Characterization by 16S rRNA sequence analysis of pseudomonads causing blotch disease of cultivated Agaricus bisporus

Bacterial blotch of Agaricus bisporus has typically been identified as bein g caused by either Pseudomonas tolaasii (brown blotch) or Pseudomonas ginge ri (ginger blotch). To address the relatedness of pseudomonads able to indu ce blotch, a pilot study was initiated in which pseudomonads were selective ly isolated from mushroom farms throughout New Zealand. Thirty-three pseudo monad isolates were identified as being capable of causing different degree s of discoloration (separable into nine categories) of A. bisporus tissue i n a bioassay. These isolates were also identified as unique using repetitiv e extragenic palindromic PCR and biochemical analysis. Relationships betwee n these 33 blotch-causing organisms (BCO) and a further 22 selected pseudom onad species were inferred by phylogenetic analyses of near-full-length 16S rRNA gene nucleotide sequences. The 33 BCO isolates were observed to be di stributed throughout the Pseudomonas fluorescens intrageneric cluster. Thes e results show that in addition to known BCO (P. tolaasii, P. gingeri, and Pseudomonas reactans), a number of diverse pseudomonad species also have th e ability to cause blotch diseases with various discolorations. Furthermore , observation of ginger blotch discoloration of A. bisporus being independe ntly caused by many different pseudomonad species impacts on the homogeneit y and classification of the previously described A gingeri.