Identification and Characterization of Hfq and Hfq-Regulated Small RNAs in Erwinia amylovora

Small RNAs (sRNAs) are important post-transcriptional regulators in bacteria. Hfq is an RNA chaperone that enhances the stability of sRNAs and facilitates their regulatory function. In this study, we identified Hfq as a critical virulence regulator in E. amylovora. The deletion of hfq in E. amylovora Ea1189 resulted in significantly reduced virulence in immature pears and apple shoots. Hfq also regulates multiple virulence determinants, including motility, amylovoran production, attachment, and type III secretion and translocation. Genome-wide identification of sRNAs was conducted combining both bioinformatics analysis and Illumina deep sequencing. A total of 42 sRNAs, including 30 novel sRNAs, were identified in E. amylovora. Mutagenesis and virulence testing of 22 sRNAs showed that two of them, ArcZ and RprA, are required for the full virulence of E. amylovora. The regulatory mechanism of motility by ArcZ and Hfq was also characterized. ArcZ and Hfq both negatively control motility. We determined that this negative regulation occurs through the repression of the translation of flhDC and rcsC. An eleven-nucleotide base-pairing between ArcZ and the 5'-untranslated region of the flhDC mRNA was identified as the critical sequence for the translational control of flhDC by ArcZ.