Transcriptomic analysis comparing the spontaneous acapsular strain AL1114 to the parent capsulated strain AL1115

P. multocida is the causative agent of a wide range of diseases of animals, including fowl cholera in birds. Fowl cholera isolates of P. multocida generally express a capsular polysaccharide composed of hyaluronic acid. There have been reports of spontaneous capsule loss in P. multocida fowl cholera-causing strains but the mechanism by which this occurs has not been determined. In this study, we identified three independent strains that had spontaneously lost the ability to produce capsular polysaccharide. Quantitative RT-PCR showed that these strains had significantly reduced transcription of the capsule biosynthetic genes, but DNA sequence analysis identified no mutations within the cap biosynthetic locus. However, whole genome sequencing of paired capsulated and acapsular strains identified a single nucleotide polymorphism within fis that was present only in the acapsular strain. Sequencing of fis from two independently derived spontaneous acapsular strains showed that each contained a mutation within fis. Complementation of these strains with an intact copy of fis returned normal capsule expression to all strains. Therefore, expression of a functional Fis protein is absolutely required for normal capsule expression in P. multocida.DNA microarray studies comparing one of the acapsular pairs (AL114 to AL1115) identified approximately 30 genes as down-regulated in the mutant; including pfh_B2 which encodes the filamentous hemagglutinin, a known P. multocida virulence factor and the cross protective surface antigen plpE. Biological triplicates of each strain were analysed in a single colour experimental design