3,4].

Milk and dairy products represent one of the most important sources of transmission of L. monocytogenes to humans [5], driven in part by the high nutritional value of these products, characterized by the carbohydrates, fatty acids, and high-quality protein contents, as well as important micronutrients including vitamins, minerals, and trace elements [6]. L. monocytogenes control measures in dairy foods and dairy processing environments require permanent and diligent vigilance, monitoring, and corrective action [2].

L. monocytogenes is a facultative anaerobic, Gram-positive bacterium with a high case-fatality rate of up to 30% among foodborne pathogens. It is especially dangerous for pregnant women, neonates, the elderly, and immunocompromised individuals and is notorious for contaminating ready-to-eat (RTE) dairy products due to its persistence in processing environments and resistance to adverse conditions [3,5]. This pathogenic bacterium was identified as the etiologic agent in nearly all recent outbreaks in North America attributed to pasteurized dairy products [7]. In Europe, it is considered the most severe foodborne disease with the highest case fatality and hospitalization rates [3]. Listeriosis is manifested as gastroenteritis in immunocompromised people, as bacteremia and central nervous system infection in immunocompromised patients and elderly populations, and as placental and fetal infection in pregnant women [3]. Therefore, the establishment of preventive and control measures to avoid its presence in milk and RTE dairy products is of relevance.

L. monocytogenes contamination may occur due to cross-contamination during processing, even after pasteurization [4]. Proper and validated cleaning and sanitation protocols for food contact surfaces and non-food contact surfaces aligned with a microbial environmental monitoring program are, therefore, essential tools for reducing contamination risks [8]. The literature reports an association between hypervirulent L. monocytogenes clones and dairy products manufactured from raw milk [3]. Moreover, milk and dairy processing facilities often show organic residues and wet conditions that facilitate Listeria survival and growth. External factors may also contribute to the introduction of Listeria into processing environments, such as contaminated raw materials, wild and farm animals that may be asymptomatic carriers of L. monocytogenes, and rodents and insects, all of which are well-identified carriers and vehicles of transmission of this pathogen. Fecal shedding of Listeria by dairy cows represents a common route of entry of Listeria into dairy processing facilities, and floors, drains, conveyor belts, slicers, and tables are common locations where Listeria spp. persist in food manufacturing environments. Furthermore, Listeria may be easily spread throughout the processing environment through inappropriate personnel movements and practices, contaminated personal protective equipment, and inadequate processing workflows [8].

Conventional methods such as thermal treatments are commonly used listericidal processing approaches in dairy products; however, pasteurization and sterilization may have detrimental effects on the sensorial and nutritional quality of foods, such as heat-induced protein denaturalization, which decreases nutritional value and increases the level of undesirable aroma compounds [9,10]. As a consequence, there is growing interest in developing non-thermal processing alternatives, such as high-pressure homogenization, high-pressure carbon dioxide, high-pressure processing (HPP), pulsed electric fields (PEFs), high-intensity ultrasound, and cold plasma, for the inactivation of pathogenic bacteria and the shelf life extension of dairy foods [9,10]. Emerging non-thermal technologies are promising approaches as pathogen control hurdles, with better retention of nutrients and the fresh-like characteristics of milk components [11].

High-pressure processing is the application of pressure between 400 and 600 MPa, leading to microbiological inactivation due to cell injury and protein denaturation [12]. Pulsed electric fields, on the other hand, are the application of a high voltage (20–50 kV/cm) with short pulses at a pulse-defined frequency, which will increase the temperature through liquid foods, causing the electrical breakdown of cell membranes [11]. UV-C at a wavelength of 253.7 nm prevents the growth of bacteria, viruses, molds, and other microorganisms by introducing lethal mutations in the genomes of these microorganisms; its germicidal effect directly relates to the radiation dose and exposure time [10]. Cold plasma is based on exposing foods to plasma (the fourth state of matter) at low temperature obtained by transforming gas into ionized gas containing atoms, ions, and electrons, by providing sufficient energy. The effectiveness of cold plasma is based on the production of UV radiation, reactive oxygen species, and reactive nitrogen species; its efficacy inactivating microorganisms is highly dependent on the conductivity and viscosity of the liquid food, as well as the electric field strength, treatment time, pulse repetition frequency, process temperature, and the design and composition of the electrodes [6]. In ohmic heating (OH), electrical energy is converted into thermal energy. When an electric current passes through a food that acts as an electrical resistor, electrons collide with other electrons, atoms, and ions, and then the electrical resistance is generated and raises the temperature of the food [13].

