Open Access Te Herenga Waka-Victoria University of Wellington
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Genomic Organisation of Meningococcal pilS in Carriage and Disease

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posted on 2024-05-14, 22:53 authored by Paloma Gea

Neisseria meningitidis (meningococcus) is an obligate human pathogen that can cause invasive meningococcal disease. Meningococci asymptomatically colonise the nasopharyngeal mucosa as commensals in up to 30% of the population. Disease occurs when the bacteria adhere to epithelial cells and invade deeper tissues, rapidly replicating in the bloodstream or crossing the blood-brain barrier. This is a severe disease with serious long term sequelae. Vaccines have been developed against the capsule surrounding the bacteria and other recombinant proteins, but despite this, rates of invasive meningococcal disease are still high. The vaccines are not as effective against carriage and the pathogenesis of the bacterium is still poorly understood. The type IV meningococcal pilus is a known virulence factor that facilitates adhesion to different cell types. The pilus is made up of repeating subunits of the PilE protein, encoded by the pilE gene. This gene undergoes extensive recombination with a genomic locus called pilS. However, due to the long and repetitive nature of the pilS region, previous studies using short-read sequencing have been unable to accurately resolve this region. The overarching goal of this project was to use long read sequencing to resolve the pilS region and compare the genomic structure of this region between closely related carriage and disease isolates. These isolates were taken during the New Zealand meningococcal disease epidemic from 1991-2008. Isolates were taken from the index disease patient and their healthy household contacts. Healthy household contacts were often found to be asymptomatically carrying N. meningitidis of the same sequence type as the index disease patient. These carriage and disease pairs are indistinguishable by common laboratory typing, however a previous study showed the carriage isolates from one household had a truncated pilS compared to the disease isolate [1]. The aim was to look for consistent differences in the pilS region between closely related carriage and disease isolates. Long read Oxford Nanopore sequencing was used to generate assemblies for each of the isolates that mapped across repetitive regions. A method to annotate the highly repetitive pilS region was developed, including the promoter sequence and cis regulatory elements. Bioinformatic pipelines were developed to compare the pilS region between carriage and disease isolates. The variation in the expressed pilE gene was also investigated, showing conserved and highly variable regions, which corroborates what has been described in the literature. In this work, the genomes of 23 N. meningitidis isolates have been sequenced and assembled. The pilS region of all the isolates has been annotated. No consistent differences in pilS were found between carriage and disease isolates. However, by using Oxford Nanopore sequencing, novel deletions have been detected in the pilS region that have not been described in the literature. These deletions have been validated bioinformatically and experimentally by isolating and sequencing single colonies of N. meningitidis. A bioinformatic pipeline has been developed to use Oxford Nanopore sequencing to identify regions of high variation, which can be applied to other recombinant genomic loci. Understanding the genomic variation of the pilS region is important to understanding how the pilus contributes to invasive meningococcal disease.


Copyright Date


Date of Award



Te Herenga Waka—Victoria University of Wellington

Rights License

Author Retains Copyright

Degree Discipline

Molecular Microbiology

Degree Grantor

Te Herenga Waka—Victoria University of Wellington

Degree Level


Degree Name

Master of Science

ANZSRC Type Of Activity code

1 Pure basic research

Victoria University of Wellington Item Type

Awarded Research Masters Thesis



Victoria University of Wellington School

School of Biological Sciences


MacKichan, Joanna; Ren, Xiaoyun