A SNP-based phylogenetic analysis of Corynebacterium diphtheriae in Malaysia.

OBJECTIVE: There is a lack of study in Corynebacterium diphtheriae isolates in Malaysia. The alarming surge of cases in year 2016 lead us to evaluate the local clinical C. diphtheriae strains in Malaysia. We conducted single nucleotide polymorphism phylogenetic analysis on the core and pan-genome as well as toxin and diphtheria toxin repressor (DtxR) genes of Malaysian C. diphtheriae isolates from the year 1986-2016. RESULTS: The comparison between core and pan-genomic comparison showed variation in the distribution of C. diphtheriae. The local isolates portrayed a heterogenous trait and a close relationship between Malaysia's and Belarus's, Africa's and India's strains were observed. A toxigenic C. diphtheriae clone was noted to be circulating in the Malaysian population for nearly 30 years and from our study, the non-toxigenic and toxigenic C. diphtheriae strains can be differentiated significantly into two large clusters, A and B respectively. Analysis against vaccine strain, PW8 portrayed that the amino acid composition of toxin and DtxR in Malaysia's local strains are well-conserved and there was no functional defect noted. Hence, the change in efficacy of the currently used toxoid vaccine is unlikely to occur.

Bovine monocyte derived dendritic cell based assay for measuring vaccine immunogenicity in vitro.

During both human and animal vaccine development phases, animal testing is necessary to demonstrate vaccine efficacy. Since the number of antigen candidates for testing is usually large when developing a potential vaccine, it is too costly, time consuming and would involve higher risks to carry out selection using in vivo models. The currently available in vitro assays that measure immunogenicity do not adequately reproduce the in vivo state and this is especially true for vaccine research in livestock species. With this in mind, we have developed a bovine monocyte derived dendritic cell (MODC)s based assay to prime CD4 and CD8 lymphocytes in order to investigate vaccine immunogenicity in vitro. MODCs were generated, pulsed with diphtheria toxoid (DT) and co-cultured with lymphocytes for priming. Immunogenicity was measured through antigen recall when antigen pulsed MODC were re-introduced to the co-culture and proliferation of CD4 and CD8 positive lymphocytes were quantified using expressed Ki-67. Having developed the protocol for the assay, we then employed two licenced vaccines against blue tongue virus and rabies virus to validate the assay. Our results show the ability of the assay to satisfactorily measure immunogenicity in cattle. The assay could be used to identify antigens that induce CD4 and CD8 T cell responses prior to embarking on in vivo experiments and can also be used for the quality control of established vaccines in vaccine production facilities as a supplement for in vivo experiments.

Immunogenicity and efficacy of a rationally designed vaccine against vascular endothelial growth factor in mouse solid tumor models.

Vascular endothelial growth factor (VEGF) plays an important role in the progression of various cancers. The VEGF-specific antibody bevacizumab combined with chemotherapy was shown to significantly improve progression-free survival in certain cancers. However, repeated administration is necessary for effective suppression of VEGF, thereby making the therapy expensive and cumbersome. Thus, it is urgent to develop alternative reagents such as VEGF vaccines. Here we report that DTT-VEGF, a VEGF-based antigen consisting of the receptor-binding domain of VEGF and diphtheria toxin T domain (DTT), not only stimulated neutralizing antibody response, but also induced type 1 immune response as well as anti-tumor cytotoxic T lymphocytes in mice when administered with aluminum hydroxide adjuvant. The antibodies triggered by DTT-VEGF immunization inhibited the binding of VEGF to VEGF receptor and downregulated the serum VEGF levels in tumor-bearing mice. VEGF-specific IgG2a and IgG2b antibodies as well as type 1 cytokines were stimulated by DTT-VEGF vaccination. The splenocytes from DTT-VEGF-immunized mice showed cytotoxic activity against B16-F10 cells expressing VEGF. Extensive necrosis with severe hemorrhage and enhanced CD8+ T cell infiltration were observed in tumors from DTT-VEGF-immunized mice. The percentages of CD31+ vascular areas in the tumor sections from DTT-VEGF-immunized mice were significantly lower than those of control mice. DTT-VEGF significantly inhibited tumor growth in preventive and therapeutic vaccination settings in mouse models. Our data suggest that DTT is an effective antigen carrier to break immune self-tolerance and our vaccine design has potential to be used for human cancer therapy.

