...funding research, sharing discoveries.
Lawrence C Paoletti
Associate Professor of Medicine, Brigham and Women's Hospital
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Human vaccines 2009 Jun; 5(7)
A fluorescence-based opsonophagocytosis assay to measure the functional activity of antibody to group B Streptococcus.
An in vitro assay designed to measure the functional activity of vaccine-induced antibody is a necessary component of any vaccine development program. Because traditional efficacy studies of vaccines to prevent neonatal diseases caused by group B Str... expand abstracteptococcus (GBS) are unlikely given the effectiveness of current antibiotics and screen-based surveillance practices, the ability to efficiently and effectively measure functional antibody responses may be of particular importance. GBS, like other encapsulated bacterial pathogens, are susceptible to opsonization by specific antibody and complement and subsequent killing by the host's effector cells. The in vitro opsonophagocytosis and killing assay (OPA) mimics this in vivo defense strategy and has been used for decades to measure the functionality of natural and/or vaccine-induced GBS-specific antibody. Here we describe a fluorescence-based OPA (flOPA) that measures the ability of specific antibody to opsonize fixed, fluorescently labeled GBS or antigen-coated fluorescent microspheres for uptake by differentiated HL-60 cells in the presence of complement. Compared to the classical OPA, the flOPA is standardized with respect to effector cells, complement and antigenic targets. The GBS flOPA is also less time-intensive and has the potential to measure antibody to multiple antigens simultaneously. Quantitative functional antibody determinations using the flOPA may serve as a surrogate measure of GBS vaccine effectiveness in lieu of traditional phase 3 efficacy trials. collapse abstract
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Human vaccines 4(6)
Neither antibody to a group B streptococcal conjugate vaccine nor the vaccine itself is teratogenic in rabbits.
Group B Streptococcus (GBS) is a leading cause of human neonatal bacterial disease, resulting in pneumonia, sepsis, meningitis and sometimes, death. Supportive preclinical studies of GBS capsular polysaccharide (CPS)-protein conjugate vaccines have l... expand abstracted to several phase 1 and phase 2 trials in healthy, non-pregnant adults, which demonstrated that the vaccines, produced at the Channing Laboratory, were safe and immunogenic. However, evaluation of the safety and immunogenicity of a GBS conjugate vaccine administered to pregnant women demanded that it be manufactured under current good manufacturing practices (cGMP) and that it undergo developmental toxicity evaluation. In this report, we describe a GBS type III CPS-tetanus toxoid (III-TT) vaccine lot 3-1-96 manufactured and vialed under cGMP and our evaluation of the effect of this vaccine and of GBS type III CPS-specific antibody on conception and early- and late-stage fetal development in rabbits. III-TT lot 3-1-96 was compositionally similar to prototype III-TT lot 91-1, produced under non-GMP, and was potent in a mouse maternal vaccination-neonatal pup challenge model of GBS disease. Four groups of 30 female rabbits each were randomized to receive III-TT lot 3-1-96 vaccine, saline-alum, or combinations of these treatments before and after insemination. The dose of conjugated CPS on a weight basis was 1 microg/kg, mimicking the anticipated actual human dose. Based on the weight of the rabbits, this was 20- to 100-fold greater than the expected human dose. Does were pre-assigned to deliver litters naturally or have their kits delivered by Caesarean-section at gestation day 29, to assess late fetal development. Sera from does and kits were collected, and the presence of type III CPS-specific IgG was confirmed by quantitative ELISA. Based on all assessments, GBS type III-TT lot 3-1-96, nor antibody to it did not affect embryo fetal viability, sex ratio, growth or cause malformations (i.e., it was non-teratogenic). In addition, that III-TT lot 3-1-96 was found to be safe and immunogenic in two clinical studies involving healthy non-pregnant adults supports a clinical evaluation of this vaccine in pregnant women. collapse abstract
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Human vaccines 4(5)
Functional activity of antisera to group B streptococcal conjugate vaccines measured with an opsonophagocytosis assay and HL-60 effector cells.
