Adaptive microevolution in CC17 Streptococcus agalactiae from a mother-neonate infection

The hypervirulent CC17 GBS is a major cause of neonatal invasive disease. Although this lineage can acquire mutations during host adaptation, the functional significance of such microvariations for bacterial pathogenesis remains unclear.

The GBS strains were isolated from a mother-neonate pair, with clinical data obtained from electronic medical record. MALDI-TOF MS was used for species identification. Antimicrobial susceptibility testing was performed using K-B method and interpreted by CLSI criteria. Serotyping and MLST were used for molecular epidemiological analysis. Growth curve, hemolysin activity, serum inhibition and hyaluronic acid survival assays were performed to evaluate phenotypic differences between strains. Cell invasion assays and Galleria mellonella larval infection model were used to assess the invasiveness and in vivo virulence of GBS strains. Whole-genome sequencing was applied to compare the genetic backgrounds of the strains. Transcriptome sequencing and rtPCR were conducted to identify and validate DEGs between isolates.

A total of 5 GBS strains were collected and identified, and all were susceptible to penicillin, but resistant to clindamycin and erythromycin. The strains could be classified into ST17 and ST2247. The two types of strains showed no significant differences in growth curve, hemolysis activity, or serum inhibition. In hyaluronic acid solution, the ST2247 strain exhibited higher viability compared to the ST17 (P<0.05). The invasive ability of the ST2247 strain against hUVEC, HTR-8/svneo, or hCMEC/D3 was weaker than that of the ST17 (P<0.05). However, ST2247-infected Galleria mellonella larval had a significantly lower vitality index and higher mortality rate than ST17-infected ones (P<0.05). ST17 and ST2247 exhibited consistent carriage of virulence genes, resistance genes and prophage elements, while only 5 SNPs and 3 INDELs were identified between them, with one single-nucleotide insertion (C>CA) locates 75 bp upstream of the ptsG start codon. Transcriptome sequencing and rt-PCR showed that the expression level of the PTSIIBC gene in ST2247 strain was much higher than that in ST17 strain (P<0.05).

ST17 GBS can experience microevolution within maternal and infant hosts, evolving through minor genomic mutations into new strains. Variations in PTSIIBC gene may associated to the divergence in adaptability and virulence of CC17 GBS strains.