Use Of Phage Cocktails Research Articles

Bacteriophage cocktail as a substitute for antimicrobials in companion animal dermatology

The present study focuses on the use of phage cocktails as a substitute for antibiotics in companion animal dermatology. For this purpose, a systematic search was carried out in the Scopus database, with the search criteria: "veterinary" and "bacteriophage" and "dermatology" in article title, abstract and keywords during the period 2010-2021. Seven in vitro studies and one in vivo study in companion animals, for which those carried out in laboratory animals were added. In this review, the use of non-transducing lytic phage cocktails as therapeutics for pyodermas is discussed and projected, as well as the resistance to phages and the strategies to overcome it, the comparison with antibiotics, the use of cocktails in other animal species, as well as the use of individual phages and cocktails in veterinary dermatology, and autochthonous phages as a strategy when phage collections from previous studies do not have the desired effects. It is concluded that non-transducing lytic autophage cocktails are an alter-native against antimicrobial resistance in companion animal dermatology. Finally, it is recommended to compare the use of these cocktails with other antibiotic substitutes and evaluate their possible syner-gism to reduce pathogenic bacteria on the skin.

Review of bacteriophage and its applications

The most prevalent and ubiquitous organisms in the world are bacteria-killing viruses called bacteriophages. The aim of this paper is to highlight some application areas of bacteriophages. There are both virulent and temperate bacteriophages in the environment, but only virulent bacteriophages are used for treatment, known as phage therapy. Since their discovery, bacteriophages have been considered a vital weapon to fight human and animal illnesses of bacterial origin. Currently, the emergence of growing microbial resistance to antibiotics and attention to bacteriophage use in treatment has all but grown up again. Correspondingly, the applications of bacteriophages for biocontrol purposes have become a remarkable opportunity in a number of fields, including farms and food production. Despite their treatment effects, bacteriophages also maintain a vital relationship with their hosts through moderating microbial populations and promoting their evolution by horizontal gene transfer. Additionally, they serve as vehicles for the transfer of vaccinations, the detection of dangerous microorganisms and systems for displaying proteins and antibodies. Bacteriophages are a varied collection of viruses that are simple to handle, making them suitable for use in treatments and biotechnology research. Hence, the expansion of various phage companies for phage production and the use of phage cocktails for the treatment of various bacterial diseases at different stages is recommended.

Use of Phages to Treat Antimicrobial-Resistant Salmonella Infections in Poultry.

Simple SummarySalmonellosis, an infection in humans and animals caused by Salmonella spp., poses a major concern to public health and food safety worldwide. Antibiotics are mostly prescribed to treat salmonellosis. Unfortunately, indiscriminate use of antibiotics leads to the emergence and transmission of multidrug-resistant Salmonella spp. As antibiotics are becoming increasingly ineffective, infections caused by MDR strains will be difficult to manage. The search for an alternative to antibiotics has led scientists to give renewed attention on phage therapy. Though commercial use of phages for controlling Salmonella in poultry is still in its early stage, the use of lytic phages is considered an environmentally friendly, cost-effective, and sustainable antimicrobial approach. Moreover, it provides advantages over antibiotics in terms of specificity, cost of development, resistance, and genetic amenability. Studies on laboratory and field scale use show promise on the effectiveness of phages against MDR Salmonella spp. However, inadequate data on safety of phage use, phage stability, and lack of regulatory framework remain major obstacles in the commercial application of phages. Our article provides a comprehensive overview on global prevalence and antimicrobial resistance of Salmonella in poultry, the efforts to control Salmonella using phage therapy, and challenges as well as future prospects of phage therapy.Salmonellosis is one of the most common bacterial infections that impacts both human health and poultry production. Although antibiotics are usually recommended for treating Salmonella infections, their misuse results in the evolution and spread of multidrug-resistant (MDR) bacteria. To minimize the health and economic burdens associated with antimicrobial resistance, a novel antibacterial strategy that can obliterate pathogens without any adverse effects on humans and animals is urgently required. Therefore, therapeutic supplementation of phages has gained renewed attention because of their unique ability to lyse specific hosts, cost-effective production, environmentally-friendly properties, and other potential advantages over antibiotics. In addition, the safety and efficacy of phage therapy for controlling poultry-associated Salmonella have already been proven through experimental studies. Phages can be applied at every stage of poultry production, processing, and distribution through different modes of application. Despite having a few limitations, the optimized and regulated use of phage cocktails may prove to be an effective option to combat infections caused by MDR pathogens in the post-antibiotic era. This article mainly focuses on the occurrence of salmonellosis in poultry and its reduction with the aid of bacteriophages. We particularly discuss the prevalence of Salmonella infections in poultry and poultry products; review the trends in antibiotic resistance; and summarize the application, challenges, and prospects of phage therapy in the poultry industry.

