CAMP-negative Streptococcus agalactiae strains exhibited complete or partial chromosomal deletions of the CAMP-factor encoding gene cfb.

Universal antepartum group B streptococcus (GBS) screening and intrapartum antibiotic prophylaxis (IAP) have effectively reduced early-onset GBS infections. However, GBS strains with chromosomal deletions affecting the cfb gene may produce false negatives in both the CAMP test and cfb-based molecular diagnostics, potentially increasing the risk of neonatal infections. Vaginal swabs were collected from pregnant women at 35-37 weeks of gestation in our hospital and cultured on agar. Suspected GBS strains were initially identified using the CAMP test and then confirmed with the VITEK-2 system. CAMP-negative GBS strains underwent additional testing by qPCR, 16S rDNA, serotyping, and multilocus sequence typing (MLST). PCR for the cfb gene and whole-genome sequencing were performed on CAMP-negative strains. From 5,794 samples, 526 (9.1%) GBS strains, including 19 (3.6%) CAMP-negative strains and 2 strains from the same patient, were isolated. All 19 CAMP-negative strains were serotypes III and ST862. Among these strains, only one strain was cfb positive by qPCR, whereas all tested positive with a multitarget qPCR kit for cfb and cps. PCR amplification upstream of the cfb gene produced a specific band in strain PP669713 only, suggesting N-terminal cfb gene retention in PP669713 and complete cfb loss in the other strains. Whole-genome sequencing confirmed a chromosomal deletion in PP669713. Antibiotic susceptibility testing revealed no resistance to penicillin. However, CAMP-positive strains presented a greater prevalence of resistance to ciprofloxacin, and levofloxacin than CAMP-negative strains did. Our study highlights the potential risk of missed GBS detection using CAMP tests and cfb-targeted molecular assays.

Importance: Our work makes several novel contributions to the field. (i) We report the first documented case of a C-terminal deletion of the cfb gene in a CAMP-negative GBS strain, demonstrating that both N-terminal and C-terminal regions are essential for cohemolytic activity. (ii) Our findings reveal that CAMP-negative GBS strains (3.6% of isolates) are more prevalent than previously recognized, with most cases resulting from complete chromosomal deletions of the cfb gene. (iii) We provide evidence that single-target molecular assays targeting only the cfb gene may miss GBS detection, highlighting the necessity for multi-target approaches in clinical diagnostics. (iv) We demonstrate a unique antibiotic resistance pattern in CAMP-negative strains, showing significantly lower resistance to certain antibiotics compared to CAMP-positive strains.