This systematic literature review aims to address the potential application of selected non-thermal processing technologies as novel listericidal approaches to ensure the safety of milk and dairy products. The paper discusses their applications in controlling L. monocytogenes in RTE dairy foods and provides feasible suggestions for their application as listericidal processing alternatives, outlining the potential directions for the advancement and application of novel technologies in the dairy industry. Specifically, this review paper focuses on the control of Listeria monocytogenes in milk and dairy products by using high-pressure processing, pulsed electric fields, ultraviolet light, pulsed UV-light, cold atmospheric plasma, ultrasound, and ohmic heating. This literature compilation highlights alternative non-thermal processing approaches. It includes hurdles to technological applications that aim to control L. monocytogenes in milk and other ready-to-eat dairy products (L. monocytogenes can support the growth of this pathogen of public health concern), which is a critical focus and food safety challenge. A further aim of this work is to clarify the available processing technologies applicable to milk and dairy food that may be implemented by the dairy industry to mitigate Listeria cross-contamination risks after milk pasteurization.

2. Methodology

2.1. Design

This systematic review conforms to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA 2020) statement [14]. According to the PRISMA checklist (Supplementary Materials Table S1), registration and protocol (24a, 24b, 24c in Supplementary Materials Table S1) [14] should be considered on the basis of the systematic review. Registration was made using Open-Ended Registration at OSF https://osf.io/285sd/?view_only=08e03257d6714ae7b6c5f42412505139 (accessed on 17 July 2025) [14]. A review protocol was followed by the authors, and amendments were made. Initially, three reports were assessed for eligibility; however, during the assessment by an independent reviewer, those reports were excluded due to the original article being reported in a review published before 2020 (n = 1), being a duplicate record (n = 1), and involving thermal treatment (n = 1) (Figure 1).

2.2. Eligibility Criteria

The eligibility criteria were established considering novel technologies for the control of Listeria sp. Criteria were studies focusing on applications at laboratory or pilot plant scales and dairy that could be purchased or produced on-site. Studies may focus on different pathogens, but must include Listeria sp. Other criteria were publications available in full text and peer-reviewed publications in English or Spanish. The search was limited to 2020 to April 2025. Exclusion criteria included studies with applications on plant-based products or non-dairy or not inoculated with Listeria. Conference abstracts, corrections, editorials, letters, news, and scientific opinions were excluded as well. Reviews were used to extract the original article; in cases where they were published before 2020, such studies were excluded. One review found included two references from before 2020 and one reference from 2021; the reported item referred to the original article.

2.3. Search Strategy

The databases searched were Web of Science, Scopus, and ProQuest. All sources were last consulted in April 2025. Given the aim of the review, different search strategies were used on databases; nevertheless, the search string was the same. In the Web of Science, the search was within the title, for Scopus, it was within the title, abstract, and keywords, and for ProQuest, it was within anywhere except the full text (NOFT). The search string was determined by selecting the technologies categorized as novel by a preliminary search and considering the probability of application in milk or dairy. The search string used was (‘non-thermal OR non-thermal OR non-conventional OR ultraviolet OR sonication OR pulsed AND electric AND field OR cold AND plasma OR membrane AND filtration OR x AND ray OR ohmic AND heating OR high AND pressure AND processing’) AND (‘milk’) AND (‘Listeria’). Zotero (Corporation for Digital Scholarship) and Excel (Microsoft) were used for reference management. Duplicates were removed.

2.4. Selection and Data Collection Process

Eligibility criteria, the search string, and databases were selected by consensus by the reviewers. The three databases were screened by one of the authors, and the data were extracted by the same person. A second reviewer independently checked the results. When doubts were presented, they were solved with the other two reviewers.

