PsaPhageKill - isolation, characterisation and use of lytic bacteriophages against Pseudomonas syringae PV. actinidiae to fight the cancer of kiwifruit: an efficient and ecofriendly alternative

Abstract

The indiscriminate use of antibiotics combined with the adaptive capacity of microorganisms allowed the emergence of microbial strains with varying and increasing levels of antimicrobial resistance (Berquó et al., 2004; Davey et al., 2006; Silva, 2008). The increase in bacterial resistance against various antimicrobial agents limits the therapeutic options available for the effective treatment against infections.In the last decade, the worldwide production of kiwi fruit has been highly affected by Pseudomonas syringae pv. actinidiae (Psa), a phytopathogenic bacterium; this has led to severe economic losses that are seriously affecting the kiwi fruit trade. The available treatments for this disease are still scarce, with the most common involving frequently spraying the orchards with copper derivatives, in particular cuprous oxide (Cu2O). However, these copper formulations should be avoided due to their high toxicity; therefore, it is essential to search for new approaches for controlling Psa.The global population claim for new safe alternatives to antibiotics that would reduce the apparition of resistant bacteria and, at the same time, would permit a sustainable local agriculture production without the use of the traditional environmental toxic products. This research project proposal aims to develop and establish a highly accurate approach to combat and prevent the occurrence of the bacterial cancer in kiwifruit plants. To achieve that, one will use specific viruses of bacteria (bacteriophages) to eliminate the causative agent of this disease and responsible for substantial economic losses worldwide, the phytopathogen Pseudomonas syringae pv. actinidiae. To the best of my knowledge, this will be the first attempt to start the development and application of a phage therapy in the management of this emergent agricultural disease. Hence, it is expected that the results gathered will introduce novelty and significant advances in this research area. Moreover, the kiwifruit industry will benefit from this green technology, which will contribute to a sustainable growth in a highly globalized marketplace.The proposed concept is not new, existing from about 3.5 billion years, having been initiated when bacteria and bacteriophages established a predator-prey balance. Bacteriophages are the most abundant organisms on our planet (are estimated at more than 1031 particles) being completely safe for all organisms including humans, except for their target bacterial hosts. The idea embodied in the research project PsaPhageKill is unconventional and a highly creative approach, since it will join the solution (phage) with the problem (bacteria) in order to eliminate the latter. One intends to establish the proof-of-concept using either a single bacteriophage and a cocktail of bacteriophages lytic against Pseudomonas syringae pv. actinidiae, originating from environmental isolates. The physico-chemical and biological characterization of the isolated bacteriophage particles will include morphological analysis via SEM/TEM, DRX, FTIR, SDS-PAGE electrophoresis, thermal analysis by TGA and DSC, lytic spectra, detection of prophages in the host, stability and survival of the bacteriophage particles under abiotic stress (UV radiation), efficiency of plating, OSGC growth curve, adsorption curve, DNA analyses.