Development of a biopesticide based on Lysobacter capsici AZ78: formulation and fate in the environment

Microbial biological control agents applied on the phyllosphere are exposed to harsh environmental stresses that could negatively affect their persistence. Formulation can be an efficient way to protect them and obtain consistent results under field conditions. In addition, monitoring the populations on the treated plants to understand the ecological fitness of the microbial biocontrol agent requires specific quantitative molecular methods. For this reason, we designed a protocol to develop a suitable formulation for Lysobacter capsici AZ78, a Gram-negative bacterium able to control Plasmopara viticola, and developed a qPCR procedure for monitoring the fate of L. capsici AZ78 in vineyards. Cell mass production was maximized and a harvest of at least 1010 L. capsici AZ78 cells/ml was obtained in a benchtop fermenter. Shelf life of the harvested cells was assessed and viability of L. capsici AZ78 cells stored in distilled water decreased only one order of magnitude after one year of storage at 4°C. We screened compounds capable to protect the bacterial cells against desiccation and UV light irradiation, and to limit the wash-off. The use of a combination of 0.1 % polyethyleneglycol, corn steep liquor and lignosulfonate protected the L. capsici AZ78 population on grapevine leaves and its ability to control P. viticola under field conditions. A specific primer pair for qPCR was developed from a discriminating band obtained by REP-PCR fingerprinting. The qPCR procedure showed that the L. capsici AZ78 population reached 106 cells/gram of leaf after its application and revealed that the use of additives in the tank mix enhanced the persistence of L. capsici AZ78 cells in vineyards.