Contribution of phase variation of Opa proteins to persistent carriage and immune evasion of Neisseria meningitides
thesisposted on 20.11.2018, 11:30 by Ali Abdulwahid Abed Al-Rubaiawi
Neisseria meningitidis is one of the main causes of bacterial meningitis and septicaemia but colonizes the upper respiratory tract of humans asymptomatically as a normal commensal. Phase variation (PV) in the surface antigens is proposed as an effective mechanism to enable these bacteria to adapt and persist in the human host. Opa proteins are expressed on the outer surface of meningococcal cells playing an important role in the pathogenicity by mediating the adhesion to and invasion of human cells. These proteins are encoded by three/four loci and each locus is phase variable due to pentameric repeats within the coding region. The phase variability in Opa proteins was investigated in meningococcal isolates from 19 carriers and time points representing up to six months of asymptomatic carriage. Changes in repeat tracts were analyzed by GeneScan, and a high frequency of PV was observed in at least two loci with a rate of 0.06 mutations/gene/month during colonization. The expression state of Opa was confirmed by Western blotting indicating expression of a limited number of Opa variants. Around 70% of the isolates expressed only one Opa and none simultaneously expressed four Opa. Intergenic and intragenic recombination was detected in two carriers, leading to new opa alleles with functions differing from the previous alleles. These results revealed that persistent carriage was correlated with a high rate of variation and switching between different Opa variants with stable expression of one or more alleles that may maintain Opa-mediated adhesion. The study also highlights the role of Opa proteins in mediating in vitro escape of N. meningitidis strain MC58 from anti-Opa bactericidal antibodies while selection for populations expressing other Opa variants was also observed in vivo indicating the importance of switching between the different variants for immune evasion and maintaining the function of this protein. Four recombinant Opa proteins were generated in this study and used for developing bactericidal polyclonal antibodies that can be used for further investigations of in vitro and in vivo immune escape.