Molecular genetic modifications to biocontrol agents
A very wide variety of genetic approaches have been used to genetically engineer BCAs with improved biocontrol or plant colonising ability, and these approaches can be grouped in three categories: i) deletion or mutation of existing genes; ii) alteration of gene regulation; iii) introduction of heterologous genes. Selected examples of these approaches are given below.
Deletion or mutation of existing genes
Agrobacterium radiobacter K84 is a well-described BCA of crown gall that is sold worldwide (Kerr, 1980). A transfer (Tra-) mutant of Agrobacterium K84 (designated K1026) was constructed to prevent the possible transfer of pAgK84 encoding agrocin 84 to Agrobacterium tumefaciens, which could result in the pathogen becoming resistant to the BCA (Jones et al., 1988). The recombinant strain K1026 is as effective as the wild type and is used commercially (Jones and Kerr, 1989).
Another excellent example of this type of genetic modification involves biocontrol of ice nucleating bacteria by an ice nucleating deficient Pseudomonas syringae (Hirano and Upper, 2000). An Ice strain of Pseudomonas syringae was constructed by deleting a fragment of the ice gene, followed by marker exchange of the mutated gene into the wild type. This engineered derivative was the first recombinant microbe deliberately released into the environment. Application of Ice- mutants reduced populations of Ice+ Pseudomonas syringae on potato and strawberry 50-fold by pre-emptive exclusion and reduced frost damage in the field (Lindow, 1995; Lindow and Panopoulos, 1988). The Ice- strain faced a difficult path through regulatory, social and political obstacles prior to field release, which contrasted strikingly with the release of Agrobacterium K1026, which faced little resistance.
Finally, Barahona et al. (2011) constructed a triple mutant of Pseudomonas fluorescens F113 in the genes sadB, wspR and kinB, resulting in hypermotility and better root colonisation. In addition, the mutant strain had improved biocontrol activity against Fusarium oxysporum f. sp. Radicis-lycopersici on tomato and Phytophthora cactorum on strawberry as compared to F113.