Development of chitosan-pullulan composite nanoparticles for nasal delivery of vaccines: in vivo studies.

Here, we aimed at developing chitosan/pullulan composite nanoparticles and testing their potential as novel systems for the nasal delivery of diphtheria toxoid (DT). All the chitosan derivatives [N-trimethyl (TMC), chloride and glutamate] and carboxymethyl pullulan (CMP) were synthesised and antigen-loaded composites were prepared by polyion complexation of chitosan and pullulan derivatives (particle size: 239-405 nm; surface charge: +18 and +27 mV). Their immunological effects after intranasal administration to mice were compared to intramuscular route. Composite nanoparticles induced higher levels of IgG responses than particles formed with chitosan derivative and antigen. Nasally administered TMC-pullulan composites showed higher DT serum IgG titre when compared with the other composites. Co-encapsulation of CpG ODN within TMC-CMP-DT nanoparticles resulted in a balanced Th1/Th2 response. TMC/pullulan composite nanoparticles also induced highest cytokine levels compared to those of chitosan salts. These findings demonstrated that TMC-CMP-DT composite nanoparticles are promising delivery system for nasal vaccination.

Divalent toxoids loaded stable chitosan-glucomannan nanoassemblies for efficient systemic, mucosal and cellular immunostimulatory response following oral administration.

The present study reports dual tetanus and diphtheria toxoids loaded stable chitosan-glucomannan nanoassemblies (sCh-GM-NAs) formulated using tandem ionic gelation technique for oral mucosal immunization. The stable, lyophilized sCh-GM-NAs exhibited ~152 nm particle size and ~85% EE of both the toxoids. The lyophilized sCh-GM-NAs displayed excellent stability in biomimetic media and preserved chemical, conformation and biological stability of encapsulated toxoids. The higher intracellular APCs uptake of sCh-GM-NAs was concentration and time dependent which may be attributed to the receptor mediated endocytosis via mannose and glucose receptor. The higher Caco-2 uptake of sCh-GM-NAs was further confirmed by ex vivo intestinal uptake studies. The in vivo evaluation revealed that sCh-GM-NAs posed significantly (p<0.001) higher humoral, mucosal and cellular immune response than other counterparts by eliciting complete protective levels of anti-TT and anti-DT (~0.1 IU/mL) antibodies. Importantly, commercial 'Dual antigen' vaccine administered through oral or intramuscular route was unable to elicit all type of immune response. Conclusively, sCh-GM-NAs could be considered as promising vaccine adjuvant for oral mucosal immunization.

Primer caso de difteria en España después de 30 años. Lecciones aprendidas / No disponible

No disponible

Effects of immunological castration and distiller's dried grains with solubles on carcass cutability and commercial bacon slicing yields of barrows slaughtered at two time points.

