The opportunistic pathogen Propionibacterium acnes: insights into typing, human disease, clonal diversification and CAMP factor evolution

Andrew McDowell, István Nagy, Márta Magyari, Emma Barnard, Sheila Patrick, Andrew McDowell, István Nagy, Márta Magyari, Emma Barnard, Sheila Patrick

Abstract

We previously described a Multilocus Sequence Typing (MLST) scheme based on eight genes that facilitates population genetic and evolutionary analysis of P. acnes. While MLST is a portable method for unambiguous typing of bacteria, it is expensive and labour intensive. Against this background, we now describe a refined version of this scheme based on two housekeeping (aroE; guaA) and two putative virulence (tly; camp2) genes (MLST4) that correctly predicted the phylogroup (IA1, IA2, IB, IC, II, III), clonal complex (CC) and sequence type (ST) (novel or described) status for 91% isolates (n = 372) via cross-referencing of the four gene allelic profiles to the full eight gene versions available in the MLST database (http://pubmlst.org/pacnes/). Even in the small number of cases where specific STs were not completely resolved, the MLST4 method still correctly determined phylogroup and CC membership. Examination of nucleotide changes within all the MLST loci provides evidence that point mutations generate new alleles approximately 1.5 times as frequently as recombination; although the latter still plays an important role in the bacterium's evolution. The secreted/cell-associated 'virulence' factors tly and camp2 show no clear evidence of episodic or pervasive positive selection and have diversified at a rate similar to housekeeping loci. The co-evolution of these genes with the core genome might also indicate a role in commensal/normal existence constraining their diversity and preventing their loss from the P. acnes population. The possibility that members of the expanded CAMP factor protein family, including camp2, may have been lost from other propionibacteria, but not P. acnes, would further argue for a possible role in niche/host adaption leading to their retention within the genome. These evolutionary insights may prove important for discussions surrounding camp2 as an immunotherapy target for acne, and the effect such treatments may have on commensal lineages.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Sliding window analysis of nucleotide…
Figure 1. Sliding window analysis of nucleotide diversity (π) within the concatenated sequence of all eight MLST loci (4253 bp).
Analysis was carried out using a window size of 200(aroE, guaA, sodA, tly and camp2) selected for further analysis are indicated.
Figure 2. Neighbour-net splits graph of allelic…
Figure 2. Neighbour-net splits graph of allelic profiles from all 38 STs identified upon MLST analysis of 87 P. acnes isolates.
STs representing all phylogroups were identified from this cohort of isolates. Parallelogram structures indicative of recombination events are evident within the major type I and II divisions.
Figure 3. Association of P. acnes phylogroups…
Figure 3. Association of P. acnes phylogroups with different clinical conditions and normal skin.
Data was obtained from MLST analysis of 372 isolates.
Figure 4. eBURST population snapshot of the…
Figure 4. eBURST population snapshot of the current P. acnes MLST database.
To date, a total of nine clonal complexes, where the isolates share 7/8 loci with at least one other ST in the group, and 21 singletons have been identified. The frequency of each ST within the isolate database is indicated by the size of each circle. Founding genotypes are highlighted in blue and sub-founders in yellow. The previously described separate complexes CC1 and CC4 now form a large single complex with ST1 as the founder (100% bootstrap) and ST4 (98% bootstrap) as a sub-founder; these two clusters are linked via ST105. Note, the spacing between the singletons and clonal complexes is not related to the genetic distance between them.
Figure 5. Alignment of CAMP factor 2…
Figure 5. Alignment of CAMP factor 2 amino acid sequences from P. acnes type strain NCTC737 (type IA1; ST1; CC1) and P. humerusii strains HL044PA1, HL037PA2, HL037PA3 and P08.
Overall identity between the homologues from these two species based on their protein sequence is 92%. The N-terminal putative signal sequence cleavage site occurs at the AHA-VE sequence (start residue position, 26).
Figure 6. MAUVE visualisation of synteny between…
Figure 6. MAUVE visualisation of synteny between P. acnes, P. avidum and P. humerusii genomes.
Pairwise alignments of genomes were generated using the progressive MAUVE algorithm (v2.3.1). The sequence similarity in the pairwise alignment of P. acnes (6609, type IB ST5, CC5) and P. avidum (ATCC25577) was 53.2%. The similarity between P. acnes (6609) and P. humerusii (P08) was 69.3%, and between P. avidum (ATCC25577) and P. humerusii (P08) 52.7%.

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