Abstract

Two haemoglobin-binding proteins, HmbR and HpuAB, contribute to iron acquisition by Neisseria meningitidis. These receptors are subject to high frequency, reversible switches in gene expression - phase variation (PV) - due to mutations in homopolymeric (poly-G) repeats present in the open reading frame. The distribution and PV state of these receptors was assessed for a representative collection of isolates from invasive meningococcal disease patients of England, Wales and Northern Ireland. Most of the major clonal complexes had only the HmbR receptor whilst the recently expanding ST-275-centred cluster of the ST-269 clonal complex had both receptors. At least one of the receptors was in an ‘ON’ configuration in 76.3% of the isolates, a finding that was largely consistent with phenotypic analyses. As PV status may change during isolation and culture of meningococci, a PCR-based protocol was utilised to confirm the expression status of the receptors within contemporaneously acquired clinical specimens (blood/cerebrospinal fluid) from the respective patients. The expression state was confirmed for all isolate/specimen pairs with <15 tract repeats indicating that the PV status of these receptors is stable during isolation. This study therefore establishes a protocol for determining in vivo PV status to aid in determining the contributions of phase variable genes to invasive meningococcal disease. Furthermore, the results of the study support a putative but non-essential role of the meningococcal haemoglobin receptors as virulence factors whilst further highlighting their vaccine candidacy.

Highlights

  • Neisseria meningitidis is a commensal of the human nasopharynx with an overall prevalence of approximately 10% [1]

  • The ST-269 clonal complex was split between isolates comprising an hmbR-only (45.5%; all centered around sequence type (ST)-269 by eBURST analysis [19], table S1) or an hmbR:hpuA genotype (50%; of which 90.9% were centered around ST-275 [19])

  • Sequence analysis of homopolymeric tract regions of hmbR and hpuA among invasive meningococcal disease (IMD) isolates Homopolymeric tracts are susceptible to strand slippage during PCR amplification and this susceptibility increases with tract length and the number of PCR cycles [22]

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Summary

Introduction

Neisseria meningitidis (the meningococcus) is a commensal of the human nasopharynx with an overall prevalence of approximately 10% [1]. It is a leading cause of meningitis and septicaemia globally and is associated with significant morbidity and mortality [2]. There are five main capsular serogroups associated with invasive meningococcal disease (IMD; serogroups A, B, C, W and Y) of which serogroups A, C, W and Y are preventable by means of efficacious glycoconjugate vaccines [3]. The polysaccharide of serogroup B meningococci (MenB), a leading cause of IMD in industrialized nations, is poorly immunogenic in humans [4].

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