Conjugate vaccines against group B Streptococcus (GBS), which is a leading cause of bacterial disease among newborns and the elderly with underlying illnesses, have progressed from animal studies to phase 1 and 2 clinical trials in healthy adults. Du... expand abstracte to the wide-spread use of antibiotics to treat at-risk deliveries, a phase 3 efficacy trial of a GBS vaccine to prevent neonatal disease in the United States is unlikely. A viable approach to assess a vaccine's efficacy is to use a surrogate of protection which in the case of GBS is the opsonizing activity of serum antibody. The opsonophagocytosis assay (OPA) measures the ability of serum antibody to opsonize GBS for killing by effector cells in the presence of complement. In this report we demonstrate that differentiated HL-60 cells can substitute for human peripheral blood leukocytes (hPMNLs) in the OPA. Antisera to GBS type Ia CPS and type III CPS conjugate vaccines opsonized homologous GBS for killing at effector cells to GBS ratios of 2-4:1 regardless of whether HL-60 or hPMNLs were used. These results represent the first important step in developing a standardized, high-throughput OPA that could be used to assess the functional activity of vaccine-induced antibody and potentially serve as a surrogate of efficacy. collapse abstract
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Proceedings of the National Academy of Sciences of the United States of America 2008 Sep; 105(35)
Role of glycan synthesis in colonization of the mammalian gut by the bacterial symbiont Bacteroides fragilis.
Bacteroides species are the most abundant Gram-negative bacteria of the human colonic microbiota. These endogenous organisms are unique in that they synthesize an extensive number of phase-variable surface polysaccharides. Pathogenic bacteria phase v... expand abstractary expression of surface molecules for immune evasion, but the importance of the synthesis of multiple phase-variable polysaccharides to these commensal bacteria is unknown. We previously showed that a Bacteroides fragilis mutant unable to synthesize 4 of the 8 capsular polysaccharides and unable to glycosylate proteins properly is rapidly outcompeted by the wild-type strain for colonization of the gnotobiotic mouse intestine. In the present study, we constructed mutants defective only in capsule polysaccharide synthesis to define better the importance of these surface molecules to intestinal colonization. We discovered a key enzymatic activity required for synthesis of 7 of the 8 capsular polysaccharides. Deletion of its gene resulted in the first B. fragilis mutant able to synthesize only one phase-variable polysaccharide, and further mutation resulted in a stable acapsular mutant. We show that the acapsular mutant is rapidly outcompeted, but synthesis of a single polysaccharide is sufficient for the organism to colonize the gnotobiotic intestine competitively. These data demonstrate that initial colonization of the gnotobiotic mouse intestine by B. fragilis requires that the organism synthesize only a single polysaccharide and suggest that the synthesis of multiple phase-variable polysaccharides is important for the bacteria's long-term maintenance in the normally complex and competitive ecosystem. collapse abstract
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Clinical and vaccine immunology : CVI 2008 Jun; 15(7)
Recombinant group B Streptococcus alpha-like protein 3 is an effective immunogen and carrier protein.
Conjugate vaccines against pathogens of multiple serotypes are optimized when all components induce functional antibody, resulting in broadened coverage. While most clinical studies of vaccines against group B Streptococcus (GBS) have evaluated conju... expand abstractgates composed of capsular polysaccharide (CPS) coupled to tetanus toxoid, conjugates prepared with GBS proteins as carriers have also been efficacious in animals. Here, we report that recombinant GBS alpha-like protein 3 (rAlp3) is both a strong immunogen and a viable carrier protein for type III CPS. The type III CPS-specific immunoglobulin G (IgG) titer rose from <100 to 64,000 among mice that received type III CPS coupled to rAlp3 (III-rAlp3) compared with an absence of a specific response among mice that received an uncoupled mixture. Most (94%) newborn pups born to III-rAlp-vaccinated dams survived challenge with viable type III GBS, compared with 43% survival among those born to dams that received the uncoupled mixture (P < 0.0001). A tricomponent conjugate of type III CPS, rAlp3, and a GBS recombinant beta C protein lacking its IgA binding site (III-rAlp3-rBCP(DeltaIgA)) provided protection against a serotype III strain and a serotype Ia strain bearing beta C protein. High-titered anti-rAlp3 rabbit serum opsonized Alp3-containing strains of two GBS serotypes (types V and VIII) and invasive type III strains bearing the cross-reactive Rib protein for in vitro killing by human peripheral blood leukocytes. Thus, the potential exists for the inclusion of rAlp3 in a GBS vaccine formulated to provide multiserotype coverage. collapse abstract
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Proceedings of the National Academy of Sciences of the United States of America 2008 Apr; 105(15)
Rational chemical design of the carbohydrate in a glycoconjugate vaccine enhances IgM-to-IgG switching.
