Evidence of horizontal gene transfer within porB in 19 018 whole-genome Neisseria spp. isolates: a global phylogenetic analysis

Abstract

The PorB porins are the major pore-forming proteins in the genus Neisseria . The trimeric PorB porins consist of 16 highly conserved transmembrane domains that form an amphipathic β-sheet connected by short periplasmic turns and eight extracellular hydrophilic loops. These loops are immunogenic and also play an important role in mediating antimicrobial influx. This study sought to (i) characterize the variations in Neisserial loop 3(355–438 bp) associated with intermediate resistance to penicillin/tetracycline and (ii) evaluate if there was evidence of horizontal gene transfer in any of the loops. We collated an integrated database consisting of 19 018 Neisseria spp. genomes – 17 882 Neisseria gonorrhoeae , 114 Neisseria meningitidis and 1022 commensal Neisseria spp. To identify the porB alleles, a gene-by-gene approach (chewBBACA) was employed. To evaluate the presence of recombination events, the Recombination Detection Programme (RDP4) was used. In total, 3885 porB alleles were detected. Paralogues were identified in 17 Neisseria isolates. Putative recombination was identified in loop regions. Intraspecies recombination among N. gonorrhoeae isolates and interspecies recombination between N. meningitidis and commensal Neisseria spp., and N. gonorrhoeae and N. lactamica were identified. Here, we present a large-scale study of 19 018 Neisseria isolates to describe recombination and variation in the porB gene. Importantly, we found putative recombination in loop regions between the pathogenic and non-pathogenic Neisseria spp. These findings suggest the need for pheno- and genotypic surveillance of antimicrobial susceptibility in commensal Neisseria spp. to prevent the emergence of AMR in the pathogenic Neisseria . This article contains data hosted by Microreact.

Keywords: Neisseria; PorB1a; Porb1b; commensal Neisseria; phylogenetics; porin.

Fig. 1.

(a) Phylogenetic tree of porB alleles (n=3885) (b) Phylogenetic tree of PorB1a allele clade (n=698). The figure was generated using iTOL. Blue, magenta, cyan and yellow colour nodes denotes N. gonorrhoeae, N. meningitidis, N. lactamica and N. subflava, respectively, and the other colours denote other Neisseria spp. Red and green filled circles denote the porB1a and porB1b alleles, respectively. Black stars denote evidence of horizontal gene transfer.

Fig. 2.

(a) Minimum spanning tree comparing core-genome allelic profiles in association with serogroups. (b) Global distribution of serogroups. Percentage of number of isolates is given in brackets. Isolates are displayed as circles. The size of each circle indicates the number of isolates of this particular type. Red and blue nodes denotes the PorB1a and PorB1B serogroup, respectively. Interactive map is available at https://microreact.org/project/3xqrzaAdwarJK8YnGwuzbi-neisserialporins. NTNG – non-typeable N. gonorrhoeae .

Fig. 3.

Minimum spanning tree comparing core-genome allelic profiles in association with substitutions at (a) G120 and (b) A121 amino acid positions. (c) Global distribution of substitutions. Percentage of number of isolates is given in brackets. Isolates are displayed as circles. Inset – percentage of Neisseria spp. with A121G substitution. The size of each circle indicates the number of isolates of a particular type. Interactive map is available at https://microreact.org/project/3xqrzaAdwarJK8YnGwuzbi-neisserialporins

Fig. 4.

MIC distribution of (a, c) penicillin and (b, d) tetracycline for G120/A121 variants of porb1a and porb1b serogroups, respectively. Box plots and statistical analyses for all the isolates with MIC are shown. Statistical significance between variants and MIC distributions was assessed by Mann–Whitney U est: *<I>P<0.01, **<I>P<0.001, ***<I>P<0.0001. Blue colour box plot denotes the wild type. The line inside the box marks the median. The upper and the lower hinges corresponds to the 25th and 75th percentiles.

Fig. 5.

Time of emergence of resistance-associated mutations at positions 120 (columns) and 121 (scatter points) of (a) porB1a and (b) porb1b serogroups. X and Y axis denotes the year and the relative abundance of resistance-associated mutations in the N. gonorrhoeae isolates, respectively.