Virulence-Associated Characteristics of Serotype 14 and Serogroup 9 Streptococcus pneumoniae Clones Circulating in Brazil: Association of Penicillin Non-susceptibility With Transparent Colony Phenotype Variants

Abstract

Streptococcus pneumoniae remains a major agent of invasive diseases, especially in children and the elderly. The presence of pneumococcal capsule, pneumococcal surface protein A (PspA), and pilus type 1 (PI-1) and the ability of colony phase variation are assumed to play important roles in the virulence potential of this microorganism. Differences in the capsular polysaccharide allow the characterization of more than 90 pneumococcal serotypes; among them, serotype 14 and serogroup 9 stand out due to their prevalence in the pre-pneumococcal conjugate vaccine era and frequent association with penicillin non-susceptibility. Here we investigated the distribution of PI-1 and pspA genes and colony phase variants among 315 S. pneumoniae isolates belonging to serotype 14 and serogroup 9, recovered over 20 years in Brazil, and correlated these characteristics with penicillin susceptibility and genotype as determined by multilocus sequence typing. All strains were shown to carry pspA genes, with those of family 2 (pspA2) being the most common, and nearly half of the strains harbored P1-1 genes. The pspA gene family and the presence of PI-1 genes were conserved features among strains belonging to a given clone. A trend for increasing the occurrence of pspA2 and PI-1 genes over the period of investigation was observed, and it coincided with the dissemination of CC156 (Spain 9V-3) clone in Brazil, suggesting a role for these virulence attributes in the establishment and the persistence of this successful clone. Opaque variant was the colony phenotype most frequently observed, regardless of clonal type. On the other hand, the transparent variant was more commonly associated with penicillin-non-susceptible pneumococci and with strains presenting evidence of recombination events involving the genes coding for polysaccharide capsule and PspA,

Clinical features relating to pneumococcal colony phase variation in hospitalized adults with pneumonia

“…The minimally (2–3 times) passaged pneumococci from frozen stocks were directly grown in Todd-Hewitt broth with 0.5 % yeast extract and plated on Tryptic Soy Agar (Nippon Beckton-Dickinson, Tokyo, Japan) onto which 100 µl of catalase (3000 U) (MP Biomedicals) was added per plate. After incubation for 16 h at 37 °C with 5 % CO 2 [12–14], colony morphology was evaluated by a stereomicroscope (EVOS FL System), as described previously [7].…”

“…Japan) onto which 100 µl of catalase (3000 U) (MP Biomedicals) was added per plate. After incubation for 16 h at 37 °C with 5 % CO 2 [12][13][14], colony morphology was evaluated by a stereomicroscope (EVOS FL System), as described previously [7].…”