Many pathogens are sheltered from host immunity by surface polysaccharides that would be ideal as vaccines except that they are too similar to host antigens to be immunogenic. The production of functional IgG is a desirable response to vaccines; beca... expand abstractuse IgG is the only isotype that crosses the placenta, it is of particular importance in maternal vaccines against neonatal disease due to group B Streptococcus (GBS). Clinical studies found a substantially lower proportion of IgG-relative to IgM-among antibodies elicited by conjugates prepared with purified GBS type V capsular polysaccharide (CPS) than among those evoked by CPSs of other GBS serotypes. The epitope specificity of IgG elicited in humans by a conjugate prepared with type V CPS is for chemically desialylated type V CPS (dV CPS). We studied desialylation as a mechanism for enhancing the ability of type V CPS to induce IgM-to-IgG switching. Desialylation did not affect the structural conformation of type V CPS. Rhesus macaques, whose isotype responses to GBS conjugates match those of humans, produced functionally active IgG in response to a dV CPS-tetanus toxoid conjugate (dV-TT), and 98% of neonatal mice born to dams vaccinated with dV-TT survived lethal challenge with viable GBS. Targeted chemical engineering of a carbohydrate to create a molecule less like host self may be a rational approach for improving other glycoconjugates. collapse abstract
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Infection and immunity 2007 Jun; 75(7)
Recombinant group B streptococcus Beta C protein and a variant with the deletion of its immunoglobulin A-binding site are protective mouse maternal vaccines and effective carriers in conjugate vaccines.
Immunogenic vaccines against group B Streptococcus (GBS) have been created by coupling the GBS capsular polysaccharides (CPS) to carrier proteins. The GBS beta C protein (BCP) serves as an effective carrier while inducing protective immunity against ... expand abstractBCP-expressing strains. BCP also binds human immunoglobulin A (IgA), a characteristic that may be undesirable for use in humans. Here, we examined the immunogenicity and protective efficacy of a recombinant GBS BCP (rBCP), an rBCP modified to eliminate its IgA-binding site (rBCP(DeltaIgA)), and their corresponding GBS serotype III CPS conjugates (III-rBCP and III-rBCP(DeltaIgA)). Deletion of the IgA-binding site or conjugation to CPS did not alter antigenic BCP epitopes. Recombinant proteins and conjugates elicited specific, high-titered IgG in mice. Antisera to rBCP, rBCP(DeltaIgA), III-rBCP, and III-rBCP(DeltaIgA) opsonized GBS strains A909 (Ia/BCP(+)) and H36B (Ib/BCP(+)) for killing by HL-60 cells; antiserum to III-rBCP and III-rBCP(DeltaIgA) also opsonized strain M781 (III/BCP(-)). Vaccination of female mice with either rBCP or rBCP(DeltaIgA) protected approximately 40% of their pups challenged with GBS strain A909. Pups born to III-rBCP- or III-rBCP(DeltaIgA)-vaccinated dams survived at rates of 56% and 66%, respectively. Over 90% of pups born to dams that received the type III CPS conjugates survived challenge with GBS strain M781. In summary, rBCP and rBCP(DeltaIgA) proteins and the conjugates containing them were immunogenic in mice, inducing both CPS- and protein-specific functional IgG. These results suggest that the rBCP(DeltaIgA) could be used as a carrier to augment the immunogenicity of the CPS while expanding coverage to GBS strains bearing BCP. collapse abstract
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Infection and immunity 2007 Feb; 75(3)
Transcriptional and proteomic profiles of group B Streptococcus type V reveal potential adherence proteins associated with high-level invasion.
Group B Streptococcus (GBS) is an opportunistic organism that can harmlessly colonize the human gut, vagina, and rectum but can also cause pneumonia, sepsis, and meningitis in neonates born to colonized mothers. We have shown previously that growth r... expand abstractate and oxygen level regulate the ability of GBS to invade eukaryotic cells in vitro. Herein we extend and expand on these observations to show that GBS type V, an emergent serotype, grown in a chemostat at a cell mass-doubling time (t(d)) of 1.8 h with oxygen invaded human ME-180 cervical epithelial cells in large numbers compared with those grown at the same t(d) without oxygen or at a slower t(d) of 11.0 h. The fact that several GBS type V cell wall-associated and membrane proteins were expressed exclusively under the invasive growth condition prompted an investigation, using genomics and proteomics, of all upregulated genes and proteins. Several proteins with potential roles in adherence were identified, including an undefined surface antigen (SAG1350), a lipoprotein (SAG0971), penicillin-binding protein 2b (SAG0765), glyceraldehyde-3-phosphate dehydrogenase (SAG0823), and an iron-binding protein (SAG1007). Mouse antisera to these five proteins inhibited binding of GBS type V to ME-180 cells by > or =85%. Recombinant undefined surface antigen (SAG1350), lipoprotein (SAG0971), and penicillin-binding protein 2b (SAG0765) each bound to ME-180 cells in a dose-dependent fashion, confirming their ability to act as ligands. Collectively, these data increase the number of potential GBS adherence factors and also suggest a role for these surface-associated proteins in initial pathogenic events. collapse abstract
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Nature reviews. Microbiology 2006 Nov; 4(12)
Group B Streptococcus: global incidence and vaccine development.
