Updated genome assembly and annotation of Paenibacillus larvae, the agent of American foulbrood disease of honey bees

Abstract

Background: As scientists continue to pursue various 'omics-based research, there is a need for high quality data for the most fundamental 'omics of all: genomics. The bacterium Paenibacillus larvae is the causative agent of the honey bee disease American foulbrood. If untreated, it can lead to the demise of an entire hive; the highly social nature of bees also leads to easy disease spread, between both individuals and colonies. Biologists have studied this organism since the early 1900s, and a century later, the molecular mechanism of infection remains elusive. Transcriptomics and proteomics, because of their ability to analyze multiple genes and proteins in a high-throughput manner, may be very helpful to its study. However, the power of these methodologies is severely limited without a complete genome; we undertake to address that deficiency here.

Results: We used the Illumina GAIIx platform and conventional Sanger sequencing to generate a 182-fold sequence coverage of the P. larvae genome, and assembled the data using ABySS into a total of 388 contigs spanning 4.5 Mbp. Comparative genomics analysis against fully-sequenced soil bacteria P. JDR2 and P. vortex showed that regions of poor conservation may contain putative virulence factors. We used GLIMMER to predict 3568 gene models, and named them based on homology revealed by BLAST searches; proteases, hemolytic factors, toxins, and antibiotic resistance enzymes were identified in this way. Finally, mass spectrometry was used to provide experimental evidence that at least 35% of the genes are expressed at the protein level.

Conclusions: This update on the genome of P. larvae and annotation represents an immense advancement from what we had previously known about this species. We provide here a reliable resource that can be used to elucidate the mechanism of infection, and by extension, more effective methods to control and cure this widespread honey bee disease.

Figure 1

Self-alignment of 353 P. larvae contigs of assembly ADZY01000000.(a) MUMmer-generated graph of the P. larvae contigs aligned with themselves under default parameters. Red dots indicate sense matches and blue dots represent antisense matches. (b) Enlarged view of a sample region to demonstrate the common occurrence of alignments at the ends of contigs.

Figure 2

Comparative genomics for aligning P. larvae contigs.MUMmer-generated dot-plot showing P. larvae contigs (y-axis) aligned with the fully-sequenced genomes of (a) P. JDR2 and (b) P. vortex (x-axes). Proteins sequences were used for matching. Red dots indicate same-direction matches and blue dots represent antisense matches. Magnified views of regions from (a) exemplify P. larvae contigs that are (c) highly conserved and (d) poorly conserved with P. JDR2; the same contigs are also compared in P. vortex (e).

Figure 3

Flagellar proteins. Distribution of flagellar proteins (excluding chemotaxis proteins) among flagellated Gram-positive (A) and Gram-negative (B) bacteria. The proteins encoded by the P. larvae genome are boxed in green (the ones that are lacking: in red).