High quality genome annotation and expression visualisation of a mupirocin-producing bacterium.

Abstract

Pseudomonas strain NCIMB10586, in the P. fluorescens subgroup, produces the polyketide antibiotic mupirocin, and has potential as a host for industrial production of a range of valuable products. To underpin further studies on its genetics and physiology, we have used a combination of standard and atypical approaches to achieve a quality of the genome sequence and annotation, above current standards for automated pathways. Assembly of Illumina reads to a PacBio genome sequence created a retrospectively hybrid assembly, identifying and fixing 415 sequencing errors which would otherwise affect almost 5% of annotated coding regions. Our annotation pipeline combined automation based on related well-annotated genomes and stringent, partially manual, tests for functional features. The strain was close to P. synxantha and P. libaniensis and was found to be highly similar to a strain being developed as a weed-pest control agent in Canada. Since mupirocin is a secondary metabolite whose production is switched on late in exponential phase, we carried out RNAseq analysis over an 18 h growth period and have developed a method to normalise RNAseq samples as a group, rather than pair-wise. To review such data we have developed an easily interpreted way to present the expression profiles across a region, or the whole genome at a glance. At the 2-hour granularity of our time-course, the mupirocin cluster increases in expression as an essentially uniform bloc, although the mupirocin resistance gene stands out as being expressed at all the time points.