by Marta M. D. C. Vila 1,*,Edjane C. Cinto 1,Arthur O. Pereira 1,Denicezar Â. Baldo 2,José M. Oliveira Jr. 2 andVictor M. Balcão 1,3
VBlab—Laboratory of Bacterial Viruses, University of Sorocaba, Sorocaba 18023-000, SP, Brazil
LaFiNAU—Laboratory of Applied Nuclear Physics, University of Sorocaba, Sorocaba 18023-000, SP, Brazil
Department of Biology and CESAM, University of Aveiro, Campus Universitário de Santiago, P-3810-193 Aveiro, Portugal
Author to whom correspondence should be addressed.
Polymers 2024, 16(5), 680; https://doi.org/10.3390/polym16050680
Abstract
The goal of this research was to create an antibacterial biopolymeric coating integrating lytic bacteriophages against Salmonella enterica for use in ripened cheese. Salmonella enterica is the main pathogen that contaminates food products and the food industry. The food sector still uses costly and non-selective decontamination and disease control methods. Therefore, it is necessary to look for novel pathogen biocontrol technologies. Bacteriophage-based biocontrol seems like a viable option in this situation. The results obtained show promise for food applications since the edible packaging developed (EdiPhage) was successful in maintaining lytic phage viability while preventing the contamination of foodstuff with the aforementioned bacterial pathogen.
Keywords:
lytic bacteriophage particles; Salmonella enterica; antibacterial edible biopolymeric coating (EdiPhage); bacteriophage structural and functional stabilization
1. Introduction
Foodborne illnesses, or foodborne diseases, are a major cause of morbidity and mortality and a major public health problem worldwide. According to estimates from the World Health Organization (WHO), eating food tainted with dangerous microorganisms results in the deaths of 1.9 million children annually [1,2]. The three primary bacteria that cause foodborne infections are Salmonella enterica, Escherichia coli, and Staphylococcus aureus [3,4].
Over 50% of the recognized serotypes of Salmonella are caused by the Salmonella enterica species, which is responsible for most human infections with the bacteria [5,6]. Daniel Salmon identified and described Salmonella, a bacillus Gram-negative member of the Enterobacteriaceae family, in 1885 [5]. Salmonella is typically split into two species: Salmonella enterica and Salmonella bongori. There are over 2500 recognized serotypes of Salmonella. The contamination of food by Salmonella occurs through various factors, such as exposure time to the environment during the manufacturing process, preparation, and/or storage [7]. Salmonella is the most prevalent foodborne pathogen causing more than 93 million cases of salmonellosis and 150,000 fatalities every year [5]. These bacillus outbreaks in recent years have been linked to a variety of items, including raw tuna, cabbage, chicken, eggs, pistachios, cucumbers, and pre-cut melons [4]. Many proposals have been evaluated to control the main pathogens causing food poisoning. The developing technologies are anticipated to be sustainable and to have the least negative effects on nutrients and food quality, all the while taking into account the difficulties of effectively inactivating pathogenic microorganisms in various food matrices [8].
Contamination by bacteria can occur during the slaughter, milking, fermentation, processing, storage, or filling among other processes [7]. The increasing demand for high-quality, shelf life-extended, ready-to-eat food products has led to the development of new processing technologies that ensure that the product’s natural attributes and appearance are not significantly compromised [9].
Among the various strategies used to minimize the microbial load of some foods, the use of antibiotics has been explored [6]. However, antibiotic substances present restricted use due to both the negative impact on human antimicrobial therapies as well as the selection of more resistant microorganisms [10]. The use of physical methods, such as superheated steam, dry heat, and UV light, can lead to product acceptability problems and the deterioration of the organoleptic properties of foods [11,12]. In addition, some approaches often used in processed foods to reduce contamination by foodborne pathogens cannot be directly applied to fresh fruits, vegetables, and ready-to-eat products [13]. New processing technologies such as gamma-ray irradiation, plasma processing, high-pressure processing, pulsed electric field, and ultrasound, can be efficient, but have high costs [14]. Therefore, the development of new processing strategies to reduce bacterial pathogens in food while still meeting consumer demands for minimally processed foods, with low concentrations of chemical preservatives, has been more and more urgent [1].
In this context, bacteriophages (or phages) have emerged as a bacterial biocontrol tool with enormous potential in the fight to reduce the burden of infectious diseases [2,14]. Bacteriophages were discovered in the mid-1910s by British scientist Frederick Twort [15]. During his research with virus cultivation, Twort realized that plates contaminated with some bacteria showed zones of lysis, and therefore, he assumed that there was possibly some microorganism capable of lysing bacterial cultures. The official discovery was made in 1917 by the French-Canadian microbiologist Felix D’ Herelle, who used lytic viruses from Shiguella dysenteriae to treat his dysentery [15]. D’ Herelle named this virus a bacteriophage (or phage) and was congratulated as the “father” of modern virology [10]. Bacteriophages are viruses that solely and exclusively infect susceptible bacterial cells and have no metabolic machinery of their own, hence being intracellular parasites requiring a bacterial host cell to replicate [16,17,18]. Many applications of bacteriophages in the control of foodborne pathogens have been proposed over the years, with relative success [14]. Bacterial biocontrol using bacteriophages has the unique advantage that phage particles are natural antibacterial agents, self-multiplying and highly specific [16]. Another interesting property is the remarkable stability of bacteriophages in foodstuff [11]. Furthermore, according to Sillankorva et al. [19] and Sahu et al. [16], bacteriophages have several other advantages as biocontrol agents in foodstuff, including (but not limited to): (i) specificity to reaching their bacterial host cells while keeping the local microbiota unaffected; (ii) self-replication and self-limitation, such as in multiplying while the target host cells are still present and viable; (iii) adaptation to the defense mechanisms of the bacterial cells; (iv) a very low inherent toxicity, since they are formed basically of nucleic acids and proteins; (v) a very low cost of isolation and simplicity of handling; and (vi) tolerance to various food conditions. In this way, researchers have therefore attempted to employ these bacterial viruses to combat a variety of bacterial illnesses in humans and animals [5,16]. Phage particles can be used to battle foodborne pathogens at every stage of manufacturing across the food chain. Bacteriophages are appropriate in (i) stopping or lessening illness and colonization in livestock; (ii) cleaning up carcasses and other unprocessed goods like fresh produce, eggs, and fruits; (iii) the decontamination of surfaces and equipment; and (iv) increasing the shelf life of perishable industrialized foods [7,20].
Keeping in mind everything described above, the main objectives of the research presented were to isolate lytic bacteriophages for Salmonella enterica from ambient sources and to characterize them from both physicochemical and biological points of view, aiming at producing an edible biopolymeric film integrating a phage cocktail containing the isolated phages with the potential for controlling Salmonella enterica in foodstuff, using matured cheese as a food matrix model.