Genotypic and phenotypic modifications of Neisseria meningitidis after an accidental human passage.
Bottom Line: By comparing the number of SNP in all three isolates and knowing the number of passages between Z5463 and Z5463PI, we concluded that around 25 bacterial divisions occurred in the human body.Different pilin variants were found after the in vivo passage, which expressed different properties of adhesion.Furthermore the deletion of one gene involved in LOS biosynthesis (lgtB) results in Z5463BC expressing a different LOS than the L9 immunotype of Z2491.
Affiliation: INSERM U1002, Paris, France.
A scientist in our laboratory was accidentally infected while working with Z5463, a Neisseria meningitidis serogroup A strain. She developed severe symptoms (fever, meningism, purpuric lesions) that fortunately evolved with antibiotic treatment to complete recovery. Pulse-field gel electrophoresis confirmed that the isolate obtained from the blood culture (Z5463BC) was identical to Z5463, more precisely to a fourth subculture of this strain used the week before the contamination (Z5463PI). In order to get some insights into genomic modifications that can occur in vivo, we sequenced these three isolates. All the strains contained a mutated mutS allele and therefore displayed an hypermutator phenotype, consistent with the high number of mutations (SNP, Single Nucleotide Polymorphism) detected in the three strains. By comparing the number of SNP in all three isolates and knowing the number of passages between Z5463 and Z5463PI, we concluded that around 25 bacterial divisions occurred in the human body. As expected, the in vivo passage is responsible for several modifications of phase variable genes. This genomic study has been completed by transcriptomic and phenotypic studies, showing that the blood strain used a different haemoglobin-linked iron receptor (HpuA/B) than the parental strains (HmbR). Different pilin variants were found after the in vivo passage, which expressed different properties of adhesion. Furthermore the deletion of one gene involved in LOS biosynthesis (lgtB) results in Z5463BC expressing a different LOS than the L9 immunotype of Z2491. The in vivo passage, despite the small numbers of divisions, permits the selection of numerous genomic modifications that may account for the high capacity of the strain to disseminate.
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Mentions: Excluding the pilin region, the number of genes that could encode a protein with a different amino-acid sequence in Z5463 than in Z2491 is 64. In addition, 3 genes could have their expression modified by a deletion/insertion in a phase variable sequence in their promoter region. Table 2 summarizes the functional groups to which the products of these genes belong. Surprisingly, this analysis revealed the deletion of one bp at position 2478 of the mutS gene of Z5463, introducing a stop codon in the sequence, which would lead to the production of a truncated MutS protein (Fig. 3A). In order to address the possibility that this strain had a mutator phenotype, the ability of Z5463 to generate mutants resistant to rifampicin was assessed. Z5463 has a rate of mutations (10−7/generation) similar to that of a genetically engineered mutS mutant (data not shown). The functional mutS allele of Z2491 was then introduced into Z5463 as described in the experimental procedure section, which reverse this hypermutator phenotype (Fig. 3B), demonstrating that the high mutation rate observed with Z5463 was indeed due to the defective mutS allele.