EMGM Strain Characterization Working Group

Martin Maiden, University of Oxford, United Kingdom, martin.maiden@zoo.ox.ac.uk
Muhamed-Kheir Taha, Institut Pasteur, France, mktaha@pasteur.fr
Julio Vázquez, Reference Laboratory for Neisserias, National Center for Microbiology, Institute of Health Carlos III, Madrid, Spain, jvazquez@isciii.es
Georgina Tzanakaki, National School of Public Health, Athens, Greece, gtzanakaki@esdy.edu.gr
Paola Stefanelli, Istituto Superiore di Sanità, Rome, Italy, paola.stefanelli@iss.it

This working group has formed from a merger of the Molecular Typing and Antibiotic Resistance working groups.

Molecular Typing

Harmonising the characterisation of meningococcal isolates across Europe has been a major focus of the EMGM's activities since its inception in 1993. The MTWG achieves this by conducting multi-laboratory studies, promoting the sharing of reagents and methods, and organising workshops and training sessions.

Landmark achievements of members of this group include:

1992 - present: The production and distribution of monoclonal antibodies for meningococcal typing.
1996 - 1998: The development of Multilocus sequence typing (MLST).
2000: The establishment of the European Meningococcal MLST Centre as part of the EuMenNet project and the organisation of training workshops.
2003 - 2004: Implementation of unified typing nomenclatures for meningococcal PorA and FetA.
2005: The adoption of PorA and FetA sequence based typing as the future paradigm for European meningococcal isolate characterisation.
2006 - 2007: The development of penA sequencing for the characterisation of meningococcal isolates with reduced susceptibility to penicillin G.
2007: The development of European Meningococcal Epidemiology in Real Time (EMERT).
2009: Development of fHbp typing scheme and database on PubMLST.
2009: Introduction of BIGSdb genomics platform to PubMLST allowing the inclusion of a vast variety of markers.
2010: The development of rpoB sequencing for the characterisation of meningococcal isolates resistant to rifampicin.
2010: The development of tools for whole genome sequencing analysis and comparisons.
2012: Development of ribosomal MLST (rMLST) universal typing scheme.
2013: The development of gyrA sequencing for the characterisation of meningococcal isolates resistant to ciprofloxacin.
2013: Description of Neisseria meningitidis capsule locus with recommendations for update of nomenclature.
2013: Initiation of IBDLabNet European WGS strain collection project.
2014: The development of rplF sequencing for the identification of Neisseria species.

Linked to publications

Bennett JS, Watkins ER, Jolley KA, Harrison OB, Maiden MC. Identifying Neisseria species using the 50S ribosomal protein L6 (rplF) gene. J Clin Microbiol 2014, [Epub ahead of print]
Harrison OB, Claus H, Jiang Y, Bennett JS, Bratcher HB, Jolley KA, Corton C, Care R, Poolman JT, Zollinger WD, Frasch CE, Stephens DS, Feavers I, Frosch M, Parkhill J, Vogel U, Quail MA, Bentley SD, Maiden MC. Description and nomenclature of Neisseria meningitidis capsule locus. Emerg Infect Dis 2013, 19:566-73
Hong E, Thulin Hedberg S, Abad R, Fazio C, Enríquez R, Deghmane AE, Jolley KA, Stefanelli P, Unemo M, Vazquez JA, Veyrier FJ, Taha MK. Target gene sequencing to define the susceptibility of Neisseria meningitidis to ciprofloxacin. Antimicrob Agents Chemother 2013, 57:1961-4
Jolley KA, Bliss CM, Bennett JS, Bratcher HB, Brehony C, Colles FM, Wimalarathna H, Harrison OB, Sheppard SK, Cody AJ, Maiden MC. Ribosomal multilocus sequence typing: universal characterization of bacteria from domain to strain. Microbiology 2012, 158:1005-15
Jolley KA, Maiden MC. BIGSdb: Scalable analysis of bacterial genome variation at the population level. BMC Bioinformatics 2010, 11:595
Brehony C, Wilson DJ, Maiden MC. Variation of the factor H-binding protein of Neisseria meningitidis. Microbiology 2009, 155:4155-69
Brehony C, Jolley KA, Maiden MC. Multilocus sequence typing for global surveillance of meningococcal disease. FEMS Microbiol Rev 2007, 31:15-26
Jolley KA, Brehony C, Maiden MC. Molecular typing of meningococci: recommendations for target choice and nomenclature. FEMS Microbiol Rev 2007, 31:89-96
Russell JE, Jolley KA, Feavers IM, Maiden MC, Suker J. PorA variable regions of Neisseria meningitidis. Emerg Infect Dis 2004, 10:674-8
Thompson EA, Feavers IM, Maiden MC. Antigenic diversity of meningococcal enterobactin receptor FetA, a vaccine component. Microbiology 2003, 149:1849-58
Maiden MC, Bygraves JA, Feil E, Morelli G, Russell JE, Urwin R, Zhang Q, Zhou J, Zurth K, Caugant DA, Feavers IM, Achtman M, Spratt BG. Multilocus sequence typing: a portable approach to the identification of clones within populations of pathogenic microorganisms. Proc Natl Acad Sci USA 1998, 95:3140-5