An ongoing public health challenge is to develop vaccines that are effective against infectious diseases that have global relevance. Vaccines against serotypes of group B Streptococcus (GBS) that are prevalent in the United States and Europe are not ... expand abstractoptimally efficacious against serotypes common to other parts of the world. New technologies and innovative approaches are being used to identify GBS antigens that overcome serotype-specificity and that could form the basis of a globally effective vaccine against this opportunistic pathogen. This Review highlights efforts towards this goal and describes a template that can be followed to develop vaccines against other bacterial pathogens. collapse abstract
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Vaccine 2007 Jan; 25(1)
Dose-response to type V group B streptococcal polysaccharide-tetanus toxoid conjugate vaccine in healthy adults.
A phase 1, dose-escalating trial was conducted in healthy adults to evaluate immunogenicity and reactogenicity of a type V group B streptococcal (GBS) capsular polysaccharide (CPS)-tetanus toxoid (TT) conjugate vaccine. Participants received one dose... expand abstract of unconjugated V CPS (37 microg), V-TT (2.4 microg CPS/1.1 microg TT), V-TT (9.6 microg CPS/4.3 microg TT) or V-TT (38.5 microg CPS/17.0 microg TT). Each vaccine and all doses of V-TT were well-tolerated. V CPS-specific antibodies reached a peak 4-8 weeks after immunization and were significantly higher through 52 weeks post-immunization in recipients of V-TT at each dose than in uncoupled CPS vaccinees. V-TT vaccine-induced antibodies promoted opsonophagocytic killing of type V GBS and avidity maturation of V CPS-specific IgG. collapse abstract
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Journal of clinical microbiology 2006 Jun; 44(7)
Molecular characterization of nontypeable group B streptococcus.
Traditionally, the capsular polysaccharide (CPS) antigen has been used to distinguish between the nine known serotypes of group B streptococcus (GBS) by classical antibody-antigen reactions. In this study, we used PCR for all CPSs and selected protei... expand abstractn antigens, multilocus sequencing typing (MLST), and pulsed-field gel electrophoresis (PFGE) to molecularly characterize 92 clinical isolates identified as nontypeable (NT) by CPS-specific antibody-antigen reactivity. The PCR and MLST were performed on blinded, randomly numbered isolates. All isolates contained the cfb gene coding for CAMP factor. While most (56.5%) contained a single CPS-specific gene, 40 isolates contained either two or three CPS-specific genes. Type V CPS-specific gene was present in 66% of the isolates, and all serotypes except types IV, VII, and VIII were represented. Most (44.5%) of the isolates contained a single protein antigen gene (bca, bac, rib, alp1, or alp3), and the remaining isolates had multiple protein antigen genes. Of the 61 isolates that had the V CPS-specific gene, 48 (78.6%) had the alp3 gene. PFGE analysis classified the isolates into 21 profile groups, while MLST analysis divided the isolates into 16 sequence types. Forty-two (69%) of 61 isolates with the V CPS-specific gene were in PFGE profile group 4; 41 of these 42 were sequence type 1 by MLST. These data shed new light on the antigenic complexity of NT GBS isolates, information that can be valuable in the formulation of an effective GBS vaccine. collapse abstract
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Journal of medical microbiology 2006 May; 55(Pt 6)
Identification of novel cps locus polymorphisms in nontypable group B Streptococcus.
