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Neisseria meningitidis is a major cause of life-threatening, rapidly progressing meningitis and septicemia. On the other hand, it is also among the nasopharynx normal flora in up to 15% of healthy adults, who can spread the bacterium to others through saliva or respiratory secretions. Those at greatest risk of disease are infants, adolescents, and young adults, along with travelers to the “meningitis belt” in sub-Saharan Africa. Outbreaks have been associated with hypervirulent strains.
There are currently three vaccines available in the U.S. to prevent meningococcal disease for people aged 2 or older. All three are effective against serogroup A, C, Y, and W (a.k.a. W135) strains. These four, along with serogroup B strains (for which vaccines are currently lacking), were historically responsible for most disease.
N. meningitidis strain identification through serogrouping and genotyping is considered essential to any outbreak investigation, and also plays a central role in monitoring vaccine coverage and efficacy. Indeed, no less than four typing assays are recommended (e.g., by the European Meningococcal Disease Society): serogroup, porA VR, fetA VR, and MLST. Needless to say, meeting this recommendation is technically challenging and costly.
Serogroup is conventionally determined by antisera-based agglutionation assay, although DNA-based PCR assays have been developed that offer potential advantages with respect to cost, sensitivity, and specificity. The remaining three typing assays involve DNA sequencing (neisseria.org/nm/typing). MLST targets 7 housekeeping genes, yielding a sequence type (ST#) and clonal complex (cc#). PorA and FetA are outer membrane proteins with one or two immunodominant variable regions (VR) each. Due to extensive recombination and horizontal gene transfer among N. meningitidis strains, the serogroup, porA, fetA, and MLST markers are often unlinked; e.g., isolates expressing multiple serogroups share fetA VR type F5-1 (see dendrograms below).
MicrobiType has developed Nm-porA and Nm-fetA for affordable, genomics-optimized sequence typing of N. meningitidis. In addition to providing conventional porA and fetA VR type (e.g., P5,2 and F1-30, respectively, for strain FAM18), these services potentially provide greater strain discrimination by analyzing flanking sequences as well. For example, strains FAM18 and NM313 cannot be distinquished in terms of serogroup (C), porA VR (P5,2) or MLST (ST11/cc11), but differ at one nucleotide downstream of VR2 and hence form distinquishable “twigs” on the same branch of the Nm-porA dendrogram. The advantage provided by flanking sequence analysis is particularly apparent when comparing fetA VR type and the Nm-fetA dendrogram: 5 types (F1-5, F3-3, F3-6, F4-1, F5-5) shared by multiple strains are resolved into 13 branches in the dendrogram.
MicrobiType also offers Nm-MLST: through genomics optimization of PCR and sequencing primers, sequence type and clonal complex can be determined for roughly half the cost of conventional MLST. Finally, MicrobiType offers Nm-sero, using validated PCR targets with optimized primers for DNA-based serogrouping (A, B, C, W, X, and Y).