Male pigs were randomly assigned to a castration method at birth and allotted to 48 pens (28 pigs/pen). Physically castrated (PC) barrows were castrated at 2 d of age; immunologically castrated (IC) barrows were administered Improvest (GnRF analog diphtheria toxoid conjugate; Zoetis, Kalamazoo, MI) at 16 and 20 wk of age. Distiller's dried grains with solubles (DDGS) feeding strategies included either 0% DDGS (control), 30% DDGS (30% DDGS) fed from 6 wk of age to slaughter, or 30% DDGS fed from 6 wk of age to second dose of Improvest and then fed 0% DDGS until slaughter (withdrawal). Four barrows closest to the median pen weight at 4.5 wk after second dose were selected for evaluation; two were randomly selected and slaughtered at 5 wk and the other two at 7 wk after second dose. Data from each slaughter time were analyzed independently as a 2 × 3 factorial design with pen as the experimental unit. At 5 wk after second dose, bone-in lean cutting yields were 2.63% units greater (P < 0.01) in IC when compared to PC. Bellies were thicker (P < 0.01) and tended to have greater belly flop distances (P = 0.07) in PC compared to IC, however iodine values (IV) were not altered (P = 0.84). Carcass traits (P ≥ 0.10), cutting yields (P ≥ 0.43), and fresh belly characteristics (P ≥ 0.08) were minimally affected by DDGS feeding strategy. Bacon slicing yields (percentage of green weight) were 6.10% units less (P < 0.01) in IC compared with PC. At 7 wk after second dose, bone-in lean cutting yields were 1.57% units greater (P = 0.03) in IC compared with PC. Distiller's grains feeding strategy had no effect (P ≥ 0.83) on boneless carcass cutting yields in IC; while in PC, these yields were 2.32% units less (P < 0.02) in control-fed barrows when compared to other feeding strategies (castration method × feeding strategy; P = 0.03). Bellies from PC tended to be thicker (P = 0.07) and have similar flop distances (P = 0.44) and IV (P = 0.54) when compared with IC. Iodine value was greater (P = 0.03) in 30% DDGS-fed barrows compared with control-fed barrows. Bacon slicing yields (percentage of green weight) were 4.27% units less (P = 0.05) in IC compared with PC. These data suggested that while bacon slicing yield was reduced in IC barrows fed control and 30% DDGS compared with PC barrow counterparts, withdrawal of DDGS improved bacon slicing yields of IC barrows.

Innate immune pathways in afferent lymph following vaccination with poly(I:C)-containing liposomes.

Many modern vaccines use defined adjuvants to stimulate the innate immune system and shape the adaptive immune response. The exact nature of these innate signals and whether immune differentiation can originate within the periphery is not known. Here we used an ovine lymphatic cannulation model to characterise the cellular and transcriptomic profile of the afferent lymph following injection of a liposomal vaccine formulation incorporating diphtheria toxoid and the innate stimulator poly(I:C) over a 78-h period. The response to this vaccine featured an early activation of broad pro-inflammatory pathways (e.g. TLR signalling and inflammasome pathways) and the transient recruitment of granulocytes into the lymph. At 24 h a more monocytic cellular profile arose coinciding with a transition to a specific antiviral response characterised by the up-regulation of genes associated with the receptors typical for the viral mimic, poly(I:C) (e.g. TLR3, RIG-I and MDA5). At the latest time points the up-regulation of IL-17A and IL-17F suggested that Th17 cells may participate in the earliest adaptive response to this vaccine. These data provide the most comprehensive picture of the cellular and molecular mechanisms that link the periphery to the draining lymph node following vaccination, and indicate that the immune response is capable of specialising within the periphery.

An alternative method for purifying and detoxifying diphtheria toxin.

Infections caused by Corynebacterium diphtheriae frequently induce situations in which very small doses of antigens injected intradermally can cause strong inflammatory reactions. This bacterium secretes the diphtheria toxin (DT), a virulence factor that can be lethal to the human organism at doses below 0.1 µg/kg of body weight. The present work proposes alternative methods of DT purification using affinity chromatography and of DT detoxification through conjugating with the polymer methoxypolyethylene glycol activated (mPEG). Tests were performed to evaluate: the formation of edemas and the presence of dermonecrotic activity, in vitro cytotoxicity to Vero cells, the neutralizing activity of serum from guinea pigs immunized with the diphtheria toxoid inactivated with mPEG, and the immunogenic activity of the purified and modified toxin. The results indicated that purification with Blue Sepharose was an efficient method, yielding antigen purity equivalent to 2600 Lf/mg of protein nitrogen. The modification of the Purified Toxin with mPEG did not result in the formation of edema or necrosis although it was immunogenic and stimulated the formation of antibodies that could neutralize the Purified Toxin. The toxoid obtained from the purified toxin maintained its immunogenic characteristics, inducing antibodies with neutralizing activity; edema and necrosis were still observed, however.