Group B Streptococcus (GBS) is an important pathogen responsible for a variety of diseases in newborns and the elderly. A clinical GBS isolate is considered nontypable (NT) when serological methods fail to identify it as one of nine known GBS serotyp... expand abstractes. Eight clinical isolates (designated A1-A4, B1-B4) showed PFGE profiles similar to that of a GBS serotype V strain expressing R1, R4 surface proteins. These unique isolates were further characterized by immunologic and genetic methods. Rabbit sera to isolates A1 and A2 reacted weakly with concentrated HCl extracts of A1-A4 isolates, but not with those of B1-B4 isolates. In addition, a type V capsular polysaccharide (CPS) inhibition ELISA revealed that cell wall extracts from isolates A1-A4, but not from B1-B4, expressed low but measurable amounts of type V CPS. Molecular serotyping with PCR analysis showed that all eight isolates contained a type V-specific CPS gene (cpsO) and harboured the gene encoding the surface protein Alp3. Multilocus sequence typing identified isolate A1 as belonging to a new sequence type (ST) designated ST-173, whereas the other seven isolates keyed to ST-1. Sequencing of the 18 genes (17 736 bp) in the cps locus showed that each NT isolate harboured one to three unique polymorphisms, and also identified an IS1381 element in cpsE of the B4 isolate. Collectively, genetic and immunologic analyses revealed that these NT isolates expressing R1, R4 proteins have a genetic profile consistent with that of type V, an emergent, antigenically diverse and increasingly prevalent GBS serotype. collapse abstract
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Science (New York, N.Y.) 2005 Jun; 309(5731)
Identification of a universal Group B streptococcus vaccine by multiple genome screen.
Group B Streptococcus (GBS) is a multiserotype bacterial pathogen representing a major cause of life-threatening infections in newborns. To develop a broadly protective vaccine, we analyzed the genome sequences of eight GBS isolates and cloned and te... expand abstractsted 312 surface proteins as vaccines. Four proteins elicited protection in mice, and their combination proved highly protective against a large panel of strains, including all circulating serotypes. Protection also correlated with antigen accessibility on the bacterial surface and with the induction of opsonophagocytic antibodies. Multigenome analysis and screening described here represent a powerful strategy for identifying potential vaccine candidates against highly variable pathogens. collapse abstract
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Vaccine 2005 May; 23(24)
The projected health benefits of maternal group B streptococcal vaccination in the era of chemoprophylaxis.
While maternal antibiotic prophylaxis has greatly reduced early-onset group B streptococcal (GBS) disease in the United States, a GBS vaccine currently under development could potentially prevent additional GBS cases and preterm births. A decision an... expand abstractalytic model was created to compare preventive strategies using adolescent, maternal (prenatal), or postpartum vaccination with selective chemoprophylactic strategies. The current practice of culture-based chemoprophylaxis was predicted to prevent 55% of early plus late-onset GBS infections. Maternal vaccination strategies were superior to current practice, preventing 68-69% of all GBS infections and 4% of very preterm births (<32 weeks gestation). The most effective adolescent vaccination strategy combined vaccination with culture-based chemoprophylaxis for all women and prevented 66% of all GBS infections. All other strategies were similar in efficacy to current practice or inferior. Maternal GBS vaccination is predicted to prevent more cases of neonatal GBS disease than current practice and would prevent approximately one in 25 very preterm births. collapse abstract
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Canadian journal of microbiology 2005 Mar; 51(4)
Growth rate and oxygen regulate the interactions of group B Streptococcus with polarized respiratory epithelial cells.
We investigated growth conditions that regulate the ability of group B Streptococcus (GBS) to attach to, invade, and translocate through polarized human respiratory epithelial cells (RECs). GBS grown in a chemostat at a fast cell mass doubling time (... expand abstractt(d) = 1.8 h), invaded RECs from both the apical and basolateral surfaces in higher numbers compared with those held at a t(d) = 11 h. With the exception of adherence from the basolateral surface, GBS reached peak adherence to, invasion of, and translocation through RECs when held at the fast t(d) with 15% dissolved oxygen compared with 0% dissolved oxygen. Growth rate and oxygen level strongly influence the interaction of GBS with polarized RECs and likely GBS pathogenicity. collapse abstract
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Infection and immunity 2004 Jul; 72(8)
Adherence to, invasion by, and cytokine production in response to serotype VIII group B Streptococci.
