Dose effect in the dual inoculation of an ectomycorrhizal fungus and a mycorrhiza helper bacterium in two forest nurseries

Abstract

We inoculated disinfected soil at two Douglas-fir bareroot forest nurseries with three doses (8×10 5, 8×10 7 and 8×10 9 cfu m −2) of the rifampin-resistant mycorrhiza helper bacterium Pseudomonas fluorescens strain BBc6R8 and the ectomycorrhizal fungus Laccaria bicolor strain S238N. In one of the two nurseries, two doses of fungal inoculum (50 and 100 mg m −2 dry weight (DW) mycelium entrapped in alginate beads at the constant dose of 1 l m −2) were tested. For all bacterial treatments the density of P. fluorescens BBc6R8 in the soil, determined by dilution plating, dropped below the detection limit (10 −2 cfu g −1 DW soil) 2 weeks after inoculation. Fifteen weeks after inoculation, the introduced bacterium could be detected by enrichment only in the treatments inoculated with the highest bacterial dose. Two years after inoculation, P. fluorescens BBc6R8 could not be detected in the soil of any of the bacterial treatments. Five months after inoculation and sowing, bacterial inoculation significantly increased the percentage of mycorrhizal short roots on plants inoculated with either low or high amounts of L. bicolor, in one of the two nurseries. The lowest bacterial dose increased mycorrhizal colonization from 45 to 70% in plants inoculated with the low amount of fungal inoculum, and from 64 to 77% in plants inoculated with the high amount of fungal inoculum. The lowest bacterial dose increased mycorrhizal colonization more than the highest bacterial dose. The same L. bicolor mycorrhizal index (70%) was obtained with 50 mg m −2 DW mycelium plus the bacterium than with twice this fungal dose and no bacterium (64%). Two years after inoculation, the height of the mycorrhizal Douglas-firs in the other nursery was significantly increased by the lowest bacterial dose (from 40.7 to 42.6 cm). These results indicate that co-inoculating a helper bacterium together with an ectomycorrhizal fungus can be an efficient way of optimizing controlled mycorrhization techniques for the production of high-quality Douglas-fir planting stocks. They also confirm that BBc6R8 acts at a low population density (less than 10 2 cfu g −1 soil); this contrasts with most PGPR effects discussed in the literature, where the minimal inoculation dose of 10 5 cfu g −1 soil is required to obtain the beneficial effect.