Antibiotic Resistance

Convener: Muhamed-Kheir Taha, Neisseria Unit, Institut Pasteur, Paris, France, mktaha@pasteur.fr.
Co-convener: Julio Vázquez, Reference Laboratory for Neisserias, National Center for Microbiology, Institute of Health Carlos III, Madrid, Spain, jvazquez@isciii.es.

The aims of the working group on antibiotic resistance are:

Standardization of methods and break point definitions.
Implementation and standardization of molecular tools to determine meningococcal susceptibility to antibiotics.
Implementation of non-culture methods using PCR for detection of antibiotics resistance in clinical samples.

Work within the WP designed an approach to better define breakpoints for the most relevant antibiotics. The approach consists of several steps:

Sequencing of large collections of isolates to define wild type alleles for penA (penicillin G), rpoB (rifampicin), gyrA, parE and parC (ciprofloxacin).
Correlate wild-type genes to MICs to define the epidemiological cut off values.
Use of animal models to explore the impact of increased MIC during experimental infections.

Similar work is intended to characterize ß-lactamase negative ampicillin-resistant (BLNAR) invasive Haemophilus influenzae isolates through implementing of ftsI typing (method and sequences database).

This work aims to link genotypes to phenotypes in terms of antibiotic susceptibility testing.

Landmark achievements of members of this group include:

2003: Recommendations on methods and media to be used were made available to the scientific community.
2005 - 2007: Implementation of PCR-based approaches as a non-culture method to detect isolates with reduced susceptibility to penicillin on the basis of penA sequencing. A database of penA sequences was established.
2010: Implementation of PCR-based approaches as a non-culture method to detect isolates resistant to rifampicin to penicillin on the basis of rpoB sequencing. A database of rpoB sequences was established.
2013: Implementation of PCR-based approaches as a non-culture method to detect isolates resistant to rifampicin to ciprofloxacin on the basis of gyrA sequencing. A database of gyrA sequences was established.

Linked to publications

Vazquez JA, Arreaza L, Block C, et al. Interlaboratory comparison of agar dilution and Etest methods for determining the MICs of antibiotics used in management of Neisseria meningitidis infections. Antimicrob Agents Chemother 2003, 47:3430-4
Taha MK, Vazquez JA, Hong E, et al. Target gene sequencing to characterize the penicillin G susceptibility of Neisseria meningitidis. Antimicrob Agents Chemother 2007, 51:2784-92
Taha MK, Thulin Hedberg S, Szatanik M, et al. Multicenter study for defining the breakpoint for rifampin resistance in Neisseria meningitidis by rpoB sequencing. Antimicrob Agents Chemother 2010, 54:3651-8
Hong E, Thulin Hedberg S, Abad R, et al. Target gene sequencing to define the susceptibility of Neisseria meningitidis to ciprofloxacin. Antimicrob Agents Chemother 2013, 57:1961-4