Identification of Newly Described Streptococcus pneumoniae Serotype 6D by Use of the Quellung Reaction and PCR

Abstract

We tested 121 pneumococcal serogroup 6 isolates (including 30 serotype 6C and 24 serotype 6D isolates) by serotype-specific PCR and the Quellung reaction, using "old" and "new" pool B, factor 6b, and new factor 6d antisera. In combination with group B and other factor antisera, factor 6d antiserum can reliably identify the newly described serotype 6D.Serotyping of Streptococcus pneumoniae isolates is an important component of surveillance for pneumococcal disease and assessment of the impact of pneumococcal vaccines. The Quellung reaction is the accepted gold standard method and is used at the New South Wales (NSW) Pneumococcal Reference Laboratory for routine serotyping, supplemented by molecular methods (7,8,11). Recently, we developed serogroup 6 serotype-specific PCRs to identify the newly recognized serotype 6C (6, 10), which also allowed us to identify the first naturally occurring serotype 6D isolates (5).In this study, we compared these PCRs with Quellung reactions, using a new factor, antiserum 6d, which reacts with serotype 6C (Statens Serum Institute, Copenhagen, Denmark). The use of this factor antiserum for identification of serotype 6C has recently been validated (4, 9). We also compared "old" (purchased before 2009) and "new" factor 6b and pool B antisera (purchased in 2009). We tested 64 S. pneumoniae isolates from nasopharyngeal swabs from Fijian children that had been serotyped in 2008 by the Quellung reaction, using old antisera (not including factor 6d), and by PCR (5). They included (i) 24 isolates initially identified as serotype 6B by Quellung reaction but subsequently identified as 6D by PCR (5), (ii) 30 serotype 6C isolates initially identified as 6A by the Quellung reaction but subsequently identified as 6C by PCR (6), and (iii) five isolates identified by the Quellung reaction as serotypes 6A and 6B and confirmed by PCR. In addition, 57 serogroup 6 isolates, referred to the NSW Pneumococcal Reference Laboratory in 2009, were retyped by both the Quellung reaction (using new reagents) and serogroup 6 PCRs (5, 6). To avoid any bias in interpretation, all the isolates were tested in a blind manner.Previous PCR results for Fijian isolates belonging to serotypes 6A, 6B, 6C, and 6D were confirmed by Quellung reactions. Results are shown in Table 1. Serotype 6C isolates reacted with the new (absorbed), but not the old, pool B antiserum. Fourteen serotype 6D isolates did not react with the old pool B antiserum, and 10 showed weak or delayed (and easily missed) reactions; all 24 reacted with the new pool B antiserum, although reactions were very weak for three isolates.Of the 57 NSW isolates, 32 had initially been identified as unusual 6A isolates on the basis of reactions with old group 6 and factor 6b antisera but not with old pool B antiserum. Subsequently, we identified these as serotype 6C by PCR. Consistent with this, the isolates reacted with the old 6b and 6d but not with the new (absorbed) 6b or 6c antiserum (Table 1). Reactions of the remaining NSW isolates (6A and 6B) were the same as...

Multidrug-Resistant Streptococcus pneumoniae Serotype 6D Clones in South Korea

“…To date, S. pneumoniae serotype 6D isolates have been reported in several regions including Fiji, South Korea, China, Japan, Hong Kong, Denmark, Poland, Australia, and Peru (3,4,7,12,16,18,20,21,25). Especially, serotype 6D is relatively prevalent in South Korea, comprising Ͼ10% of serogroup 6 isolates (1, 8).…”

Concurrent Serotyping and Genotyping of Pneumococci by Use of PCR and Electrospray Ionization Mass Spectrometry

“…Similarly, it cannot resolve serotypes 6B and 6D; however, we have found only two serotype 6D isolates within U.S. invasive-disease surveillance within the past 5 years. There are now PCR assays available that can resolve the 4 serogroup 6 serotypes (8,20), which could be added to the PSGS assay.…”

“…The most important single parameter for epidemiological typing of this pathogen is the capsular serotype, for which there are 93 known types (5,7,16,20,22). The capsule is the organism's most important virulence factor, and current licensed vaccines effectively target subsets of specific capsular serotypes.…”

The Future of Synthetic Carbohydrate Vaccines: Immunological Studies on Streptococcus pneumoniae Type 14

“…Recently, new S. pneumoniae serotypes have been identified, e.g. serotype 6C [17], 6D [18,19], and 11E [20]. Capsular polysaccharides are large polymers (0.5-2x10 6 Da), composed of multiple repeating units of up to eight sugar residues [14].…”

“…Today more than 92 serotypes have been identified based on the different chemical structures of these polysaccharides [16,17]. This diversity determines the ability of the serotypes to survive in the bloodstream and very likely the ability to cause invasive disease, especially in [18,19], and 11E [20]. Capsular polysaccharides are large polymers (0.5-2x10 6 Da), composed of multiple repeating units of up to eight sugar residues [14].…”