Biotyping of Nontypeable Group B Streptococci (original) (raw)
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Journal of Clinical Microbiology, 2008
, 207 (12%) initially nonserotypeable isolates were tested by improved conventional serotyping methods (Lancefield antigen extraction with 0.1 and 0.2 N HCl, latex agglutination assays, and use of antisera against all known serotypes [Ia, Ib, and II to IX]) and a molecular serotype identification system (multiplex PCR-based reverse line blot [mPCR/RLB] assays targeting serotype-specific sites in the region spanning cpsH to cpsM). Serotypes were assigned to 71 (34%) of the 207 isolates by using antisera and to 204 (98.5%) of them by mPCR/RLB. Sequencing of a portion of the cpsE-cpsF-cpsG region of 141 persistently nonserotypeable isolates and 1 with discrepant conventional and molecular serotyping results was attempted. Major mutations were identified in 34 isolates (24%), including 11 (8%) from which no amplicons were obtained and 23 (16%) with sequence variation compared with published sequences; of the latter, 21 (15%) were associated with amino acid changes. By contrast, mutations were identified in only 12 (2.3%) of 516 serotypeable isolates for which this region has been sequenced previously. In summary, an improved serotyping scheme allowed serotype identification of more than one-third of the previously nonserotypeable GBS isolates. Molecular serotypes were assigned to almost all of the isolates by mPCR/RLB. Significant mutations (with no amplicons or with associated amino acid changes) were found in the cpsE-cpsF-cspG region of a higher proportion of nonserotypeable than of serotypeable isolates (32/141 versus 8/516; P < 0.001), but further investigation is needed to determine the genetic basis for most nonserotypeable GBS isolates.
Molecular Characterization of Group B Streptococcus Serotypes By Multiplex Polymerase Chain Reaction
Medical Express, 2017
OBJECTIVE: Group B Streptococcus (GBS) serotypes (Ia, Ib and II to IX) are classified based on variations in their capsular polysaccharide; their prevalence differs between different geographic areas. We examined the prevalence of all GBS serotypes in rectal and vaginal swab samples obtained from 363 pregnant women followed at a Brazilian referral center (Hospital da Mulher Professor Doutor José Aristodemo Pinotti); bacterial susceptibility to antibiotics was further determined. METHOD: Prevalence of positive GBS was evaluated by latex agglutination and by multiplex PCR analysis; bacterial susceptibility to antibiotics, such as clindamycin, erythromycin, levofloxacin, linezolid, penicillin and tetracycline was determined by the disk diffusion method. RESULTS: (a) standard GBS culture and the multiplex PCR analysis tested positive for 83 swabs, collected from 72 women (prevalence of GBS colonization: 72/363; 20%); the most prevalent Serotype was Ia (n=43/83; 52%), followed by serotype V (n=14/83; 17%); according to anatomical origin, serotype Ia accounted for 27/59 (46%) and 16/24 (67%) of the vaginal and rectal samples, respectively; PCR also identified serotypes Ib, II, III and VI. Serotype VI is rarely described and had not been previously reported in Brazil or in Latin America. (b) The latex agglutination test only identified 44 positive samples, all of which were serotyped: 34 of these samples (77%) had serotypes matching those identified by multiplex PCR. (c) Only one sample (serotype Ia) showed resistance to erythromycin and clindamycin. CONCLUSION: Regional studies on GBS serotypes prevalence are essential to guide immunoprophylactic interventions (vaccines) and the implementation of adequate antibiotic prophylaxis or treatment. In this study, the incidence of the serotype VI, a new and rare serotype of GBS was described for the first time in a Brazilian population.
Fems Microbiology Letters, 2006
We used a multiplex PCR-based reverse line blot (mPCR/RLB) hybridization assay and sequencing of a variable region of the cps cluster to identify serotypes of 140 Streptococcus agalactiae (group B Streptococcus; GBS) isolates from cattle. Only 71 (51%) isolates were typeable using antisera, but molecular serotypes (MS) were assigned to 133 (95%) and 139 (99%) isolates by partial cpsE-cpsF-cpsG sequencing and mPCR/RLB, respectively. Ninety-four isolates (67%) belonged to MS III and most belonged to a molecular serosubtype (msst) III-3, which is uncommon among GBS isolates from humans. Our results demonstrate that cps clusters of bovine GBS differ significantly from those of GBS isolates from humans.