Deploying Viruses against Phytobacteria: Potential Use of Phage Cocktails as a Multifaceted Approach to Combat Resistant Bacterial Plant Pathogens.

Plants in nature are under the persistent intimidation of severe microbial diseases, threatening a sustainable food production system. Plant-bacterial pathogens are a major concern in the contemporary era, resulting in reduced plant growth and productivity. Plant antibiotics and chemical-based bactericides have been extensively used to evade plant bacterial diseases. To counteract this pressure, bacteria have evolved an array of resistance mechanisms, including innate and adaptive immune systems. The emergence of resistant bacteria and detrimental consequences of antimicrobial compounds on the environment and human health, accentuates the development of an alternative disease evacuation strategy. The phage cocktail therapy is a multidimensional approach effectively employed for the biocontrol of diverse resistant bacterial infections without affecting the fauna and flora. Phages engage a diverse set of counter defense strategies to undermine wide-ranging anti-phage defense mechanisms of bacterial pathogens. Microbial ecology, evolution, and dynamics of the interactions between phage and plant-bacterial pathogens lead to the engineering of robust phage cocktail therapeutics for the mitigation of devastating phytobacterial diseases. In this review, we highlight the concrete and fundamental determinants in the development and application of phage cocktails and their underlying mechanism, combating resistant plant-bacterial pathogens. Additionally, we provide recent advances in the use of phage cocktail therapy against phytobacteria for the biocontrol of devastating plant diseases.

Applications of Bacteriophage Cocktails to Reduce Salmonella Contamination in Poultry Farms.

Salmonella contamination is a critical problem in poultry farms, with serious consequences for both animals and food products. The aim of this study is to investigate the use of phage cocktails to reduce Salmonella contamination in poultry farms. Within the scope of the study, Salmonella phages were isolated from chicken stool. After the host range of phages was determined, morphological characterization was performed through transmission electron microscopy analysis. Then, replication parameters and adsorption rates were determined by one-step growth curves. After that, phage cocktail was prepared, and its effectiveness was tested in three environments, which were drinking water, shavings, and plastic surfaces. The results obtained have demonstrated that the phage cocktail can reduce Salmonella count up to 2.80 log10 units in drinking water, up to 2.30 log10 units on shavings, and 2.31 log10 units on plastic surfaces. It has been determined that phage cocktails could be a successful alternative in reducing Salmonella contamination in poultry environment. This work is the first study to investigate the use of phage cocktails for reducing Salmonella contamination in poultry water and on shavings, and it is presumed that the results obtained will contribute to the fight against pathogens by making them applicable to poultry farms.

Phage and phage lysins: New era of bio-preservatives and food safety agents.

