This indicates that MNZ21 and MNZ22 do not belong to serotypes 6A and 6C. (PSs), which greatly increase its virulence (Avery & Dubos, 1931). However, antibodies to the capsule can abrogate the virulence and provide serotype-specific safety (Cole, 1913).S. pneumoniaeas a varieties can produce more than 90 different capsule types (Parket al., 2007b), but not all capsule types are equally common in human being diseases. For instance, prior to the use of conjugate vaccines, several serotypes such as 6A, 6B, 14, 19A, 19F and 23F were responsible for a large number of invasive pneumococcal diseases (Robinsonet al., 2001) and consequently have been extensively studied. Serotypes 6A and 6B have been grouped into serogroup 6 because of their serological similarities. Also, serotypes 6A and 6B consistently differ in their 17 kb capsule gene loci by only one nucleotide in thewciPgene (Mavroidiet al., 2004). Podophyllotoxin This difference generates 6A WciP having a serine at residue 195 but 6B WciP with an asparagine at the same residue (Mavroidiet al., 2004). Serotypes 6A and 6B produce capsular PSs with almost identical structures except for a different rhamnoseribitol linkage (Kamerling, 2000). Recently, we discovered a new member of serogroup 6 which we named 6C (Parket al., 2007b). Serotype 6C is definitely distinguished serologically from serotypes 6A and 6B by its unique monoclonal antibody (mAb) binding profile (Linet al., 2006). Subsequent genetic studies have shown the capsule gene loci of serotypes 6A and 6B Podophyllotoxin havewciN6A(named on the other hand aswciNhere), but the 6C capsule gene locus haswciN6C(orwciN) (Parket al., 2007a). Finally, biochemical studies exposed that 6C PS has a glucose residue in place of the galactose residue present in the 6A or 6B PSs (Parket al., 2007b). Following a finding of serotype 6C, we postulated the presence of a new serotype, which could become produced by mutating the crucial nucleotide in thewciPgene of the 6C Podophyllotoxin capsule gene locus or by inserting thewciNgene into the 6B capsule gene locus (Bratcheret al., 2009). This fresh serotype, which logically could have been named 6D, was provisionally named 6X1 because the fresh serotype had not been found in nature (Bratcheret al., 2009). However, our group continued the search and found out natural isolates expressing serotype 6D in Korea; we describe our findings below. == METHODS == == Bacterial strains and tradition. == TIGR6A, TIGR6B, TIGR6C and TIGR6X1, which are isogenic strains of TIGR4 expressing the four different capsule serotypes of serogroup 6, were used as settings for the assays (Bratcheret al., 2009). Fourteen serogroup 6 isolates, including MNZ21 and MNZ22, were from the nasopharyngeal ethnicities of 14 healthy children (less than 5 years old) going to a day-care centre in Jeju Island in Korea in 2008 and were subsequently transferred to the Nahm laboratory for further analysis. These isolates showed the typical-haemolytic colony morphology ofS. pneumococcuson blood agar plates, were susceptible to optochin, and were bile soluble. All bacteria were cultivated in ToddHewitt broth (BD Biosciences) supplemented with 0.5 % yeast extract (THY) and kept frozen at 80 C until used. == Circulation cytometry. == Aliquots of freezing bacteria were thawed, washed, resuspended in FACS buffer (PBS comprising 3 % FBS and kanadaptin 0.1 % sodium azide) and incubated with tradition supernatants of hybridomas (diluted 1 : 10 in FACS buffer) for 20 min at space temperature with shaking. After washing, the bacteria were incubated with fluorescein-conjugated goat antibody to mouse immunoglobulin for 20 min at space heat with shaking. After washing aside unbound goat antibody, the bacteria were resuspended in FACS buffer comprising Syto9 (160 nM) and examined with a circulation cytometer (FACSCalibur, Becton Dickinson). The data were then analysed.
This indicates that MNZ21 and MNZ22 do not belong to serotypes 6A and 6C