Capsular Switching and ICE Transformation Occurred in Human Streptococcus agalactiae ST19 With High Pathogenicity to Fish

Abstract

Although Streptococcus agalactiae (GBS) cross-infection between human and fish has been confirmed in experimental and clinical studies, the mechanisms underlying GBS cross-species infection remain largely unclear. We have found different human GBS ST19 strains exhibiting strong or weak pathogenic to fish (sGBS and wGBS). In this study, our objective was to identify the genetic elements responsible for GBS cross species infection based on genome sequence data and comparative genomics. The genomes of 11 sGBS strains and 11 wGBS strains were sequenced, and the genomic analysis was performed base on pan-genome, CRISPRs, phylogenetic reconstruction and genome comparison. The results from the pan-genome, CRISPRs analysis and phylogenetic reconstruction indicated that genomes between sGBS were more conservative than that of wGBS. The genomic differences between sGBS and wGBS were primarily in the Cps region (about 111 kb) and its adjacent ICE region (about 106 kb). The Cps region included the entire cps operon, and all sGBS were capsular polysaccharide (CPS) type V, while all wGBS were CPS type III. The ICE region of sGBS contained integrative and conjugative elements (ICE) with IQ element and erm(TR), and was very conserved, whereas the ICE region of wGBS contained ICE with mega elements and the variation was large. The capsular switching (III-V) and transformation of ICE adjacent to the Cps region occurred in human GBS ST19 with different pathogenicity to fish, which may be related to the capability of GBS cross-infection.

Keywords: GBS; ICE; ST19; capsular switching; cross-species infection.

Figure 1

The pan-genome and comparison of sGBS and wGBS genomes. (A) Anvi'o pangenome visualization of sGBS and wGBS genomes. The red part in outer ring showed single-copy core genes. The genomes of sGBS and wGBS were shown in purple and black, respectively, and the 2603V/R genome was shown in blue, the green histogram meaned genome length. (B) Comparison of genomes of sGBS and wGBS. The LZF006 (wGBS) genome and the LZF001 (sGBS) genome were served as the reference, respectively, and other genomes were aligned to the reference genome; each circle represented a different genome, the strain name and CPS type were labeled on each circle. The circle of reference genome was not shown; the color bar on the bottom indicated the percentage of protein sequence identity against the reference genome. The relevant areas and locus were indicated by black arrows in the figure.

Figure 2

Comparative analysis of systematic evolution and CRISPRs structure of ST19 GBS strains. (A) A Maximum Likelihood phylogenetic tree based on 1,707 single copy orthology clusters of the 23 strains, and 2603V/R strain was used as outgroup strain. The bootstrap confidence value which was 100 was not shown in the figure. (B) CRISPRs structure comparison. Direct repeat sequence was not included; only RT, ST, and spacers were represented. The spacers were numbered, and the same number highlighted with same color indicated that the spacer sequence was the same.

Figure 3

The evolutionary analysis of homologous proteins in cps operon and their structure. (A) A Maximum Likelihood phylogenetic tree based on 12 single copy orthology clusters of cps operon from 20 strains. The bootstrap confidence value which was 100 was not shown in the figure. (B) The cps operon structure of GBS serotypes Ia, Ib, III, and V. The cpsY, cpsA-cpsL, NeuA-NeuD genes were labeled with arrows, and the arrow showed the direction of transcription. The different color of arrow indicated the percent identity of the encoded protein.

Figure 4

Gene organization of putative integrative conjugative element ICESagLZF001IQ, ICESagLZF001erm, and ICESagLZF006mega. (A) Schematic representation of ICESagLZF001IQ with ICESpn529IQ as the genetic reference. (B) Schematic representation of ICESagLZF001erm with ICESp2905 as the genetic reference. (C) Schematic representation of ICESagLZF006mega with ICESpy009 as the genetic reference. The arrows indicated the direction of the transcription. The color of the arrow was correlated with the percent identity. The arrows with red border were annotated the gene information. The deletion regions were shown with red rectangle. The number in the arrow indicated the serial number of ORFs, and the number in the red square indicated the number of ORFs within the fragment.