There has been an increase in the search and application of new antimicrobial agents as alternatives to use of chemical preservatives and antibiotic-like compounds by the food industry. The massive use of antibiotic has created a reservoir of antibiotic-resistant bacteria that find their way from farm to humans. Thus, there exists an imperative need to explore new antibacterial options and bacteriophages perfectly fit into the class of safe and potent antimicrobials. Phage bio-control has come a long way owing to advances with use of phage cocktails, recombinant phages, and phage lysins; however, there still exists unmet challenges that restrict the number of phage-based products reaching the market. Hence, further studies are required to explore for more efficient phage-based bio-control strategies that can become an integral part of food safety protocols. This review thus aims to highlight the recent developments made in the application of phages and phage enzymes covering pre-harvest as well as post-harvest usage. It further focuses on the major issues in both phage and phage lysin research hindering their optimum use while detailing out the advances made by researchers lately in this direction for full exploitation of phages and phage lysins in the food sector.

Bacteriophage therapeutics to confront multidrug-resistant Acinetobacter baumannii - a global health menace.

We have already entered the post-antibiotic era as the outbreaks of numerous multidrug-resistant strains in the community as well as hospital-acquired infections are ringing alarm bells in the health sector. Acinetobacter baumannii is one such pathogen that has been considered a worldwide threat as it acquires multidrug resistance. It is one of the most challenging hospital-acquired pathogens as World Health Organization has listed carbapenem-resistant A. baumannii as a critical priority pathogen with limited therapeutic options. There is an urgent need to develop novel strategies against such pathogens to tackle the global crisis. Bacteriophages (phages), especially the lytic ones have re-emerged as a potential therapeutic approach. This review encompasses vast majority of phages against A. baumannii strains with special references related to single phage or monophage therapy, use of phage cocktails, combination therapy with antibiotics, use of phage-derived enzymes like endolysins and depolymerases to combat the pathogen and explore their therapeutic aspects. The concurrent ecological as well as evolutionary interplay between the phages and host bacteria demands in depth-research and knowledge, so as to utilize the maximum potential of the bacteriophage therapy.

Bacteriophage therapy for challenging bacterial infections: achievements, limitations and prospects for future clinical use by veterinary dermatologists.

Bacteriophages were discovered just over 100 years ago and have been used to treat bacterial infections in animals since the 1920s. The antimicrobial resistance crisis has led to a new surge of interest in the use of bacteriophage therapy as an alternative or supplement to antimicrobial therapy in humans and other animals. To describe the nature of bacteriophages and provide a critical review and update on the clinical use of bacteriophages in the treatment of challenging bacterial infections, with an emphasis on companion animal veterinary applications. The scientific literature on the subject was critically evaluated. Findings from the most pertinent papers have been presented in summary form and critiqued. Over the last 20 years there has been a considerable increase in the volume and quality of publications dealing with bacteriophage therapy. Some recent papers build on excellent work published in the 1980s and describe promising veterinary applications. Challenges related particularly to the registration and approval of phage remedies will need to be overcome before phage therapy can become a mainstream tool for use in veterinary settings. Considerably more research, particularly controlled clinical trials, needs to be done. Bacteriophage therapy is one of the most promising approaches to tackling the looming antimicrobial resistance crisis, yet substantial regulatory challenges will need to be overcome before it enters widespread use. Phage therapy also may, in the future, improve the management of challenging bacterial infections that are not necessarily multidrug-resistant.

Effective control of Shigella contamination in different foods using a novel six-phage cocktail

Shigella, the globally distributed causative agent of bacillary dysentery, is an important microbial contamination in raw and ready-to-eat foods. The recent commercial use of phage cocktails in the food industry is evidence that the method is a highly effective solution in controlling the pathogenic bacteria, especially the antibiotic-resistant strains. This research addresses the effects of a novel phage cocktail made of six different lytic bacteriophages to prevent drug resistant isolates of Shigella species in a wide variety of food products with a different matrix structure, viscosity, carbon content, and texture properties including cooked chicken, cherry tomato, yoghurt, cucumber and Chinese cabbage. After seven days of treatment with the phage cocktail, more than 99% of the bacterial count decreased compared to the control sample (P < 0.0001). The cocktail stability in dry conditions showed that the bacteriophages remain desirably active during food products' average shelf life. The prepared phage cocktail showed to be a very effective product for bio-controlling Shigella due to its high overlap of the host spectrums, the potential for use in a variety of foods, and stability in dry conditions.

