Sir, Meningococcal meningitis is caused by Neisseria meningitidis. At present, there is an effective vaccine for some of the serogroups of this organism but not for the B strains, which are the most prevalent in the UK, Europe and the USA. The serogroup B polysaccharide is poorly immunogenic, so development of new candidate vaccines has focused on the outer membrane proteins, particularly the porin protein PorA. To develop a non-culture-based subtyping system for the meningococcus, we have used the polymerase chain reaction (PCR) in combination with single-stranded conformational polymorphism analysis (SSCP) to examine the meningococcal porA gene (Newcombe et al., 1997, J Clin Microbiol35: 1809–1812). The VR1 region of porA was amplified using PCR primers that generated a PCR product of ≈ 300 bp. However, out of the 25 clinical specimens examined, one blood specimen, JP280, generated a 1.1 kb PCR product (Fig. 1). The isolate was obtained from a 12-year-old boy from the Gloucester area in the UK who presented with meningitis, was treated and went on to make an uneventful recovery. The same 1.1 kb PCR product was generated using DNA isolated from a meningococcal strain cultured from the patient's blood (Fig. 1) and was sequenced. The DNA sequence (EMBL AJ007406, AJ007407) matched the published sequences of the meningococcal porA gene (McGuinness et al., 1990, J Exp Med171: 1871–1882) but was interrupted by a single copy of the insertion sequence IS1301 (Hammerschmidt et al., 1996, EMBO J15: 192–198). The IS element had inserted within the VR1 region of porA in an identical position and orientation to that described for a type C strain recently isolated in Quebec, Canada (Arhin et al., 1998, Can J Microbiol44: 56–63). Strain JP280 was typed by the Meningococcal Reference Laboratory as a C,2a:NT (non-subtypable), it did not react with any of the PorA antibodies, indicating that insertion of IS1301 sequence led to insertional inactivation of PorA expression. We have also examined DNA from 100 microbial isolates of N. meningitidis using PCR-SSCP and found two that were identical to JP280 with regard to both IS1301 insertion into the porA gene and the C,2a:NT typing. The isolates were both from a recent outbreak of meningococcal disease (CDR Weekly, Vol. 7 no. 44, 31 October 1997). The first (M97.252455) was isolated from the cerebrospinal fluid of a patient who had died of meningococcal septicaemia and the second (M97.252456) was isolated from a throat swab obtained from a contact of the first patient. . PCR amplification of the VR1 region of meningococcus from DNA isolated from clinical specimens and microbial cultures. Lane 1, molecular size markers; lane 2, blood specimen from JP280; lane 3, blood specimen from a patient infected with a P1.16 meningococcal strain; lane 4, PCR negative control; lane 5, P1.9 meningococcal strain; lane 6, JP280 meningococcal culture. PCR was performed as described (Newcombe et al. 1997, J Clin Microbiol35: 1809–1812) and the products electrophoresed through 1.5% agarose stained with ethidium bromide and photographed under ultraviolet light. IS1301 is known to be involved in reversible capsulation in Neisseria, by insertion into and excision from the SiaA gene locus (Hammerschmidt et al., 1996, EMBO J15: 192–198). It is possible that similar excision events may restore expression of PorA in strains like JP280. However, the IS1301 insertion event was also detected in vivo within the blood of one patient (JP280), indicating that the PorA mutant strain was fully virulent and capable of causing both infections and meningitis. PorA is a major antigen of the meningococcus and a target for bactericidal antibodies. For these reasons, a number of vaccines and candidate vaccines have been developed that use PorA as a major immunogen (Ala'Aldeen and Cartwright, 1996, J Infect33: 153–157). However, meningococcal strains like JP280 with an inactivated porA gene would not be subject to any protection afforded by a PorA-based vaccine. In addition, widespread immunization with a PorA-based vaccine would risk selecting for strains, which, like JP280, have lost PorA expression but may remain fully virulent. This work was supported by the National Meningitis Trust. The use of the automated DNA sequencer was supported by the Wellcome Trust. We would like to thank the Meningococcal Reference Laboratory (Manchester Public Health Laboratory) for supplying clinical strains.
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Newcombe et al. (1998) studied this question.