Journal of Clinical Microbiology
Direct identification of group B streptococci from a selective broth medium was performed with the Phadebact streptococcus test to determine the feasibility of this technique for early detection of streptococcal colonization. Of 124 clinical isolates, 122 (98.4%) were correctly identified in less than 24 h from the selective broth medium, whereas standard cultures from blood agar plates identified, after 48 h, only 66 (53.2%). The presence of group B streptococci in mixed cultures was always detected by the Phadebact test, and no false-positive co-agglutination tests were observed in 372 cultures from which organisms other than group B streptococci were isolated.
Serotype Identification of Group B Streptococci by PCR and Sequencing
Streptococcus agalactiae) is the most common cause of neonatal and obstetric sepsis and is an increasingly important cause of septicemia in elderly individuals and immunocompromised patients. Ongoing surveillance to monitor GBS serotype distribution will be needed to guide the development and use of GBS conjugate vaccines. We designed sequencing primers based on the previously published sequences of the capsular polysaccharide (cps) gene clusters to further define partial cps gene clusters for eight of the nine GBS serotypes (serotypes Ia to VII). Subsequently, we designed and evaluated primers to identify serotypes Ia, Ib, III, IV, V, and VI directly by PCR and all eight serotypes (serotypes Ia to VII) by sequence heterogeneity. A total of 206 clinical GBS isolates were used to compare our molecular serotype (MS) identification method with conventional serotyping (CS). All clinical isolates were assigned an MS, whereas 188 of 206 (91.3%) were assigned a serotype by use of antisera. A small number of isolates (serosubtypes III-3 and III-4) showed different serotype specificities between PCR and sequencing, but the PCR results correlated with those obtained by CS. The overall agreement between the MS identification method and CS for isolates for which results of both tests were available was 100% (188 of 188 isolates). The MS identification method is a specific and practical alternative to conventional GBS serotyping and will facilitate epidemiological studies.
Journal of Clinical Microbiology, 2011
We report the results from the first international multicenter external quality assessment (EQA) studies for molecular and serological typing of group B streptococcus (GBS) strains as part of DEVANI (Design of a Vaccine against Neonatal Infections), a pan-European program. A questionnaire-based surveillance was undertaken among eight laboratories participating in DEVANI and six laboratories not participating in DEVANI from 13 countries in order to assess their current microbiological procedures for GBS screening, diagnosis, and typing. GBS strains from three EQA distributions were characterized using molecular and serological methods based on GBS capsular polysaccharide typing. Participants were asked to test the first distribution using their current serotyping and genotyping methods. The Strep-B-Latex agglutination method was the most widely used method, with a typeability value of >90%. A multiplex PCR assay for GBS capsular gene typing was also used by 2 of 14 centers, which achieved a typeability value of 93%; this assay detected only 9 of 10 GBS capsular polysaccharide genes. From the second and third EQA studies, standardized protocols were prepared for serological and molecular typing of GBS strains based on the Strep-B-Latex agglutination method and a novel multiplex PCR assay that detected all 10 GBS capsular types (Ia to IX). These standardized protocols are being used by many European laboratories, and as the use of these methods increases, it is imperative to continuously improve and assess laboratory performance and offer training to any laboratories that have technical difficulties.
Biochemical properties and whole-cell protein profiles of group G streptococci isolated from dogs
Journal of Applied Bacteriology
Whole-cell protein profiles obtained by SDS-PAGE were used in conjuction with physiological tests to differentiate strains of Streptococcus canis isolated from dogs. Fermentation of trehalose and lactose, aesculin hydrolysis together with production of b-D-glucuronidase and aD -galactosidase allowed the demonstration of nine different biotypes. However, visual analysis of the protein patterns and comparison by the coefficient of Dice showed minor differences i n band patterns among strains. Only two different profiles were observed. Although a correlation between biotyping and protein profile has been found, this kind of analysis did not provide the basis for a typing method.
A NOVEL METHOD FOR THE PRIMARY DIAGNOSIS OF GROUP A STREPTOCOCCI FROM CLINICAL SPECIMENS.
1997
A novel non-instrumental method for the primary diagnosis of group A streptococci from clinical specimens notably throat swabs is described. The method is based upon a sandwich immunoassay. The assay format consists of a dot-blotting procedure and immunodiffusion on nitrocellulose of pore sizes 0.45um. and 5um. respectively. The detection reagent consists of specific polyclonal rabbit antibodies covalently bound to colloidal carbon particles.