Draft genome sequence of Bordetella pertussis PS21, a circulating clinical isolate from India

TL;DR

This paper presents the draft genome sequence of a Bordetella pertussis strain from India, highlighting mutations that may affect vaccine recognition.

Contribution

The study provides a new genome sequence of a clinical B. pertussis isolate from India with specific mutations that could impact virulence and vaccine efficacy.

Findings

The genome contains classical virulence genes and notable polymorphisms in key toxin and regulatory genes.

Mutations in ptxA, bvgS, bapC, and ptxB may influence toxin function and immune recognition.

These findings could inform understanding of B. pertussis evolution and vaccine effectiveness in India.

Abstract

We report the draft genome sequence of Bordetella pertussis PS21, a clinical isolate obtained from a culture-positive sample from India. The 3.6 Mb genome harbors classical virulence genes and several notable polymorphisms, including a dual substitution in ptxA (Arg5Trp, Met228Ile), a Lys705Glu mutation in bvgS, a Lys97 frameshift in bapC, and a Gly45Ser substitution in ptxB. These mutations may influence pertussis toxin function and virulence regulation, potentially impacting immune recognition by pertussis vaccines.

Full text

ANNOUNCEMENT

Genomic characterization of Bordetella pertussis PS21, a clinical isolate from culture-positive sample in India, offers key insights into pertussis pathogenesis and adaptation in low- and middle-income countries. India accounts for ~26% of global pertussis cases (1) yet a culture-positive isolates are scarce, emphasizing the importance of this genomic data set for informing control strategies in a region with high primary vaccination coverage (2).

The PS21 isolate was obtained via nasopharyngeal swab from a 4-week-old child at Christian Medical College, Vellore, India. The child was admitted from 2 to 13 June 2023, with myocarditis and suspected pertussis, treated with azithromycin and later vaccinated with PENTAVAC (DTwP + Hib+ HBV) followed by two doses of QUADROVAX (DPT + Hib). Since the diagnosis preceded vaccination, PS21 likely represents a circulating B. pertussis strain. This highlights the risk to unvaccinated neonates and the continued circulation of B. pertussis despite vaccination efforts (3).

B. pertussis PS21 was isolated on Bordet–Gengou agar with 15% defibrinated horse blood and incubated at 35°C for 72 h. Identification was based on morphology and biochemical profile (gram-negative coccobacilli; greyish-white, convex, shiny colonies with “bisected pearl” or “mercury drop” appearance; no growth on MacConkey agar; opaque on charcoal blood agar; catalase/oxidase positive; motility, urease, nitrate reduction, and citrate utilization negative) and confirmed by MALDI-TOF MS (VITEK MS, bioMérieux). Ten colonies were pooled for DNA extraction (Qiagen DNeasy Blood & Tissue Kit).

Libraries (Illumina DNA Prep) were sequenced on the Illumina HiSeq platform (2×151 bp). A total of 740,470 read pairs were obtained. Quality was assessed with FastQC v0.11.9; assembly (SKESA v2.4.0) was evaluated with QUAST (4–6). The 3.6 Mb draft genome comprised 372 contigs (largest 67,885 bp; GC 68%; N50 16,241 bp; N90 5,521 bp) and was annotated using the NCBI Prokaryotic Genome Annotation Pipeline with default parameters (7).

The annotated genome comprises 3,412 protein-coding sequences, 46 tRNA genes, and three rRNA operons. Key virulence genes include the ptx operon (ptxA-E) (8), cyaA-E (adenylate cyclase toxin) (9), dnt (dermonecrotic toxin) (10), type IV secretion system (ptlA-I) (11), type III secretion system (bsc cluster and associated effectors and regulators) (12), fimbrial biogenesis cluster (fimA-D, fim2, fim3, and fimX) (13), fhaC (filamentous hemagglutinin transporter), and complement resistance genes (bapC and brkA/B) (14).

Comparative genomic analysis of B. pertussis strain PS21 was carried out using Snippy v4.6.0, with Tohama I (NC_002929) as the reference genome (15). The analysis included comparisons with six vaccine strains (J445, J446, J447, J448, Bp165, and Pelita III) (16, 17) and six clinical isolates (S1–S5, BPD1, and BPD2) (18). PS21 carried a unique dual substitution in ptxA (Arg5Trp and Met228Ile), absent in the compared strains. This mutation may change the structure or stability of the pertussis toxin, possibly affecting its ADP-ribosyltransferase activity (8). Other mutations included Lys705Glu in bvgS (global virulence regulator), a Lys97 frameshift in bapC (biofilm-associated protein), and Gly45Ser in ptxB (pertussis toxin subunit), which were also present in vaccine strains and therefore not unique to PS21 (19).

This genome characterization of PS21 offers insights into B. pertussis evolution and supports improved diagnostics and vaccines in resource-limited settings.