title: A multiplex oligonucleotide ligation-PCR as a complementary tool for subtyping of Salmonella Typhimurium.
abstract: Subtyping below the serovar level is essential for surveillance and outbreak detection and investigation of Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) and its monophasic variant 1,4,[5],12:i:- (S. 1,4,[5],12:i:-), frequent causes of foodborne infections. In an attempt to overcome the intrinsic shortcomings of currently used subtyping techniques, a multiplex oligonucleotide ligation-PCR (MOL-PCR) assay was developed which combines different types of molecular markers in a high-throughput microsphere suspension array. The 52 molecular markers include prophage genes, amplified fragment length polymorphism (AFLP) elements, Salmonella genomic island 1 (SGI1), allantoinase gene allB, MLVA locus STTR10, antibiotic resistance genes, single nucleotide polymorphisms (SNPs) and phase 2 flagellar gene fljB. The in vitro stability of these markers was confirmed in a serial passage experiment. The validation of the MOL-PCR assay for subtyping of S. Typhimurium and S. 1,4,[5],12:i:- on 519 isolates shows that the method is rapid, reproducible, flexible, accessible, easy to use and relatively inexpensive. Additionally, a 100 % typeability and a discriminatory power equivalent to that of phage typing were observed, and epidemiological concordance was assessed on isolates of 2 different outbreaks. Furthermore, a data analysis method is provided so that the MOL-PCR assay allows for objective, computerised data analysis and data interpretation of which the results can be easily exchanged between different laboratories in an international surveillance network.
authors.affiliation: Department of Microbial and Molecular Systems, KU Leuven, Kasteelpark Arenberg 20 bus 2460, 3001, Leuven, Belgium.
authors.forename: Véronique
authors.initials: V
authors.lastname: Wuyts
authors.forename: Wesley
authors.initials: W
authors.lastname: Mattheus
authors.forename: Nancy H C
authors.initials: NH
authors.lastname: Roosens
authors.forename: Kathleen
authors.initials: K
authors.lastname: Marchal
authors.forename: Sophie
authors.initials: S
authors.lastname: Bertrand
authors.forename: Sigrid C J
authors.initials: SC
authors.lastname: De Keersmaecker
mesh_terms: D054458:Amplified Fragment Length Polymorphism Analysis; D015373:Bacterial Typing Techniques / Q000379:methods*; D044404:Genomic Islands; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D009841:Oligonucleotides / Q000235:genetics; D012480:Salmonella Infections / Q000382:microbiology*; D012486:Salmonella typhimurium / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D009841:Oligonucleotides