Significant systemic and mucosal immune response induced on oral delivery of diphtheria toxoid using nano-bilosomes.

BACKGROUND AND PURPOSE: Over the last decade apprehension has been growing over the effectiveness of existing parenteral vaccines for diphtheria and has created an interest in the development of a mucosally active vaccine. Oral immunization appears to be an effective alternative, but is not without the inherent disadvantages of antigen destruction and tolerance. Therefore, our objective was to investigate the incorporation of diphtheria toxoid (DTx) into bilosomes, which could provide protection as well as aid transmucosal uptake and subsequent immunization. Another objective was to determine the oral dose that will produce serum antibody titres comparable with those produced by i.m. doses of DTx. EXPERIMENTAL APPROACH: Bilosomes containing DTx were prepared by thin film hydration and characterized in vitro for their shape, size, percent antigen entrapment and stability. In the in vivo study the anti-DTx IgG and anti-DTx sIgA response was estimated using elisa, in serum and in various body secretions, respectively, following oral immunization with different doses of DTx entrapped in nano-bilosomes. KEY RESULTS: High dose loaded nano-bilosomes (DTxNB3, 2Lf) produced comparable anti-DTx IgG levels in serum to those induced by i.m. alum-adsorbed DTx (0.5Lf). In addition, all the nano-bilosomal preparations elicited a measurable anti-DTx sIgA response in mucosal secretion, whereas i.m. alum-adsorbed DTx (0.5Lf) was unable to elicit this response. CONCLUSIONS AND IMPLICATIONS: The orally administered nano-bilosomal DTx formulation produced comparable serum antibody titres to i.m.alum-adsorbed DTx, at a fourfold higher dose and without the induction of tolerance. This approach will provide an effective and comprehensive immune protection against diphtheria with better patient compliance.

Comparison of the safety and immunogenicity of an investigational and a licensed quadrivalent meningococcal conjugate vaccine in children 2-10 years of age.

BACKGROUND: Routine administration of quadrivalent meningococcal conjugate vaccine to adolescents and certain high risk groups is recommended in the United States and Canada. We compared the immunogenicity and safety of an investigational quadrivalent meningococcal vaccine conjugated to CRM-197 (MenACWY-CRM) with a licensed quadrivalent vaccine conjugated to diphtheria toxoid (MCV4) in children aged 2-10 years. METHODS: Eligible 2-5-year-olds were randomized 1:2:2 to receive either 2 doses of MenACWY-CRM, or 1 dose of MenACWY-CRM or MCV4; 6-10-year-olds were randomized 1:1 to receive a single dose of MenACWY-CRM or MCV4. The primary immunogenicity assessment was seroresponse separately for the two age cohorts 28 days following a single dose of MenACWY-CRM or MCV4. Noninferiority and superiority criteria were predefined. Solicited injection-site and systemic reactions were collected for the 7 days postvaccination. RESULTS: A total of 2907 children were randomized to receive study vaccine. MenACWY-CRM met statistical superiority criteria vs. MCV4 for groups W and Y and was noninferior for group C in both age strata. For group A, noninferiority criteria were not met; the group A seroresponse rates for MenACWY-CRM and MCV4, respectively were 72% (95% confidence interval 68-75%) and 77% (73-80%) in 2-5-year-olds and 77% (73-80%) and 83% (79-86%) in 6-10-year-olds. When the two age strata were combined (2-10-year-old children), MenACWY-CRM was noninferior to MCV4 for all four groups, and statistically superior for groups C, W, and Y. Safety parameters were similar across age cohorts and vaccines groups. CONCLUSIONS: MenACWY-CRM and MCV4 were immunogenic and well tolerated in children aged 2-10 years. Seroresponse to MenACWY-CRM was statistically noninferior to MCV4 for all groups, and statistically superior for groups C, W, and Y. TRIAL REGISTRATION: Clinicaltrials.gov identifier: NCT00616421.

Efficient induction of immune responses through intradermal vaccination with N-trimethyl chitosan containing antigen formulations.

