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Pseudomonas syringae  pv.  phaseolicola  is the causative agent of halo blight in the common bean,  Phaseolus vulgaris .  P. syringae  pv.  phaseolicola  race 4 strain 1302A contains the avirulence gene  avrPphB  (syn.  hopAR1 ), which resides on PPHGI-1, a 106 kb genomic island. Loss of PPHGI-1 from  P. syringae  pv.  phaseolicola  1302A following exposure to the hypersensitive resistance response (HR) leads to the evolution of strains with altered virulence. Here we have used fluorescent protein reporter systems to gain insight into the mobility of PPHGI-1. Confocal imaging of dual-labelled  P. syringae  pv.  phaseolicola  1302A strain, F532 (dsRFP in chromosome and eGFP in PPHGI-1), revealed loss of PPHGI-1::eGFP encoded fluorescence during plant infection and when grown  in vitro  on extracted leaf apoplastic fluids. Fluorescence-activated cell sorting (FACS) of fluorescent and non-fluorescent PPHGI-1::eGFP F532 populations showed that cells lost fluorescence not only when the GI was deleted, but also when it had excised and was present as a circular episome. In addition to reduced expression of eGFP, quantitative PCR on sub-populations separated by FACS showed that transcription of other genes on PPHGI-1 ( avrPphB  and  xerC ) was also greatly reduced in F532 cells harbouring the excised PPHGI-1::eGFP episome. Our results show how virulence determinants located on mobile pathogenicity islands may be hidden from detection by host surveillance systems through the suppression of gene expression in the episomal state.

The Stealth Episome: Suppression of Gene Expression on the Excised Genomic Island PPHGI-1 from Pseudomonas syringae pv. phaseolicola