The adherence to and invasion of the human epithelial cell line A549 by group B streptococcus (GBS) serotype VIII strains were compared with those of serotype III strains by a conventional method and the dynamic in vitro attachment and invasion syste... expand abstractm. Twenty GBS strains, including nine vaginal isolates and one invasive isolate each of serotypes III and VIII, were used in the conventional attachment and invasion assay. Adherence to and invasion of A549 cells by serotype VIII GBS strains were significantly greater (P < 0.0001) than those by serotype III strains for both the invasive strain and vaginal isolates. Cytokine production by A549 cells following stimulation with GBS serotypes III and VIII or their purified capsular polysaccharides (CPS) was measured. Serotype III strains stimulated significantly greater tumor necrosis factor alpha (TNF-alpha) (P < 0.0001) and interleukin-10 (IL-10) (P < 0.05) production than did serotype VIII strains. IL-8 production in response to serotype VIII was significantly higher (P < 0.001) than that in response to serotype III. TNF-alpha, IL-8, and IL-10 production was greater in A549 cells infected with GBS than in the untreated control cells. TNF-alpha production was significantly greater (P < 0.005) after stimulation with purified GBS serotype III CPS than after stimulation with serotype VIII CPS, a result similar to that after stimulation with whole GBS. IL-12 production by A549 cells was observed only in response to infection with GBS serotype III, resulting in the possibility of a greater TH1 response in serotype III GBS. These results suggest differences in immune responses to infection with GBS serotypes III and VIII. collapse abstract
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The Indian journal of medical research 2004 Apr; 119 Suppl
Characterization of vaginal rectal colonization with multiple serotypes of group B streptococci using multiple colony picks.
BACKGROUND & OBJECTIVES: There is paucity of information on vaginal and rectal colonization with multiple serotypes of group B streptococci (GBS). As part of an ongoing cohort study evaluating the natural history of vaginal and rectal colonization by... expand abstract GBS, the colonization with multiple serotypes was studied in 102 non-pregnant women aged 18-30 yr. METHODS: Up to ten separate colony picks of beta-haemolytic streptococci (total 1515 isolates) were selected from vaginal and rectal primary culture plates. The colonies were identified as GBS, and their capsular polysaccharides (CPS) serotypes determined using monospecific rabbit antisera for types Ia-VIII by double immunodiffusion in agarose (DID). A colony dot immunoblot (DB) assay, using monospecific rabbit antisera to purified type polysaccharides conjugated to tetanus toxoid, was developed to serotype efficiently the multiple colony picks of GBS. RESULTS: The CPS serotype distribution, examining only the 177 "a" or first colony picks from the 102 patients, was 30.5 per cent for Ia; 28.2 per cent for type III; 15.3 per cent for type II; and 13.6 per cent for type V. Only 2.8 per cent were nontypeable. Eighty of the 102 patients (78.4%) were colonized with only one serotype; 20 (19.6%) had two serotypes and two patients (2%) had three serotypes in their vaginal and/or rectal paired cultures. Overall, 91.9 per cent of the culture sites colonized with one to three CPS types (from the total number of colonies picked) were identified with a minimum of three colony picks. In 75 patients with vaginal/rectal pairs the GBS serotype concordance of only the "a" colony was 89.3 per cent and concordance decreased to 80 per cent when the serotype concordance of the total colony picks was analyzed. INTERPRETATION & CONCLUSION: In conclusion, there was a relatively high prevalence of serotype nonconcordance in this population, and 21.6 per cent of patients had multiple GBS serotypes. collapse abstract
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The Journal of infectious diseases 2004 Mar; 189(6)
Immune response of healthy women to 2 different group B streptococcal type V capsular polysaccharide-protein conjugate vaccines.
BACKGROUND: Infections caused by group B streptococcal (GBS) type V are increasingly common. Capsular polysaccharide (CPS)-protein conjugate GBS vaccines are immunogenic in healthy adults, but type V vaccines have not previously been tested. METHODS:... expand abstract Thirty-five healthy, nonpregnant women were randomized to receive an intramuscular dose of GBS type V CPS-tetanus toxoid (TT) vaccine (n=15), GBS type V CPS-cross-reactive material (CRM(197)) conjugate vaccine (n=15), or placebo (n=5) (double-masked design). Levels of serum antibodies to type V CPS were measured by ELISA, and functional activity was measured by opsonophagocytosis. RESULTS: The vaccines were well tolerated. Significant increases in type V CPS-specific immunoglobulin G (IgG) were elicited by both vaccines, peaking at 4-8 weeks and persisting for 26 weeks. Four-fold or greater increases in type V CPS-specific IgG concentrations were noted in postimmunization serum samples obtained from 93% of subjects in each vaccine group. These concentrations persisted in > or =85% of conjugate-vaccine recipients 104 weeks later. Type V CPS-specific immunoglobulin M was a dominant isotype of immune response to each conjugate. Postimmunization serum samples promoted opsonophagocytic killing of GBS type V in vitro, whereas those from placebo recipients did not. CONCLUSION: GBS type V conjugate vaccines are safe and immunogenic and would be appropriate for inclusion in a candidate multivalent GBS vaccine. collapse abstract
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Infection and immunity 2003 Nov; 71(12)
Oxygen regulates invasiveness and virulence of group B streptococcus.