The function of N-trimethyl chitosan (TMC) in dermal immunisation is unknown. Therefore we investigated the immunogenicity of both antigen-containing TMC nanoparticles and TMC/antigen solutions after intradermal injection. Nanoparticles were prepared with a size around 200 nm and a positive zetapotential. In vitro, TMC nanoparticles increased the uptake of OVA by dendritic cells (DCs) and both nanoparticles and TMC/OVA mixtures were able to induce upregulation of MHC-II, CD83 and CD86. These activated DCs could induce a Th2 biased T cell proliferation. A solution of plain OVA did not induce DC maturation or T cell proliferation. In vivo, mice were injected thrice with TMC based formulations containing either OVA or diphtheria toxoid (DT), a more relevant antigen. All TMC-containing formulations were able to increase the IgG titres compared to unadjuvanted antigen and induced a Th2 biased immune response. When using DT-containing TMC formulations, IgG titres and neutralising antibody titres could match up to those obtained after subcutaneous injection of DT-Alum. In conclusion, both soluble TMC/antigen mixtures and TMC nanoparticles are able to induce DC maturation and enhance immune responses after intradermal injection. This demonstrates that TMC functions as an immune potentiator for antigens delivered via the skin.

Analysis of in vitro lymphocyte proliferation as a screening tool for cellular immunodeficiency.

Measuring lymphocyte response to mitogens and antigens is a mainstay of screening for cellular immunodeficiency. Few reports analyze performance as a screening tool in diverse patient cohorts. We studied proliferation assays performed at Children's Hospital Boston from 1996 to 2003 using mitogens phytohemagglutinin (PHA), concanavalin A (CONA) and pokeweed mitogen, and antigens tetanus (TT) and diphtheria (DT) toxoids, and compared a subset of patients with T cell dysfunction with adult controls using receiver operating characteristic analysis. Results were correlated with clinical data. CONA was superior to PHA in identifying patients with immunodeficiency. TT was second best. Interpretation based on raw CPM, a stimulation index, or reference to simultaneous controls all performed equally. Combining data from multiple mitogens and/or antigens did not enhance performance. Proliferation testing is a useful component of screening for cellular immunodeficiency, but is not a sensitive predictor of cellular immune compromise or risk of opportunistic infection.

[Prospects of use coryneformic bacteria drug to prevent and treat respiratory infections].

This work shows the Codivac efficiency in its aerosol form to treat children sick of influenza and acute respiratory viral infections. Course of therapy with the Codivac drug leads to improve clinical picture and rapid elimination virus from human organism as compared with children undergone traditional therapy. Recovery is accompanied with improving of indices of cell immunity. The article shows the prospects of use Codivac to prevent and treat respiratory infections.

Safety and immunogenicity of two octavalent pneumococcal conjugate vaccines in American Indian infants.

We evaluated the safety and immunogenicity of two octavalent pneumococcal polysaccharide vaccines (serotypes 3, 4, 6B, 9V, 14, 18C, 19F, and 23F) conjugated to either diphtheria toxoid (PncD) or tetanus protein (PncT) among White Mountain Apache and Gila River Indian Community infants. This was a prospective, non-randomized, open label, comparative pilot study of PncD and PncT. Since this was a pilot study, a small sample size of 60 infants was enrolled. Enrolled healthy infants received either PncD or PncT at 2, 4, and 6 months of age. Antibody concentrations were measured by enzyme-immunoassay (EIA) prior to each dose and 1 month after the last dose. Local reaction rates were similar between PncD and PncT groups. The geometric mean concentrations (GMCs) were significantly higher for PncD than PncT for serotype 3 whereas GMCs were significantly higher for PncT for serotype 4. For this pilot study, both vaccines appeared to be safe and immunogenic in this group of American Indian infants.

New drugs for patients with pancreatic cancer.