New insights on phage efficacy to control Aeromonas salmonicida in aquaculture systems: An in vitro preliminary study

A major source of financial loss for the fish-farming industry is the occurrence of bacterial infections. Phage therapy can be a useful alternative tool to conventional treatments to control bacterial infections in aquaculture. The promising results obtained with phage AS-A to control the agent of furunculosis, Aeromonas salmonicida, led us to isolate two new phages and evaluate their dynamics, in cocktails and individually, to control this pathogenic bacterium. Moreover, considering that in outdoor facilities aquaculture water is exposed to the natural variability of physical and chemical parameters, the influence of pH, salinity, temperature, solar radiation and UV radiation on phage AS-D stability was also evaluated in this study, in order to develop an effective phage therapy protocol. Phages were assigned to the family Myoviridae and revealed identical morphological characteristics. Phage AS-A presents a higher burst size and a shorter latent period, decreasing the concentration of A. salmonicida sooner than phages AS-D and AS-E. However, phage AS-D presented higher rate of bacterial reduction, inducing also less bacterial resistance. Bacterial control with the cocktails was higher, namely when phage AS-A was combined with one of the two new phages or with both, but the main difference in the bacterial control was in the treatment time. Phage cocktails decrease the concentration of A. salmonicida sooner than single suspensions. The use of phage cocktails, in general, decreased phage-resistant mutants. The survival of AS-D phage was mostly affected by sunlight exposure (decrease of 3 log PFU/mL after 12 h) and high temperatures (decrease of 3 PFU/mL after 21 days and of 7 log PFU/mL after 49 days at 37 °C, but no decrease after 21 and 49 days at 25 °C and decrease of only 1 and 2 log PFU/mL after 21 and 49 days, respectively, at ambient temperature). The high bacterial control and low development of phage-resistant bacterial clones suggest that these phages can be used to control the furunculosis in aquaculture. Nonetheless, the stability of the phages was affected by solar radiation, this can be overcome by the application of phages at the end of the day or at night.

In vivo efficacy of single phage versus phage cocktail in resolving burn wound infection in BALB/c mice

Klebsiella pneumoniae is one of the most predominant pathogens associated with burn wound infections, causing considerable morbidity and mortality. The indiscriminate usage of antibiotics has led to the development of resistant strains, which have contributed towards the inefficacy of antibiotics. Phage therapy is a promising alternative to hinder the progression of pathogenic bacteria. However, phage bacterial resistance is already well known but the use of phage cocktails can overcome this drawback. The aim of the study was to evaluate the therapeutic efficacy of monophage (Kpn1, Kpn2, Kpn3, Kpn4 and Kpn5) in comparison to phage cocktail in resolving the course of burn wound infection in mice. Although, animals receiving monophage therapy exhibited efficacy in resolving the course of infection but phage cocktail was highly effective in arresting the entire infection process (bacterial load, wound contraction, skin myeloperoxidase activity, collagen formation and histopathological analysis).In comparison to untreated control mice, a significant reduction in bacterial load to 4.32, 4.64, 4.42, 4.11 and 4.27 log CFU/ml in Kpn1, Kpn2, Kpn3 Kpn4 and Kpn5 treated animals was obtained respectively was on peak day (3rd day). However, the group receiving phage cocktail (group 7) showed maximum reduction in bacterial load in the skin tissue. The bacterial load was significantly reduced to 3.01 log CFU/ml on peak day. This accounts for a significant reduction of 6 log cycles (p < 0.01) as compared to that of untreated control animals where a peak of 8.81 log CFU/ml was seen followed by steady decrease thereafter. Thus, phage cocktail gave maximum protection against burn wound infection by K. pneumoniae B5055. Compared to any single phage, phage cocktail significantly checked the emergence of resistant mutants. Hence this approach can serve as an effective strategy in treating Klebsiella mediated burn wound infections in individuals who do not respond to conventional antibiotic therapy.