2.5. Data Items and Effect Measures

The extracted data included technology (e.g., type and conditions evaluated), strain, type of sample (e.g., type of dairy product, purchased or produced), and main results (e.g., effect of technology on strain). When data were collected, the reported methodology of the studies was assessed to ensure that the results responded to a statistical design according to the methodology and aim of the article.

2.6. Study Risk of Bias Assessment

While a formal risk of bias framework was not used, potential biases were minimized through a consistent screening process conducted independently by one reviewer and assessed afterwards by another reviewer, focusing on clear objectives, analytical validity, statistical analysis, and methodology.

2.7. Synthesis Methods

Due to the different types of applications that the search expected to find, tabulation of specific details, organized by technology and separated by individual or hurdle technologies, was selected as the synthesis method and allowed us to explore possible heterogeneity among studies. Excel (Microsoft) was used for management, data extraction, and analysis. No specialized systematic review software was used, given that Excel allows us to perform the steps of the systematic review process, resulting in complete reports. Figure 1 shows the PRISMA flow diagram with specific information on the search process.

3. Results

The initial search identified 157 studies through the three databases screened, resulting in 95 after eliminating duplicates, which were based on title, type of document, and abstract. Of the 28 reports assessed for eligibility, 22 were included in this review. The PRISMA flow diagram shown in Figure 1 summarizes the process of study selection.

The selected studies (

Table 1. Individual non-thermal technologies, processing parameters, Listeria strain, sample, and main results previously reported.

Technology	Processing Parameters	Strains	Sample	Inactivation	Reference
High-Pressure Processing (HPP)	10 °C; 350 MPa; time: 1, 2, 3, 4, 5 min. 4 °C and 10 °C; time: 0, 1, 4, 7, 10 days.	L. monocytogenes 7644 and GLM 5	Camel milk	Reductions: 2 to 3 log CFU/mL. D-value: 3.77 ± 0.36 min.	[17]
HPP	400, 500, 550, 600 MPa Time: 15 to 30 min. Room temperature. Storage at 4 ± 1 °C for 1, 4, 6, 8, and 10 days.	L. monocytogenes ATCC 7644	Ultrahigh-temperature-treated (UHT) milk (2% fat)	104 and 107 CFU/mL. at 550 MPa/15 min.	[12]
HPP	600 MPa at 20–25 °C with holding times < 5 min.	L. monocytogenes Scott A	UHT whole milk	Time of reduction of 5 log (t5): 3.6 ± 0.2 min to 7.4 ± 1.5 min, depending on the strain.	[18]
		L. monocytogenes NCTC 10527
