The impact of microbial diseases on crop production coupled with increasing antibiotic
resistance emphasizes the need for alternative agricultural methods that can reduce
environmental impact and do not rely on chemical pesticides. Microbial biocontrol agents
(mBCAs) could potentially provide effective and safe strategies to overcome current
constraints. The Greenlandic soil-bacterium Pseudomonas fluorescens In5 is a promising
biocontrol agent that has previously been shown to produce bioactive compounds against
fungal pathogens1,2. Genome sequencing and analysis of In5 identified large secondary
metabolite biosynthesis gene clusters. A combination of random and targeted mutagenesis,
together with MALDI-TOF imaging mass spectrometry, linked two non-ribosomal peptides
(NRPs) designated nunapeptin and nunamycin respectively, to antifungal activity against
Rhizoctonia solani, Pythium aphanidermatum and Fusarium graminearum1, 2. In order to
unravel the complex genetic regulation of these large NRP synthetase gene clusters,
antisense RNAs (asRNAs) and CRISPR/Cas9 based systems are being tested and
developed as tools to target transcripts of interest and elucidate gene function3, 4. To
investigate the effect of purified nunamycin and nunapeptin at the omics level against
pathogenic fungi, an NRP production platform is being developed which, could additionally
provide a source of antifungal compounds for industrial applications (e.g. food production,
pharmaceutical, personal care). Methods for direct cloning based on either linear-linear
homologous recombination (LLHR) or direct assembly methods (e.g. Gibson assembly) will
be used to capture, clone and refactor In5 secondary metabolite gene clusters5
. The overall
aim of this project is to understand how mBCAs communicate in complex communities, the
conditions under which these antifungal peptides are synthesised and the compound effects
on fungal pathogens. The ultimate goal is to develop a novel and sustainable biocontrol
technology.