High-throughput informative single nucleotide polymorphism-based typing of Neisseria gonorrhoeae using the Sequenom MassARRAY iPLEX platform

Ella Trembizki, Helen Smith, Monica M. Lahra, Marcus Chen, Basil Donovan, Christopher K. Fairley, Rebecca Guy, John Kaldor, David Regan, James Ward, Michael D. Nissen, Theo P. Sloots, David M. Whiley*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

49 Citations (Scopus)


Objectives: Neisseria gonorrhoeae antimicrobial resistance (AMR) is a global problem heightened by emerging resistance to ceftriaxone. Appropriate molecular typing methods are important for understanding the emergence and spread of N. gonorrhoeae AMR. We report on the development, validation and testing of a Sequenom MassARRAY iPLEX method for multilocus sequence typing (MLST)-style genotyping of N. gonorrhoeae isolates. Methods: An iPLEX MassARRAY method (iPLEX14SNP) was developed targeting 14 informative gonococcal single nucleotide polymorphisms (SNPs) previously shown to predict MLST types. The method was initially validated using 24 N. gonorrhoeae control isolates and was then applied to 397 test isolates collected throughout Queensland, Australia in the first half of 2012. Results: The iPLEX14SNP method provided 100% accuracy for the control isolates, correctly identifying all 14 SNPs for all 24 isolates (336/336). For the 397 test isolates, the iPLEX14SNP assigned results for 5461 of the possible 5558 SNPs (SNP call rate 98.25%), with complete 14 SNP profiles obtained for 364 isolates. Based on the complete SNP profile data, there were 49 different sequence types identified in Queensland, with 11 of the 49 SNP profiles accounting for the majority (n = 280; 77%) of isolates. AMR was dominated by several geographically clustered sequence types. Using the iPLEX14SNP method, up to 384 isolates could be tested within 1 working day for less than Aus$10 per isolate. Conclusions: The iPLEX14SNP offers an accurate and high-throughput method for the MLST-style genotyping of N. gonorrhoeae and may prove particularly useful for large-scale studies investigating the emergence and spread of gonococcal AMR.

Original languageEnglish
Pages (from-to)1526-1532
Number of pages7
JournalJournal of Antimicrobial Chemotherapy
Issue number6
Publication statusPublished - 1 Jun 2014
Externally publishedYes

Bibliographical note

Funding Information:
This study was conducted as part of the Gonorrhoea Resistance Assessment by Nucleic Acid Detection (GRAND) study funded by the NHMRC (APP1025517) and the Children’s Health Foundation Queensland.

Funding Information:
We thank Dr Amy Jennison, Christine Doyle, Vicki Hicks and John Bates of Forensic and Scientific Services, Queensland for their assistance with this study. We thank Dr Ralf Moser of Sequenom, Herston for his assistance with the design and testing of the iPLEX14SNP method. This study was conducted as part of the reference work of the National Neisseria Network, Australia and the Australian Gonococcal Surveillance Programme, which is funded by the Australian Government Department of Health and ageing.


  • Gonorrhoea
  • Resistance
  • Sequenom
  • SNP
  • Typing


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