		L. monocytogenes 4a KUEN 136	Raw whole milk
HPP	First cycle: 600 MPa/90 s. Second cycle: 600 MPa/120 s.	L. innocua ATCC 30090	Raw bovine milk	5.7 log CFU/mL.	[19]
HPP	600 MPa for 15 min.	L. monocytogenes ATCC 7644	UHT milk 2% fat	≥7 log reduction.	[20]
Cold Plasma (CP)	CP: 70 kV/15 min. Storage at 5 ± 2 °C for 7 days.	L. monocytogenes ATCC 7644	Raw Egyptian buffalo milk	3 log CFU/mL.	[9]
Atmospheric dielectric barrier discharge plasma	Input voltage: 50 V/Input power: 1000 W/Frequency: 10 kHz. Discharge gap of 5 mm between the quartz plate and sample surface. Exposure: 0, 30, 60, 90, and 120 s.	L. monocytogenes ATCC 19115 (G+)	Raw milk	Exposure > 120 s was more suitable for attenuating viability and avoiding recovery in raw milk within 7 days.	[20]
UV-C light (UV-C)	Lamp: 253.7 nm/18 W. Flow rate: 5–18 mL/min/Temperature 4–25 °C in the D-Optimal Quadratic model.	L. monocytogenes ATCC 19115	Milk	Reduction of 2.5 log CFU/mL.	[10]
UV-C	254 nm, flow rates: 30 and 100 L/h, cictinometric UV-C dose from 0 to 4169 ± 134 J/L.	L. innocua WS 2258	Raw milk and UHT milk (3.8% and 0.3% fat)	4.5 log CFU/mL.	[21]
Ohmic heating (OH)	5 V/cm, 10 V/cm, and 20 V/cm electric field. From 20 °C, to 62.5 °C, 5 min.	L. monocytogenes 4b (ATCC 13932)	UHT infant milk	20 V/cm reduced below the detection limit at the 4th min.	[1]
OH	10 V/cm and 50 Hz from 23.8 °C, to 60 °C.	L. monocytogenes ATCC 13932	Whole milk (3.1% fat), semi-skimmed milk (1.5%), and skimmed milk (0.1%)	Whole milk: 3.10 log CFU/mL. Semi-skimmed and skimmed milk: 5.30 log CFU/mL.	[22]
OH	Electric field intensities: 10 V and 20 V/cm. From 23 °C to 63 °C.	L. monocytogenes 4b (ATCC 13932)	Enriched milk with protein (2.5%, 5%, 7.5%)	OH 10 V/cm, 2.5% protein, time of reduction 9 min: <1 log CFU/mL. OH 20 V/cm, 2.5% and 5% protein, 2 min 30 s: <1 log CFU/mL.	[23]
OH	0, 4, 6, and 8 V/cm, 90–95 °C/5 min.	L. monocytogenes ATCC BAA 751	Whole raw milk (3.4% fat) for the elaboration of probiotic fermented milk	No viable cells (28 days)	[24]
Colinear-type Pulse Electric Fields (PEFs)	Pressure: 0.5 MPa with nitrogen gas. Flow rate: 10 L/h, moderate heat treatment at 60 °C. PEF treatment chambers with the g values: 1, 3, and 5 mm/1 h.	L. innocua, strain not reported	Long-life milk	7 log CFU/mL, 1 min.	[25]
Pulse Electric Fields (PEFs)	20 kV/cm, 55 °C.	L. monocytogenes, strain not reported	Skim milk	4.5 log CFU/mL.	[26]
	32 kV/cm, 20 °C.	L. innocua, strain not reported	Liquid whey protein formulation	6.5 log CFU/mL.
High-Voltage Pulsed Electric Field (HV-PEF)	From 0 to 180 kV/cm, lab setting at 27–28 °C.	L. monocytogenes, strain not reported	Milk	An increase in pulse counts (at 100 pulses) led to a decrease in Survival Rate (0.001).	[27]

