Multiplex Touchdown PCR for Rapid Typing of the Opportunistic Pathogen Propionibacterium acnes

Emma Barnard, Istvan Nagy, Judit Hunyadkurti, Sheila Patrick, Andrew McDowell

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

The opportunistic human pathogen Propionibacterium acnes is composed of a number of distinct phylogroups, designated types IA1, IA2, IB, IC, II, and III, which vary in their production of putative virulence factors, their inflammatory potential, and their biochemical, aggregative, and morphological characteristics. Although multilocus sequence typing (MLST) currently represents the gold standard for unambiguous phylogroup classification and individual strain identification, it is a labor-intensive and time-consuming technique. As a consequence, we developed a multiplex touchdown PCR assay that in a single reaction can confirm the species identity and phylogeny of an isolate based on its pattern of reaction with six primer sets that target the 16S rRNA gene (all isolates), ATPase (types IA1, IA2, and IC), sodA (types IA2 and IB), atpD (type II), and recA (type III) housekeeping genes, as well as a Fic family toxin gene (type IC). When applied to 312 P. acnes isolates previously characterized by MLST and representing types IA1 (n=145), IA2 (n=20), IB (n=65), IC (n=7), II (n=45), and III (n=30), the multiplex displayed 100% sensitivity and 100% specificity for detecting isolates within each targeted phylogroup. No cross-reactivity with isolates from other bacterial species was observed. This multiplex assay will provide researchers with a rapid, high-throughput, and technically undemanding typing method for epidemiological and phylogenetic investigations. It will facilitate studies investigating the association of lineages with various infections and clinical conditions, and it will serve as a prescreening tool to maximise the number of genetically diverse isolates selected for downstream higher-resolution sequence-based analyses.
LanguageEnglish
Pages1149-1155
Number of pages6
JournalJournal of Clinical Microbiology
Volume53
Issue number4
Early online date28 Jan 2015
DOIs
Publication statusE-pub ahead of print - 28 Jan 2015

Fingerprint

Multilocus Sequence Typing
Propionibacterium acnes
Multiplex Polymerase Chain Reaction
Epidemiologic Methods
Essential Genes
Virulence Factors
Phylogeny
rRNA Genes
Sequence Analysis
Adenosine Triphosphatases
Research Personnel
Sensitivity and Specificity
Infection
Genes
N-(phosphonomethyl)glycine trimethylsulfonium salt

Keywords

  • Propionibacterium acnes
  • multiplex PCR
  • phylogroups

Cite this

Barnard, Emma ; Nagy, Istvan ; Hunyadkurti, Judit ; Patrick, Sheila ; McDowell, Andrew. / Multiplex Touchdown PCR for Rapid Typing of the Opportunistic Pathogen Propionibacterium acnes. In: Journal of Clinical Microbiology. 2015 ; Vol. 53, No. 4. pp. 1149-1155.
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Multiplex Touchdown PCR for Rapid Typing of the Opportunistic Pathogen Propionibacterium acnes. / Barnard, Emma; Nagy, Istvan; Hunyadkurti, Judit; Patrick, Sheila; McDowell, Andrew.

In: Journal of Clinical Microbiology, Vol. 53, No. 4, 28.01.2015, p. 1149-1155.

Research output: Contribution to journalArticle

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