WGS analysis of a penicillin-resistant Neisseria meningitidis strain containing a chromosomal ROB-1 β-lactamase gene

Abstract

Objectives: Neisseria meningitidis is rarely penicillin resistant. We describe WGS analysis of a penicillin-resistant N. meningitidis collected from a case of invasive meningococcal disease.

Methods: Serogrouping, serotyping and serosubtyping were performed with specific antibodies. β-Lactamase was detected by nitrocefin. MICs were determined by Etest and agar dilution. Sequencing of N. meningitidis genomes was done on the Illumina MiSeq platform and genome data were analysed using the Bacterial Isolate Genome Sequence Database (BIGSdb) on the PubMLST Neisseria website (https://pubmlst.org/neisseria/). Transformation was used to confirm the genetic basis of the penicillin resistance.

Results: An N. meningitidis blood isolate from a female patient in her mid-50s with a painful and septic left shoulder was found to have penicillin MIC values of 3-12 mg/L. The isolate was typed as Y: 14, 19: P1.- and ST3587, and was weakly β-lactamase positive. WGS analysis identified a full-length copy of the β-lactamase gene blaROB-1, which was contained on a 1719 bp insert with a G + C content of 41.7% (versus a G + C content of N. meningitidis of 51.7%), suggesting that the blaROB-1 gene came from a different bacterial species. A GenBank analysis of the blaROB-1 gene insert found 99.77% identity with a DNA segment found in plasmid pB1000' from Haemophilus influenzae. Transformation of a penicillin-susceptible strain with the blaROB-1 gene conferred β-lactamase activity and penicillin resistance.

Conclusions: N. meningitidis serogroup Y, ST3587 can carry and express the blaROB-1 gene, leading to penicillin resistance. It is highly likely that the N. meningitidis isolate acquired the blaROB-1 gene from H. influenzae.

Figure 1.

A pan-genome map of three ST3587 meningococcal strains (NMLY144 from Alberta, Canada; M17 240005 from the UK; and 2816 from Italy), a serogroup Y strain (NMLY048 from Alberta, Canada) and a serogroup W strain (M7124 from Saudi Arabia) generated using the GView server. A full-genome view (a) and a zoomed view (b) show an area centring around the β-lactamase (bla_ROB-1) gene, labelled as penP. The innermost slot (black) shows the constructed pan-genome using all uploaded genomes. A grey line separates the pan-genome from the other genomes, aligned to show sequence homologies and the absence of genes. Gaps denote non-coding sequences. Depicted in the box is the translated protein sequence of the ROB-1 β-lactamase annotated by PROKKA as penP. This figure appears in colour in the online version of JAC and in black and white in the print version of JAC.

Figure 2.

Neighbour-net phylogenetic network comparison of four N. meningitidis ST23 CC isolates with the meningococcal serogroup W reference strain M7124 as an outlier by means of the BIGSdb genome comparator tool using the N. meningitidis cgMLST version 1.0 scheme. The scale bar denotes the number of allelic differences. This figure appears in colour in the online version of JAC and in black and white in the print version of JAC.