and

Table 2. Hurdle technologies, processing parameters, Listeria strain, sample, and main results previously reported.

Technology	Processing Parameters	Strains	Sample	Inactivation	Reference
Hydrogen peroxide (HP), caprylic acid (CA), and UV radiation	HP and CA: 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%. Exposure time: 15 min. UV: 260 nm.	L. monocytogenes CCM 5578	Tile surface	2 log CFU/mL.	[15]
		L. monocytogenes (isolated from raw milk sheep cheese ‘Bryndza’)
Ultrasound-assisted pulsed ohmic heating (POH)	40 kHz and 50 W. 45, 50, 55, and 60 °C. 335, 475, and 525 s.	L. monocytogenes ATCC 19111, ATCC 19115, and ATCC 15313	Whole milk (3.6% fat), low-fat milk (1.0% fat), and non-fat milk (0% fat)	2 log CFU/mL.	[13]
Origanum onites essential oil (OEO) and Cold Atmospheric Plasma (CAP)	OEO: 2000 and 3000 ppm. CAP: 3 and 7 min. Storage at 4 °C for 45 days.	L. monocytogenes ATCC 13076	Iranian white cheese	E3000P7: <3.5 log CFU/mL.	[16]
PEFs	Constant pulse: 50 μs, frequency: 3 Hz, electric field strength: 10 kV/cm, flow rate: 2.92 L/h. Thermal treatment at a flow rate of 10 L/h. Temperatures: 63, 66, 69, 72, and 75 °C, 2 s.	L. monocytogenes ATCC 13932	UHT milk (1.5% fat) and raw goat milk	PEFs with thermal treatment: 5 log CFU/mL. PEFs without thermal treatment: 2.9 log CFU/mL.	[11]
Mild HHP, phage ListexTM P100, and the bacteriocin pediocin PA-1	Processing pressure of 200 and 300 MPa, 5 min, 10 °C.	L. monocytogenes Scott A (clinical isolate, ATCC 49594, serotype 4b); 1751 (isolated from dairy product, LRCESB, serotype 4b); ATCC 19116 (serotype 4c) L. innocua 2030c.	UHT whole milk (3.6% fat)	Non-recovery of L. monocytogenes during the shelf-life of milk at refrigeration temperatures.	[28]
Application of PU in combination with Thurincin	Frequency: 20–25 kHz, nominal power: 150 W, ultrasonic energy density: 0.914–0.943 W/cm3, and temperature of 30 ± 5 °C in combination with thurincin H (40 μg/mL)	L. innocua ATCC 33090	Ultra-pasteurized, partially skimmed milk enriched with vitamins A and D (28 g/L of butyric fat, 31 g/L of protein, and a pH of 6.5)	0.7 log CFU/mL.	[29]
Pulsed UV light (PUV)	Emission wavelength of 200–1100 nm, flow rate at 14.3–74.9 L/h; pulse frequency of 1–5 Hz; reactor configuration, annular (AT) and coiled tube (CT). Total delivered fluence 4.46 J/cm2 in the AT reactor and 22.47 J/cm2 for the CT reactor.	L. innocua 33090 (a surrogate for L. monocytogenes, given phenotypic similarity)	Skimmed milk	>3.5 log reduction. D-values (J/cm2) = 6.44. Reduction equivalent fluence (F0,REF (J/cm2) = 10.25 (AT), 119.5 (CT).	[30]

) emphasized the importance of dairy safety, particularly regarding Listeria as a persistent environmental pathogen. Novel technologies applied to milk and dairy products aim to develop applications for industry. Nevertheless, current information recommends further research, even in the case of high-pressure processing (HPP) being one of the most advanced, with enough data for certain applications (Figure 2). Some of the studies reported hurdle methodologies (Table 2), for instance, the use of essential oils with non-thermal technologies (UV and CP) [15,16] and a combination of non-thermal technologies [13].

4. Discussion

The reported novel technologies aim to achieve a reduction in temperature applied to milk and dairy to significantly reduce the risk of foodborne illness, traditionally accomplished by pasteurization. Evaluation of the effect of technology on different strains of Listeria provides valuable information for the industry due to the difficulties related to the control of this pathogen in dairy processing plants. Table 1 and Table 2 show that most studies evaluate technology using one specific strain or different strains but, individually, nevertheless, the recommended practice for conducting microbial challenge studies when evaluating novel technologies is to consider a cocktail of two to three Listeria strains, according to different guidelines, since strain variation and virulence may affect the results; when using a mixture, the behavior will be more complex, giving more conclusive results since the worst case scenario is evaluated [31]. For instance, when evaluating HPP, some results show that Listeria OSY8578 is more resistant; meanwhile, Scott A is more sensitive [18].

Heat resistance may be affected by several factors besides strain. For instance, the presence of background microbiota, pH, and stress treatment has been related to heat resistance, which makes the control or inactivation of pathogens such as Listeria complex [32]. Evidence shows that Listeria, once settled in the processing plant, is persistent and resistant to regular cleaning and sanitation. Measures such as cross-contamination programs, personal hygiene, and environmental control have been shown to support the control of Listeria in dairy processing [8]; therefore, Good Manufacturing Practices (GMP) and Hazard Analysis and Critical Control Point (HACCP) should be basic measures in the safety assurance strategy [2].

Food safety criteria might differ between countries or regulations. For instance, in ready-to-eat foods (RTE), such as dairy, Codex Alimentarius establishes that L. monocytogenes must be absent in 25 g, and the European Union establishes the same parameter before the food has left the immediate control of the food business operator who has produced it [2,32]. Meanwhile, the European Union has established 100 CFU/g for products on the market during their shelf life. Within the European Union, alert reports regarding milk, milk products, and Listeria in the same period as the search are mainly on cheeses (raw and pasteurized), and a few on butter, milk, and yogurt (RASFF Window). Compared with the samples found in this systematic review, the results show mainly applications on milk (raw, UHT, whole, semi-skimmed, and skimmed), while only three studies used cheese. This could be explained since milk is the raw material, and pasteurization is also applied to milk as a critical control point when processing dairy. Nevertheless, contamination with L. monocytogenes is attributed to milk and inadequate hygiene conditions or practices post-pasteurization, hence the importance of evaluating the application of novel technologies on finished products as well.

