Comparative genomics of Mycoplasma pneumoniae isolated from children with pneumonia: South Korea, 2010-2016

Abstract

Background: Mycoplasma pneumoniae is a common cause of respiratory tract infections in children and adults. This study applied high-throughput whole genome sequencing (WGS) technologies to analyze the genomes of 30 M. pneumoniae strains isolated from children with pneumonia in South Korea during the two epidemics from 2010 to 2016 in comparison with a global collection of 48 M. pneumoniae strains which includes seven countries ranging from 1944 to 2017.

Results: The 30 Korean strains had approximately 40% GC content and ranged from 815,686 to 818,669 base pairs, coding for a total of 809 to 828 genes. Overall, BRIG revealed 99% to > 99% similarity among strains. The genomic similarity dropped to approximately 95% in the P1 type 2 strains when aligned to the reference M129 genome, which corresponded to the region of the p1 gene. MAUVE detected four subtype-specific insertions (three in P1 type 1 and one in P1 type 2), of which were all hypothetical proteins except one tRNA insertion in all P1 type 1 strains. The phylogenetic associations of 30 strains were generally consistent with the multilocus sequence typing results. The phylogenetic tree constructed with 78 genomes including 30 genomes from Korea formed two clusters and further divided into two sub-clusters. eBURST analysis revealed two clonal complexes according to P1 typing results showing higher diversity among P1 type 2 strains.

Conclusions: The comparative whole genome approach was able to define high genetic identity, unique structural diversity, and phylogenetic associations among the 78 M. pneumoniae strains isolated worldwide.

Keywords: Comparative genomics; Mycoplasma pneumoniae; Whole genome analysis.

Fig. 1

Overall sequence identity of the 30 sequenced strains with the reference M129 genome. Solid coloration indicates > 99% identity and transparent grey indicates approximately 95% identity. Location in the reference genome is indicated by numeration on the inside of the ring. GC content in the reference genome is indicated by the black bar graphs between the genomic coordinates and the colored rings (bars pointing toward the outside of the circle indicate high GC content)

Fig. 2

Whole genome alignment of the 30 sequenced strains with 6 reference sequences using MAUVE. Regions colored in MAUVE are conserved across all strains. a Two 1 Kbp (approximate) insertions are noticed in the P1 type 1 groups at 169–170 Kb and 178–179 Kb. b A 2 Kbp (approximate) insertion is noticed in the P1 type 1 groups at 558–560 Kb. c A 6 Kbp (approximate) insertion is noticed in the P1 type 2 groups at 708 Kb. All positions are based on M129 reference strain

Fig. 3

Heatmap of protein families of 30 sequenced genomes with reference genome M. pneumoniae M129. Cell color represents the number of proteins from a specific genome in a given protein family. Note that P1 types 2 (10–1385, 11–174, 11–949, 11–1384, 14–637 and 16–734) are distinguishable from P1 types 1

Fig. 4

Phylogenetic tree based on whole genome alignment of the 30 sequenced strains with 48 M. pneumoniae genomes accessed from NCBI. The tree was built through 500 bootstraps using the maximum composite likelihood approach based on neighbor-joining algorithms. Branch length designates actual distance. Bootstrapping values over 50 are represented on the tree. Blue colored strains are from this study and red colored strains are the 6 references. Strains are grouped into four distinct clades. ST, sequence type

Fig. 5

Mycoplasma pneumoniae sequence type (ST) relationship by eBURST analysis including 30 strains from this study, 48 strains from NCBI, and previously reported STs from PubMLST (http://pubmlst.org/mpneumoniae/). Two main CCs were defined with two singletons (ST12 and ST22). ST3 and ST2 were the predicted founder of each CC. The size of each circle correlates with the number of isolates of each ST. STs in gray are previously reported, but not included in the investigation of this study. CC, clonal complex