Genomic analysis of endemic clones of toxigenic and non-toxigenic Corynebacterium diphtheriae in Belarus during and after the major epidemic in 1990s

Abstract Background Diphtheria remains a major public health concern with multiple recent outbreaks around the world. Moreover, invasive non-toxigenic strains have emerged globally causing severe infections. A diphtheria epidemic in the former Soviet Union in the 1990s resulted in ~5000 deaths. In this study, we analysed the genome sequences of a collection of 93 C. diphtheriae strains collected during and after this outbreak (1996 – 2014) in a former Soviet State, Belarus to understand the evolutionary dynamics and virulence capacities of these strains. Results C. diphtheriae strains from Belarus belong to ten sequence types (STs). Two major clones, non-toxigenic ST5 and toxigenic ST8, encompassed 76% of the isolates that are associated with sore throat and diphtheria in patients, respectively. Core genomic diversity is limited within outbreak-associated ST8 with relatively higher mutation rates (8.9 × 10−7 substitutions per strain per year) than ST5 (5.6 × 10−7 substitutions per strain per year) where most of the diversity was introduced by recombination. A variation in the virulence gene repertoire including the presence of tox gene is likely responsible for pathogenic differences between different strains. However, strains with similar virulence potential can cause disease in some individuals and remain asymptomatic in others. Eight synonymous single nucleotide polymorphisms were observed between the tox genes of the vaccine strain PW8 and other toxigenic strains of ST8, ST25, ST28, ST41 and non-toxigenic tox gene-bearing (NTTB) ST40 strains. A single nucleotide deletion at position 52 in the tox gene resulted in the frameshift in ST40 isolates, converting them into NTTB strains. Conclusions Non-toxigenic C. diphtheriae ST5 and toxigenic ST8 strains have been endemic in Belarus both during and after the epidemic in 1990s. A high vaccine coverage has effectively controlled diphtheria in Belarus; however, non-toxigenic strains continue to circulate in the population. Recombination is an important evolutionary force in shaping the genomic diversity in C. diphtheriae. However, the relative role of recombination and mutations in diversification varies between different clones

Additional file 1: Table S1. of Genomic analysis of endemic clones of toxigenic and non-toxigenic Corynebacterium diphtheriae in Belarus during and after the major epidemic in 1990s

Details of C. diphtheriae strains analysed in this study. (XLSX 21 kb

Additional file 3: Figure S1. of Genomic analysis of endemic clones of toxigenic and non-toxigenic Corynebacterium diphtheriae in Belarus during and after the major epidemic in 1990s

A ML tree from core genomic alignment of ST5 strains; Figure S2. Gubbins analysis of recombination in the core genome of C. diphtheriae. Predicted recombination events on internal branches are shown in red and those occurred at terminal branches are shown in blue; Figure S3. A plot of root-to-tip divergence (Y-axis) against the sampling dates (X-axis); A. within ST5 and B. within ST8; Figure S4. A. CDS BLAST map of ST5 using strain ISS 4060 as the reference. B. CDS BLAST map of ST8 using strain NCTC 13129 as the reference; Figure S5. A ML tree from the binary data of the presence or absence of genes in the accessory genome. The scale bar with a distance of 0.1 represents the difference of 341.7 genes; Figure S6. A plot showing the average global vaccine coverage, reported vaccination in Belarus and the reported number of diphtheria cases between 1992 and 2015. (PDF 5948 kb

Additional file 2: Table S2. of Genomic analysis of endemic clones of toxigenic and non-toxigenic Corynebacterium diphtheriae in Belarus during and after the major epidemic in 1990s

Number and toxigenicity of isolates collected between 1996 and 2014 in Belarus and those selected for genomic analyses in this study; Table S3. Distribution of isolates from epidemic (≤ year 2000) and postepidemic period (≥ year 2001) in major groups. All minor groups are pooled together for statistical analysis; Table S4. Distribution of isolates from asymptomatic carriage, diphtheria and sore throat patients in major groups. All minor groups are pooled together for statistical analysis; Table S5. Allelic variation in the direct repeat and spacer sequences among the CRISPR loci of ST8 and ST5 isolates. (PDF 396 kb