Cross-contamination with L. monocytogenes from the environment is a recognized hazard in RTE food [33]. Therefore, cleaning, sanitization, and monitoring programs are important in food facilities to control the risk of disease outbreaks or product recalls. Persistence of L. monocytogenes in the environment, the capability to form biofilms, and resistance to stressful conditions present a difficult scenario when facing the presence of this pathogen in dairy facilities [15]. The combined effect of chemicals was evaluated, showing that caprylic acid with hydrogen peroxide at concentrations lower than those used in separate applications was effective against L. monocytogenes CCM 5578, probably caused by faster and better penetration of the chemicals through the bacterial cell membrane [15], showing a potential use for the food industry.

Table 1 shows differences in the inactivation of L. monocytogenes obtained by means of CAP.UV-C, OH, PEFs, and HPP. It is difficult to compare the results since differences in experimental design are observed, mainly regarding strain, physicochemical characteristics, and equipment designs. For instance, results found for PEFs do not report strain; meanwhile, in the application of OH, three out of four studies use the same strain, which gives more consistent results. Evaluation of combined effects seems to be necessary for some applications where the individual effect does not reach an effective inactivation of Listeria. A synergistic effect is reported when combining power ultrasound with thurincin H (bacteriocin) [29], ultrasound, and pulsed ohmic heating [13], and the addition of OEO and Cold Atmospheric Plasma [16]. With the present results, HPP and OH appear to present the most robust evidence; meanwhile, CAP and UV require further optimization. PEFs show potential when varying equipment design and treatment conditions.

Regarding technologies, HPP achieved higher inactivation (5–7 log CFU/mL) when using 600 MPa on different strains; the time of reduction varies with strain; for instance, a study reported a time range from 3.6 to 7.4 min at the same pressure but varying from L. monocytogenes Scott A, NCTC 10527, and 4a KUEN 136 [18]. A limitation of the selected studies reporting HPP is that each one used single-strain applications. The results appear to be consistent in different samples of milk, from camel to bovine milk, whole and 2% fat, as shown in Table 1. Given the advances in HPP research, it may be possible that the application of combined technologies is not required for better results. Table 2 shows only one report of hurdle technologies using high-pressure processing and phage P100; in this case, the synergetic effect allows the use of a lower pressure, which may be valuable for preserving physicochemical characteristics.

The results for CAP and UV-C seem limited; only two studies were selected for each technology (Table 1), and differences in methodology, processing parameters, strain, and sample selection are observed. One hurdle technology was reported for CAP with OEO [16] and one for UV with HP and CA [15]. Inactivation was similar between studies (2–4.5 log UFC/mL). Regardless of this, it must be noted that in CAP technology, with OEO, the authors confirmed viability attenuation of L. monocytogenes in raw milk, with potential resuscitation, which could be risky during storage. Recommendations are made on the treatment of more than 120 s, as a parameter to avoid recovery within 7 days [16].

Our results show that researchers are moving towards the application of vegetable compounds and bacteriocins as hurdle technology [15,16,28,29], enhancing the non-thermal technologies with natural antimicrobials; more research seems to be needed for a better understanding of the effects.

Studies selected for OH were consistent with the selection of the strain; most of the studies evaluated similar processing parameters, including 10 and 20 V/cm from 20 °C to 63 °C. Processing time varies from 4 min to 9 min; the authors conclude that reductions are below the detection limit when 20 V/cm is applied in different samples [1,23]. Lower reductions were obtained in whole milk at 10 V/cm [22]. One study applied lower parameters of OH with higher temperatures [24], which gave good results regarding inactivation during storage (Table 1); nevertheless, application of higher temperatures (90–95 °C) takes away the final objective of non-thermal technologies.