Efficacy of two Staphylococcus aureus phage cocktails in cheese production

Staphylococcus aureus is one of the most prevalent pathogenic bacteria contaminating dairy products. In an effort to reduce food safety risks, virulent phages are investigated as antibacterial agents to control foodborne pathogens. The aim of this study was to compare sets of virulent phages, design phage cocktails, and use them in a cocktail to control pathogenic staphylococci in cheese. Six selected phages belonging to the three Caudovirales families (Myoviridae, Siphoviridae, Podoviridae) were strictly lytic, had a broad host range, and did not carry genes coding for virulence traits in their genomes. However, they were sensitive to pasteurization. At MOI levels of 15, 45, and 150, two anti-S. aureus phage cocktails, each containing three phages, one from each of the three phage families, eradicated a 106CFU/g S. aureus population after 14days of Cheddar cheese curd ripening at 4°C. The use of these phages did not trigger over-production of S. aureus enterotoxin C. The use of phage cocktails and their rotation may prevent the emergence of phage resistant bacterial strains.

Efficiency of phage cocktails in the inactivation of Vibrio in aquaculture

Abstract Aquaculture is one of the fastest growing sectors in the world, having a very important role in the economy. However, the losses associated with bacterial infections such as vibriosis, lead to huge economic costs. The regular use of antibiotics in aquaculture has resulted in the development of resistant strains, which have contributed to the inefficacy of antibiotics. To reduce the risk of the development and spreading of microbial resistance and to control the fish diseases in aquaculture, alternative strategies must be developed. Phage therapy can be an eco-friendly alternative to prevent and control pathogenic bacteria in aquaculture. However, phage bacterial resistance is already well documented but the use of phage cocktails can overcome this drawback. The aim of this study was to evaluate the efficiency of cocktails of two and three phages of Vibrio parahaemolyticus (VP-1, VP-2 and VP-3) to control Vibrio in aquaculture. All phages were effective against V. parahaemolyticus, however, the VP-3 phage was the most efficient one (additional reduction of more 2 log when compared with the other two phages). The use of cocktails with two and three phages was significantly more effective (reductions of 4 log after 2 h) than the use of VP-1 and VP-2 phages alone (reductions of 0.8 log after 2 h), however, the efficiency of VP-3 phage was similar for the phage alone and for the phage in the cocktails (reduction of 3.8 log and 4.2 log for VP-3 phage alone and in cocktails, respectively, after 8 and 6 h). All phages remained viable for a long time (at least 5–7 months) in marine water. The VP-3 phage presented a larger burst size and a shorter latent period (42 and 40 min, respectively) than the other two phages (9 and 120 min and 15 and 90 min, respectively, for VP-1 and VP-2 phages). Overall, the use of phage cocktails of two or three phages increased the efficiency of phage therapy against Vibrio (more efficient and faster bacterial inactivation), delaying the development of resistance by the bacteria and the use of Vibrio phages with high burst sizes and short lytic cycles also increases the efficiency of phage therapy.

Bacteriophages as an Alternative Strategy for Fighting Biofilm Development

The ability of microbes to form biofilms is an important element of their pathogenicity, and biofilm formation is a serious challenge for today's medicine. Fighting the clinical complications associated with biofilm formation is very difficult and linked to a high risk of failure, especially in a time of increasing bacterial resistance to antibiotics. Bacterial species most commonly isolated from biofilms include coagulase-negative staphylococci, Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Proteus mirabilis, Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter spp. The frequent failure of antibiotic therapy led researchers to look for alternative methods and experiment with the use of antibacterial factors with a mechanism of action different from that of antibiotics. Experimental studies with bacteriophages and mixtures thereof, expressing lytic properties against numerous biofilm-forming bacterial species showed that bacteriophages may both prevent biofilm formation and contribute to eradication of biofilm bacteria. A specific role is played here by phage depolymerases, which facilitate the degradation of extracellular polymeric substances (EPS) and thus the permeation of bacteriophages into deeper biofilm layers and lysis of the susceptible bacterial cells. Much hope is placed in genetic modifications of bacteriophages that would allow the equipping bacteriophages with the function of depolymerase synthesis. The use of phage cocktails prevents the development of phage-resistant bacteria.

