Increased hyphal branching and growth of ectomycorrhizal fungus Lactarius rufus by the helper bacterium Paenibacillus sp.

Abstract

Paenibacillus sp. EJP₇₃ has been previously demonstrated as a mycorrhization helper bacterium (MHB) for the Lactarius rufus–Pinus sylvestris symbiosis in both laboratory and glasshouse experiments. In the present study, the effect of Paenibacillus sp. EJP₇₃ metabolites on L. rufus EO₃ pre-symbiotic growth was tested in two agar plate-based systems. Specifically, volatile metabolites were investigated using a dual plate system, in which the presence of strain EJP₇₃ resulted in a significant negative effect on L. rufus EO₃ hyphal radial growth but enhanced hyphal branching and reduced internode distance. Soluble metabolites produced by strain EJP₇₃ were tested on L. rufus EO₃ growth in single-agar plate assays by incorporating bacterial cell-free whole or molecular weight fraction spent broth into the agar. Whole spent broth had a negative effect on hyphal growth, whereas a low molecular weight fraction (100–1,000) promoted colony radial growth. Headspace and spent broth analysis of strain EJP₇₃ cultures revealed 2,5-diisopropylpyrazine to be the most significant component. Synthesised 2,5-diisopropylpyrazine and elevated CO2 (2,000 ppm) were tested as specific volatile metabolites in the dual plate system, but neither produced the response shown when strain EJP₇₃ was present. Increased pre-symbiotic hyphal branching leading to increased likelihood of plant infection may be an important MHB mechanism for strain EJP₇₃ . Although the precise signal molecules could not be identified, the work suggests a number of metabolites may work synergistically to increase L. rufus root colonisation.