The facultative anaerobe group B Streptococcus (GBS) is an opportunistic pathogen of pregnant women, newborns, and the elderly. Although several virulence factors have been identified, environmental factors that regulate the pathogenicity of GBS have... expand abstract not been well characterized. Using the dynamic in vitro attachment and invasion system (DIVAS), we examined the effect of oxygen on the ability of GBS to invade immortalized human epithelial cells. GBS type III strain M781 invaded human epithelial cells of primitive neurons, the cervix, the vagina, and the endometrium in 5- to 400-fold higher numbers when cultured at a cell mass doubling time (t(d)) of 1.8 h than at a slower t(d) of 11 h. Invasion was optimal when GBS was cultured at a t(d) of 1.8 h in the presence of >or=5% oxygen and was significantly reduced without oxygen. Moreover, GBS grown in a chemostat under highly invasive conditions (t(d) of 1.8 h, with oxygen) was more virulent in neonatal mice than was GBS grown under suboptimal invasion conditions (t(d) of 1.8 h, without oxygen), suggesting a positive association between in vitro invasiveness with DIVAS and virulence. collapse abstract
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Expert opinion on biological therapy 2003 Aug; 3(6)
Glycoconjugate vaccines to prevent group B streptococcal infections.
Group B Streptococcus (GBS) is an opportunistic pathogen of humans. At-risk populations include neonates born to colonised mothers, peripartum women, diabetics, and the elderly with underlying illnesses. Vaccines to prevent GBS disease have been deve... expand abstractloped by coupling purified capsular polysaccharide (CPS) antigen of GBS with an immunogenic protein carrier. Glycoconjugate vaccines against all nine currently identified GBS serotypes have been synthesised and shown to be immunogenic in mice, rabbits and baboons in preclinical trials. Healthy adults have safely received conjugate vaccines prepared with GBS types Ia, Ib, II, III, and V CPSs in Phase I and II clinical trials. These vaccines elicited CPS-specific antibody that opsonised GBS for in vitro killing by human peripheral blood leukocytes in the presence of complement. Results from these preclinical and clinical studies strongly suggest that GBS conjugate vaccines will be effective in preventing diseases caused by GBS. collapse abstract
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The Journal of infectious diseases 2003 Jun; 188(1)
Safety and immunogenicity of a bivalent group B streptococcal conjugate vaccine for serotypes II and III.
To determine whether 2 monovalent group B streptococcus (GBS) serotype II or III capsular polysaccharide (CPS)-tetanus toxoid (TT) conjugate vaccines combined in a single intramuscular dose would elicit immune responses comparable to those of monoval... expand abstractent vaccines, 75 healthy adults were randomized to receive GBS II-TT (3.6 micro g of CPS), GBS III-TT (12.5 micro g of CPS), or a bivalent mixture of GBS II-TT/III-TT vaccine (double-masked design). Vaccines were well tolerated. Four-fold or greater increases in GBS II or III CPS-specific IgG, respectively, were noted in postimmunization serum samples from 80%-90% of bivalent conjugate vaccine recipients, and these responses were similar to those of recipients of GBS II-TT or GBS III-TT monovalent vaccines. Immune serum samples promoted the opsonophagocytic killing of types II and III GBS in vitro. Unexpectedly, some recipients of these vaccines developed cross-reactive antibodies to the structurally similar heterologous polysaccharide. These results support the feasibility of a multivalent vaccine for the 5 prevalent invasive disease-causing GBS CPS serotypes. collapse abstract
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The Journal of infectious diseases 2002 Nov; 186(11)
Neutralizing antibody induced in mice by novel glycoconjugates of human immunodeficiency virus type 1 gp120 and env2-3.
We sought to determine whether conjugation methods could improve the immunogenicity of human immunodeficiency virus (HIV)-1 gp120 and env2-3, a nonglycosylated form of gp120. Conjugate vaccines were created by coupling tetanus toxoid (TT), gp120, and... expand abstract/or env2-3 with group B streptococcal type III polysaccharide (III) by reductive amination. Size-exclusion chromatography revealed polymers larger than those of the uncoupled proteins after conjugation. Geometric mean titers of gp120-specific immunoglobulin G (IgG) in serum from mice given env2-3-TT-III or env2-3-gp120-TT-III conjugates increased from <50 before to 3031 and 4756, respectively, after vaccination, whereas an uncoupled mixture of gp120, TT, and III, and III-TT not coupled with gp120 or env2-3 were poorly immunogenic. Pooled serum diluted 1rcolon;20 from mice given env2-3-gp120-TT-III inhibited infection of HIV-1(MN) of Sup-T1 cells and demonstrated neutralizing activity against infection with primary isolate HIV-1(BR014). HIV-1 gp120-specific IgG with neutralizing activity against HIV-1 can be induced with conjugates containing gp120, env2-3, III, and TT. collapse abstract
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Seminars in neonatology : SN 2002 Jul; 7(4)
Vaccines to prevent neonatal GBS infection.
