Guidelines to Compose an Ideal Bacteriophage Cocktail

Abstract

Correctly designed bacteriophage therapeutics are the cornerstone for a successful outcome of bacteriophage therapy. Here we overview strategies on how to choose bacteriophages and their bacterial hosts at different steps of a bacteriophage cocktail development in order to comply with all quality and safety requirements based on the already existing essentially empirical experience in bacteriophage therapy and current accomplishments in modern biomedical sciences. A modification of the classic Appelmans' method (1922) to assess stability of bacteriophage activity in liquid media is presented in order to improve the overall performance of therapeutic bacteriophages individually and collectively in the cocktail.

Bacteriophage Therapy: Developments and Directions

“…The fixed phage cocktail approach consists of formulating a set combination of phages to address the diversity within a single bacterial pathogen or multiple bacterial pathogens. This approach is employed to develop a phage preparation as an off-the-shelf antibacterial that can be used for treatment and/or prophylaxis [34][35][36][37]. This approach is currently being pursued commercially in Western Europe and the United States because it is most compatible with existing regulatory paths for the clinical development of antimicrobials.…”

“…The laboratory adaptation and genetic engineering of phages for therapeutic purposes is likely to be a significant force that will transform phage therapy [13,36,45,46], and the latter is already a commercial focus in the United States for companies such as Locus Biosciences and Armata Pharmaceuticals. In vitro evolution (or phage adaptation, phage training) to expand or optimize host range is an approach to modify and improve phages that has been used extensively in former Soviet countries and Eastern Europe, and was recently described in the Western literature [37,[47][48][49]. Alteration of phage host range by this approach can also potentially be used to discover novel molecular mechanisms that can be exploited for genetic modification strategies [50].…”

Bacteriophages for Chronic Wound Treatment: From Traditional to Novel Delivery Systems

“…A recently published guideline gathers some strategies to select the phages and their bacterial hosts for the development of an ideal phage cocktail [150]. According to the guideline, a phage cocktail formulation should be: (i) from a natural source (e.g., isolated from different environments such as water, sewage, soil, clinical samples, among others), without genetic manipulation; (ii) constituted of only lytic phages to avoid horizontal transfer of genes of potentially damaging genetic factors, and be well characterized; (iii) active against a broad range of target bacteria (e.g., it is sufficient if the phage lyses 70-80% of the target clinical isolates, but if an individual phage only infects 40-50% of the target strains, mixing with different phages enlarges this range); (iv) able to replicate on the target bacteria and have high rates of adsorption, a short latent period, and a large burst size; (v) constituted of phages with different bacterial cell wall receptor recognition sites to overcome potential bacterial resistance mechanisms; (vi) capable of maintaining the killing ability throughout treatments and storage.…”

“…The authors of these guidelines draw attention to the fact that the mixtures should not be of phages randomly chosen since that combination can result in interference between them, causing even antagonistic effects. The Appelmans' method for the assessment of the complementary activity of phages should be taken into consideration when selecting phages for the cocktail formulations [150]. This method compares the optical density (OD 600 ) of a given bacterial strain infected with a phage mixture against the OD obtained after infection with the individual phages.…”

“…When the OD values of mixtures are lower than the individual phage OD values, the phage mixture shows a positive outcome and, therefore, the phages can be combined. Also, the formulations used for therapy must be produced in a constant process of renovation and adjustment since new bacteria can arise from infection derived from different environments and geographic areas [150].…”