PspA family typing and PCR-based DNA fingerprinting with BOX A1R primer of pneumococci from the blood of patients in the USA with and without sickle cell disease

Abstract

Disease and mortality rates for Streptococcus pneumoniae infections are much higher in patients with sickle cell disease (SCD) than in age-matched patients without SCD. Pneumococcal surface protein A (PspA) has been proposed as a component in human vaccines against S. pneumoniae to provide greater breadth of coverage than can be obtained with the 7-valent conjugate vaccine. The cross-reactivity of PspA is associated with the 'PspA family' structure. In this study we examined strains of S. pneumoniae from patients with and without SCD to determine whether the strains infecting the hypersusceptible population of SCD patients were limited to the same two PspA families already known to comprise over 95% of strains infecting non-SCD patients. Each strain was also evaluated according to the presence or absence of specific PCR fragments based on repetitive BOX elements to screen for possible SCD-associated clonal structure. Strains from SCD and non-SCD patients were similarly dispersed among the most common BOX PCR groups and strains from both groups expressed a similar distribution of PspA variants. Thus, a PspA vaccine designed for the population at large should also be appropriate for patients with SCD.

Genetic Diversity of PspA Types among Nasopharyngeal Isolates Collected during an Ongoing Surveillance Study of Children in Brazil

“…These findings are in accordance with those of previous reports on invasive pneumococcal strains, in which the prevalence of PspA Fam1 ranged from 44.8 to 51.9% and that of Fam2 ranged from 43.2 to 55.2% (3,8,26). However, the predominance of PspA Fam1 (15,17,27) and the high prevalence of PspA Fam2 (85.7%) have been detected in some studies (2), mainly with adult populations.…”

“…Our results showed no statistically significant difference between the prevalence of NP PspA families and clades in healthy and ill children, as recently reported in studies that were conducted with invasive pneumococcal isolates and that compared patients with and without sickle cell disease (26). Therefore, we could hypothesize a similar prevalence of PspA types in NP pneumococcal colonizers and invasive strains, reinforcing the usefulness of PspA as a promising candidate for the pneumococcal vaccine.…”

“…DNA standards from strains Rx1 (clade 2) and BG11703 (clade 4) were used as controls for Fam1 and Fam2, respectively, in each set of PCRs. The PCR was performed with oligonucleotide primers synthesized on the basis of the published DNA sequences for PspA Fam1 (primers LSM12 and SKH63); PspA Fam2 (primers LSM12 and SKH52); and PspA Fam1, Fam2, and Fam3 (primers LSM12 and SKH2) (15,26). The volume of the PCR mixture was 25 l and contained PCR buffer, 3.5 mM MgCl 2 , 200 M deoxyribonucleotides, 50 pmol of each primer, and 1 U of recombinant Taq polymerase (Invitrogen).…”

“…Few studies have investigated the genetic diversity of PspA (15,23,26,27), and scarce information focusing on PspA from strains derived from carriage pneumococci has been published (8). We therefore conducted this study to investigate the PspA families and clades collected from nasopharyngeal carriage isolates of community-acquired S. pneumoniae in children during an ongoing surveillance.…”

Immunization of Mice with Single PspA Fragments Induces Antibodies Capable of Mediating Complement Deposition on Different Pneumococcal Strains and Cross-Protection

“…A66.1 was first reported to express only PspA2 (1), but subsequent work has shown it to express both PspA1 and PspA2 (3). We have amplified a single pspA band from this strain using primers LSM12 and SKH2 (17), and sequencing of cloned fragments confirmed only pspA2 clones. The expression of only PspA2 by A66.1 would be in accordance with the data showing protection after immunization with PspA2, but not with PspA1.…”