Group B Streptococcus (GBS) remains the leading bacterial cause of neonatal sepsis and meningitis in the United States. Although antibiotic prophylaxis has decreased the infection rate, the best long-term solution lies in the development of effective... expand abstract vaccines. The GBS capsular polysaccharide (CPS) is a major target of antibody-mediated immunity. While antibody to CPS is protective, uncoupled CPS is variably immunogenic in humans, a finding that led to the development of GBS CPS-protein conjugate vaccines. GBS CPS-protein conjugate vaccines of all clinically important serotypes have been produced and tested in animals. Mice and baboons immunized with CPS conjugates transplacentally transferred functionally active GBS-specific IgG to their offspring. Phase 1 and phase 2 clinical trials have shown that GBS conjugate vaccines are safe, well-tolerated and immunogenic in healthy adults. Moreover, human antibodies elicited by the conjugate vaccines are functionally active both in vitro and in animal models of invasive GBS disease. collapse abstract
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The Journal of infectious diseases 2002 Jun; 186(1)
Conjugate vaccines against group B Streptococcus types IV and VII.
Although rarely encountered, group B Streptococcus (GBS) types IV and VII have been isolated from infants and adults with invasive disease. This study was designed to determine the immunogenicity and efficacy in animals of conjugate vaccines prepared... expand abstract with GBS types IV and VII capsular polysaccharide (CPS). Despite the striking similarities in structure of these 2 carbohydrate antigens, high-titer rabbit antiserum to each conjugate vaccine was serotype specific. Active vaccination of female mice with the conjugate vaccines induced type-specific IgG and resulted in survival of >90% of newborn pups challenged with viable GBS of homologous serotype. If needed, types IV and VII CPS conjugate vaccines of the design described can be added to the formulation of a multivalent GBS vaccine. collapse abstract
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Infection and immunity 2002 Mar; 70(4)
Type III group B streptococcal polysaccharide induces antibodies that cross-react with Streptococcus pneumoniae type 14.
Covalent linkage of a bacterial polysaccharide to a protein greatly enhances the carbohydrate's immunogenicity and its binding to solid surfaces in immunoassays. These findings have spurred the development of glycoconjugate vaccines to prevent seriou... expand abstracts bacterial infections as well as the use of glycoconjugates as coating antigens in bioassays. We evaluated sera from women immunized with unconjugated group B streptococcal (GBS) type III (GBS III) polysaccharide (IIIPS) or with IIIPS covalently linked to tetanus toxoid to assess specificity, sensitivity, and parallelism in dilution curves in two GBS III enzyme-linked immunosorbent assays (ELISAs). One assay used IIIPS mixed with methylated human serum albumin (IIIPS + mHSA) as the coating antigen, and the other used IIIPS covalently linked to HSA (III-HSA). Each coating antigen was associated with a highly specific GBS III bioassay. The sensitivity was higher in the III-HSA ELISA, in which conjugated IIIPS is bound to the plates. Parallelism in titration curves was observed in the III-HSA but not in the IIIPS + mHSA ELISA. The excellent correlation between the concentrations of GBS IIIPS-specific immunoglobulin G (IgG) and the opsonophagocytic activity of these antibodies indicated that the III-HSA assay can predict functionality of vaccine-induced IgG against GBS III disease. The structure of the repeating unit of the capsular polysaccharide of GBS III differs from that of Streptococcus pneumoniae type 14 (Pn14 PS) only by the presence on GBS III of a sialic acid residue at the end of the side chain. The majority of healthy adults responding to GBS III vaccines with a fourfold or greater increase in GBS III-specific IgG antibodies developed antibodies cross-reacting with Pn14 PS (i.e., desialylated GBS IIIPS). The proportion of GBS vaccine responders who developed IgG to the desialylated IIIPS did not depend on whether IIIPS was given in the unconjugated or conjugated form. When present, these vaccine-induced cross-reacting antibodies conferred in vitro antibody-mediated opsonophagocytosis and killing of both GBS III and Pn14, two pathogens that cause invasive disease in young infants. collapse abstract
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