05); these observations correlated
with a significant reduction in lesion intensity (p < 0.001) on mushrooms treated with 2.9 × 106 and 1.4 × 107 PFU B. bacteriovorus Nutlin-3a clinical trial HD100 (mean = 0.010 1/PV in both cases) compared with mushrooms inoculated with P. tolaasii 2192T alone (mean = 0.014 1/PV). Despite this significant reduction in lesion intensity, the total number of CFU recovered from B. bacteriovorus HD100 treated mushrooms onto King’s Medium B was high, suggesting that the bacteria recovered from seemingly similar, beige-coloured colonies on the King’s Medium B plates were not solely pathogenic P. tolaasii 2192T, but might include other species indigenous to the mushroom pileus surface that are not well preyed upon by B. bacteriovorus HD100, as observed in SEM images of mushroom tissue to which King’s medium B broth was added alone. Figure 4 Bacterial CFU numbers recovered from P. tolaasii -inoculated mushrooms in the presence and absence of Bdellovibrio . Lesion intensities and number of bacterial colony forming units (CFU) recovered from mushroom pilei subject to three different treatments detailed to the right. Each P. tolaasii
2192T inoculation contained 1.7 × 106 CFU. Images of mushrooms with typical: high, mean, and low intensity lesions in each group are shown below the graph. Horizontal black bars indicate the mean values for Crenolanib mouse lesion intensity/CFU count in each treatment group. Student’s t-test of significance between B .bacteriovorus-treated and non-treated mushrooms inoculated with P. tolaasii 2192T: *p <0.05, ***p <0.001. Enterobacterspecies are present on the surface of some commercially produced supermarket mushrooms The number of CFU recovered from the mushrooms that were treated with B. bacteriovorus HD100 after inoculation
with P. tolaasii was relatively high compared to mushrooms inoculated with P. tolaasii alone. To confirm the identity of the bacteria seen in Figures 3d and e and recovered from supermarket mushroom tissue pre-treated with B. bacteriovorus HD100 before P. tolaasii 2192T at both 2.9 × 106 and 1.4 × 107 PFU ml−1, 20 colonies taken from the King’s medium B agar plates used to enumerate bacterial CFU, recovered from the treated mushroom tissue of two mushrooms in each group, were grown on buy PF-02341066 Coliform Chromogenic agar (oxoid). This agar contains two chromogenic substrates that turn almost purple when cleaved by the enzymes glucorinidase and galactosidase, which are both present in coliforms such as E. coli, and absent from Pseudomonads (including P. tolaasii); all 20 colonies were pigmented purple indicating them as coliform, closely related to E. coli, and therefore as indigenous species to the mushroom pileus, and distinctly different to P. tolaasii 2192T , which produced straw coloured colonies on the agar. Three of these coliform isolates were identified by 16 s rDNA sequencing as members of the Enterobacter genus using the BLAST online tool (http://blast.ncbi.nlm.nih.gov/Blast.