Bacteriophage-Resistant Mutants in Yersinia pestis: Identification of Phage Receptors and Attenuation for Mice

BackgroundBacteriophages specific for Yersinia pestis are routinely used for plague diagnostics and could be an alternative to antibiotics in case of drug-resistant plague. A major concern of bacteriophage therapy is the emergence of phage-resistant mutants. The use of phage cocktails can overcome this problem but only if the phages exploit different receptors. Some phage-resistant mutants lose virulence and therefore should not complicate bacteriophage therapy.Methodology/Principal FindingsThe purpose of this work was to identify Y. pestis phage receptors using site-directed mutagenesis and trans-complementation and to determine potential attenuation of phage-resistant mutants for mice. Six receptors for eight phages were found in different parts of the lipopolysaccharide (LPS) inner and outer core. The receptor for R phage was localized beyond the LPS core. Most spontaneous and defined phage-resistant mutants of Y. pestis were attenuated, showing increase in LD50 and time to death. The loss of different LPS core biosynthesis enzymes resulted in the reduction of Y. pestis virulence and there was a correlation between the degree of core truncation and the impact on virulence. The yrbH and waaA mutants completely lost their virulence.Conclusions/SignificanceWe identified Y. pestis receptors for eight bacteriophages. Nine phages together use at least seven different Y. pestis receptors that makes some of them promising for formulation of plague therapeutic cocktails. Most phage-resistant Y. pestis mutants become attenuated and thus should not pose a serious problem for bacteriophage therapy of plague. LPS is a critical virulence factor of Y. pestis.

Bacteriophage for Biocontrol of Foodborne Pathogens: Calculations and Considerations

The use of phage or phage products in food production has recently become an option for the food industry as a novel method for biocontrol of unwanted pathogens, enhancing the safety of especially fresh and ready-to-eat food products. While it can be expected that many more phage products currently under development might become available in the future, several questions may be raised concerning the use of such products, regarding both immediate and long-term efficacy, consumer safety, and application methods. The available evidence suggests that, with a few caveats, safety concerns have been satisfactorily addressed. Answers concerning efficacy are more complex, depending on particular applications or the target pathogens. To ensure long-term efficacy beyond what can be tested on a laboratory scale, food safety concepts employing phages will have to be well-thought out and may involve rotation schemes as used with bacterial starter cultures, the use of phage cocktails, or application of phages combined with other antimicrobials. This review will discuss these issues on the basis of the available literature as well as providing an outlook on the potential of phages in future applications.

T4 phages against Escherichia coli diarrhea: Potential and problems

A combination of in vitro and in vivo experiments with comparative phage genomics was used for the rational design of a phage cocktail against E. coli diarrhea. Orally applied T4 coliphages representing three different subgroups (T4-, RB49- and JS98-like phages) had no negative impact on the murine gut microbiota. T4 phages were found with high titers in the cecum and colon and lower titers in the small intestine, but were not detected in the blood, liver or spleen. No adverse effects were observed after one-month exposure to phage nor were serum anti-T4 antibodies detected. T4 phages belonging to the same subgroup showed closely related genomes that differed by 12 (phage JS10 vs. JS98 reference) to 17 (phage JSE vs. RB49 reference) insertion/deletions mostly representing single small ORFs. Bioinformatic analysis did not reveal undesired genes in the T4 genomes. Sequence variability was seen over the tail fibre genes, but the variability did not correlate with phage host range. The investigated T4 phages were not only species- but also strain-specific, necessitating the use of phage cocktails consisting of 10 and 16 T4 phage isolates to cover half to two thirds of E. coli strains representing the five main pathotypes isolated from diarrhea patients.