Adapting a Phage to Combat Phage Resistance

Abstract

Phage therapy is becoming a widely recognized alternative for fighting pathogenic bacteria due to increasing antibiotic resistance problems. However, one of the common concerns related to the use of phages is the evolution of bacterial resistance against the phages, putatively disabling the treatment. Experimental adaptation of the phage (phage training) to infect a resistant host has been used to combat this problem. Yet, there is very little information on the trade-offs of phage infectivity and host range. Here we co-cultured a myophage FCV-1 with its host, the fish pathogen Flavobacterium columnare, in lake water and monitored the interaction for a one-month period. Phage resistance was detected within one day of co-culture in the majority of the bacterial isolates (16 out of the 18 co-evolved clones). The primary phage resistance mechanism suggests defense via surface modifications, as the phage numbers rose in the first two days of the experiment and remained stable thereafter. However, one bacterial isolate had acquired a spacer in its CRISPR (Clustered Regularly Interspaced Short Palindromic Repeat)-Cas locus, indicating that also CRISPR-Cas defense was employed in the phage-host interactions. After a week of co-culture, a phage isolate was obtained that was able to infect 18 out of the 32 otherwise resistant clones isolated during the experiment. Phage genome sequencing revealed several mutations in two open reading frames (ORFs) likely to be involved in the regained infectivity of the evolved phage. Their location in the genome suggests that they encode tail genes. Characterization of this evolved phage, however, showed a direct cost for the ability to infect several otherwise resistant clones—adsorption was significantly lower than in the ancestral phage. This work describes a method for adapting the phage to overcome phage resistance in a fish pathogenic system.

Coevolutionary phage training leads to greater bacterial suppression and delays the evolution of phage resistance

“…Altogether, these results contribute to a small but growing body of work (23,48,49) in support of coevolutionary phage training as a means to improve the efficacy of phage therapeutics. Furthermore, they reveal the mechanisms by which this improved efficacy can be achieved: bacteria appear less able to resist trained phage and trained phage are more able to adapt to counter their hosts.…”

“…By reciprocally adapting to changes in their hosts (coevolution), phages have maintained the ability to infect their hosts for millennia. Many have proposed harnessing this inherent, evolutionary potential by preemptively coevolving phages with target bacterial prey (22)(23)(24). Proponents of this "phage training" approach suggest that, by experiencing the ways their host can evolve resistance, trained phages will evolve to counter host defenses.…”

Prevalence of genetically similarFlavobacterium columnarephages across aquaculture environments reveals a strong potential for pathogen control

“…FCOV-F28 was able to infect genetic groups C and G, and comparison to FCOV-F25 revealed several non-synonymous nucleotide changes in the previously speculated tail fiber genes, ORFs35 and 36 (Laanto et al, 2017). In our previous study these same ORFs accumulated several mutations during a co-culture of F. columnare and phage FCV-1, leading to change in host range (Laanto et al, 2020). Previous data with other species also suggests point mutations in tail fiber proteins increase phage infectivity (Uchiyama et al, 2011;Boon et al, 2020).…”

Bacteriophages as Potential Tools for Use in Antimicrobial Therapy and Vaccine Development

“…An alternative solution to the growing resistance of bacteria to antibiotics can be phage therapy, based on lytic phages or a combination of phages and antibiotics [ 4 , 5 , 6 ]. This antibacterial technology has been known for 100 years and can be useful not only in combating antibiotic-resistant and tolerant bacteria but also in treating infections related to biofilm formation as well as against spore formers [ 7 , 8 , 9 , 10 , 11 ].…”