Several studies have reported the effect of macronutrients on the performance of non-thermal technologies. For instance, when evaluating POH in milk with different fat content, the authors concluded that fat reduced the efficacy of the treatment due to its low electrical conductivity and sonoprotective effect [13], with this being congruent with the results shown for OH when applied as a single technology [22], on the other hand, one study applied UV-C and concluded that, with this technology, fat and optical density did not influence the inactivation rate [21]; finally, from the selected reports, one evaluated OH on milk with different protein content [23], concluding that at higher protein content, higher processing time is required to obtain desired reductions, due to a decrease in electrical conductivity.

The search period and specificity of the search strategy show that non-thermal technologies are in development, and recommendations are made to continue research considering multiple factors that may affect L. monocytogenes mechanisms. Some of the studies included in the review analyze, besides safety, the effect of the technology on quality properties, mainly during storage, which gives valuable information since consumers are expecting the same product despite the process, and the industry is requiring extended shelf life.

Regarding OH processing, one study proved that L. monocytogenes inactivation was achieved in a shorter time when high electric field intensity was applied [23]; the authors reported that there was no effect on the physicochemical properties of the milk, more specifically on pH and color measurement, providing conditions that may be used to produce protein-enriched milk, which responds to consumers’ demands for the nutritional composition of dairy products.

One study monitored color and pH during OH treatment, showing promising results since no significant change was observed under the studied conditions [22], making these conditions worthy of further research since both inactivation and quality parameters complied with expectations.

The antioxidant and antimicrobial properties of cheese were studied during 45 days of storage at 4 °C, and the investigation authors concluded that when applying 2500 ppm OEO with CAP (3 min, 25 kV), an increase in shelf life and overall quality of the product was observed [16]. When analyzing Table 1, the same study reported that a greater reduction in L. monocytogenes was obtained at 3000 ppm of OEO and 7 min CAP, which may not be the optimum conditions for the preservation of characteristics, since the results show that OEO and CAP affected pH values, and lower OEO concentrations resulted in a higher preference by panelists.

A study included in this review evaluated the growth of L. monocytogenes after HPP treatment during 10 days of storage at both regular temperature (4 °C) and at a higher temperature (10 °C), showing a “real world” scenario [17], since on the dairy supply chain variability of storage temperature is frequent due to continuous opening or malfunctioning of cold chambers, as well as a lack of temperature control. The results strengthen the importance of cold chains as a barrier after treatment, particularly due to the proliferation of spoilage microorganisms such as lactic acid bacteria, yeast, and molds, which also proves that microorganisms are not destroyed but damaged.

Regarding technologies reported in this review, there are some limitations to consider when deciding whether to migrate from traditional thermal technologies to non-thermal technologies. For example, as mentioned before, HPP is well established as a useful technology for dairy safety. Meanwhile, in CP dose control, the time of exposure and the methodology in which plasma is applied are still under investigation, which represents limitations to the technology [20]. On the other hand, UV application represents a challenge for fluids such as milk, due to its high absorbance [21]. Authors are cautious about recommendations since further research is needed to confirm the efficiency of these emerging processing approaches after the scaling up of technologies. Limitations were observed through the review. For instance, when using an unreported strain, a lack of information regarding the initial load of inoculum and a lack of information regarding expected log reductions. Data are important for a better understanding of the effect of technology and decision-making. It appears to be clear that, since applications are novel and technologies are starting to be tested against specific strains and specific food matrices, the equipment reported is mostly at a laboratory scale, at the moment, except for HPP.

5. Conclusions

Challenges seem to be focused on equipment design and the optimization of parameters since studies show differences related to fat and protein content. Nevertheless, non-thermal technologies present a promising option in the control of Listeria in milk and dairy products. Authors consider that for a better comparison of non-thermal technologies, evaluation of different scales of production, inoculation of strain cocktails instead of individual strains, and standardized initial inoculum should be considered in future investigations to obtain better conclusions. A relevant remark that researchers and industry should consider is that, despite technological performance, the basic principles of GMP, HACCP, and cold-chain control in dairy processing are of special importance in safety assurance.