A rapid multiplex DNA suspension array method for Salmonella typhimurium subtyping using prophage-related markers.
In this study we developed a preliminary proof of concept of method for Salmonella typhimurium subtyping using multiplex PCR-based phage locus typing and a multiplex Luminex DNA suspension array for product detection. Thirty markers were selected from prophages ST64B, ST64T, ST104, P22, Gifsy-1, sopEΦ and mostly phage-related AFLP fragments, and organised into two multiplex PCRs of 15 markers each. A two-group DNA suspension array was developed using a combination of flow cytometry and Luminex xMAP® technology. To assess its subtyping capability the method was applied to 438 non-epidemiological related S. typhimurium isolates of 56 phage types. Eighty-one profiles were generated. Isolates were divided into sixteen main prophage marker profile types. There was a strong tendency for isolates with the same phage type to have the same or closely related profiles and for groups of phage types to share the same profile. The discriminatory power of this method expressed as the Simpson's Index of Diversity (D) was 0.954. A panel of 12 selected markers achieved almost the same D value (0.952) as the 30 markers. This new method provides an alternative typing scheme for S. typhimurium epidemiological investigations. The developed array is in a high-throughput format which could easily be semi-automated, making the test fast and economical.
Public Health Microbiology Laboratory, Division of Communicable Diseases, Queensland Health Forensic and Scientific Services, 39 Kessels Road, Coopers Plains, Qld 4108, Australia. fangningxia@hotmail.com
Ning-Xia
NX
Fang
Bixing
B
Huang
Lester
L
Hiley
John
J
Bates
John
J
Savill
D015373:Bacterial Typing Techniques; D001434:Bacteriophage Typing / Q000379:methods*; D004269:DNA, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D039002:Prophages / Q000235:genetics* / Q000502:physiology; D012483:Salmonella Phages / Q000235:genetics* / Q000502:physiology; D012486:Salmonella typhimurium / Q000145:classification / Q000235:genetics / Q000302:isolation & purification* / Q000821:virology; D014764:Viral Proteins / Q000235:genetics*
D004269:DNA, Bacterial; D014764:Viral Proteins