In image e, B. bacteriovorus HD100 (blue) are shown attached at one pole to P. tolaasii 2192T (yellow), a crucial first step in the predatory process. Images d and e both show rounded P. tolaasii 2192T cells, characteristic of the bdelloplast structures formed after Bdellovibrio invades the host cell and begins replication. 1 μm scale bar shown. Where B. bacteriovorus HD100 was added to the mushroom surface both before (Figure 3e) and after P. tolaasii 2192T (Figure 3d), B. bacteriovorus

HD100 attachment CP-690550 to P. tolaasii 2192T cells was observed: a crucial first step in the predatory process. In addition, bdelloplasts, the rounded, dead P. tolaasii structures in which Bdellovibrio establish, grow and replicate after attachment and invasion, were also observed where B. bacteriovorus HD100 was added before or after P. tolaasii 2192T. Although a valid statistical survey is not possible in these SEM samples, AZD0156 price bdelloplasts were most clearly visible on the mushroom surface where B. bacteriovorus HD100 was added before

P. tolaasii 2192T (Figure 3d). This correlates with the greater reduction in lesion intensity measurements on mushrooms where B. bacteriovorus HD100 was added before P. tolaasii 2192T (Figure 2): Bdellovibrio attachment to prey and subsequent bdelloplast formation may be easier, and occur more rapidly, where P. tolaasii cells have not had time to accumulate, adapt and adhere to the mushroom surface, preventing P. tolaasii from producing as much tolaasin, and thus reducing the extent of the characteristic brown blotch symptoms. A King’s Medium B control addition to LY2835219 the pileus resulted in the growth of different types of bacterial cells, with different morphologies that were distinct from that of P. tolaasii 2192T & B. bacteriovorus HD100 (Figure 3f); however, typically, no bacterial cells were observed on untreated mushroom tissue (Figure 3a). This indicates that the supermarket mushrooms carry a small, indigenous bacterial microflora that replicates about readily in added growth medium, which may impact upon P. tolaasii CFU numbers recovered from experimentally inoculated tissue, as described

below. Application of Bdellovibriobefore inoculation with P. tolaasiireduced the number of P. tolaasiiin infected mushroom tissue To determine whether the reduction in lesion intensity after treatment with B. bacteriovorus HD100 correlated with a reduction in P. tolaasii 2192T cell numbers, CFU were recovered and enumerated from mushroom tissue that had been inoculated with P. tolaasii 2192T and pre-treated with B. bacteriovorus HD100, compared with a P. tolaasii 2192T inoculated, non-B. bacteriovorus HD100 treated control (Figure 4). A mean number of 4.5 × 107 and 3.9 × 107 CFU were recovered from mushrooms pre-treated with 2.9 × 106 or 1.4 × 107 PFU live B. bacteriovorus HD100 respectively, which were both significantly lower than the mean 1.9 × 108 CFU recovered from mushrooms inoculated with P.