This past year has proved to be a relatively disappointing one for the development of agents that could improve the survival rates of patients with advanced pancreatic cancer. A well designed randomized trial of treatment of patients with gemcitabine with or without a farnesyl transferase inhibitor (tried because pancreatic cancers have a high incidence of K- abnormalities) showed no improvement in survival rates. A definitive randomized controlled trial with a histone deacetylase inhibitor also proved negative. There are some signs of hope in that in early nonrandomized studies there are some new agents that appear to have some activity against the disease. These agents include the thymidylate synthase inhibitor capecitabine (which is possibly activated at the tumor site), the antigastrin immunogen G17DT (which is an immunization designed to neutralize the pancreatic growth factor gastrin), and the topoisomerase I inhibitor 9-nitrocamptothecin. In addition, the combination of the new agent oxaliplatin to high-dose 5FU plus leucovorin, which gave a median survival rate of 12.5 months, is also worthy of further study. Supportive care findings of interest for the patient with advanced pancreatic cancer of note include: the study in which eicosapentaenoic acid (fish oil) caused a modest weight gain (median of 1 kg), and the finding that ofloxacin plus ursodeoxycholic acid was not superior to ursodeoxycholic acid alone for the prevention or occlusion of biliary stents.

Gastrimmune-induced antigastrin-17 antibodies inhibit acid secretion in a rat fistula model.

BACKGROUND: Gastrimmune is an immunogenic form of gastrin. It raises in situ antibodies against two proliferative forms of gastrin: amidated and glycine-extended gastrin-17. It has been shown to have a therapeutic action in several in vivo tumour models. Following immunization, due to the complex equilibrium that exists between the antibodies and gastrin, it is not technically feasible to assay for free gastrin. AIM: To determine the effect of Gastrimmune-induced antigastrin antibodies on acid secretion. METHOD: A rat gastric fistula model was used. Animals (six per group) were immunized with a control immunogen or ascending doses of Gastrimmune. Acid output was measured following infusion of increasing doses of gastrin-17 and pentagastrin. RESULTS: Gastrimmune-induced antibodies significantly reduced gastrin-17-stimulated acid output compared to control animals (Gastrimmune at 200 microg/rat vs. control; acid output following 30 ng gastrin-17, 0.01 vs. 0.16, P < 0.001; following 120 ng gastrin-17, 0.022 vs. 0.29, P < 0.001). CONCLUSIONS: Gastrimmune significantly inhibits gastrin-17-stimulated acid output. This biological assay suggests that the antigastrin antibodies effectively bind gastrin-17. In addition to its use as an antineoplastic agent, Gastrimmune may have a role as an acid-decreasing agent in oesophagogastric pathology.

Immunogenicity and reactogenicity of a single dose of a diphtheria--tetanus--acellular pertussis component vaccine (DTaP) compared to a diphtheria--tetanus toxoid (Td) and a diphtheria toxoid vaccine (d) in adults.

We compared immunogenicity and reactogenicity of a single dose of DTaP vaccine (containing tetanus and diphtheria toxoids and four acellular pertussis antigens) with conventional Td- or d-vaccines in 180 German adults. Antibody values against diphtheria and tetanus toxin and against the pertussis antigens fimbriae (FIM), filamentous hemagglutinin (FHA) and pertussis toxin (PT) were measured in pre- and post-immunization sera. Reactogenicity was determined by a patient diary card. Pre-immunization antibody values against diphtheria toxin were low in all three vaccine groups. After immunization, > or = 80% of the vaccinees in all three groups were fully protected (> or = 0.1 IU/ml), but geometric mean values were significantly higher in DTaP recipients compared to Td or d recipients (1.65 vs. 0.44 and 0.48, respectively; both P < 0.05). Pre-immunization antibody values against tetanus toxin were high in all three groups, and after immunization 100% of the vaccinees were protected (> or = 0.1 IU/ml). Furthermore, substantial antibody responses against pertussis antigens were elicited in DTaP recipients with geometric mean rises of 22.5, 4.1 and 7.5 for antibodies against FHA, fimbriae and PT, respectively. All three vaccines were well tolerated. Frequency and severity of local reactions were similar between DTaP and Td recipients and even less common in d recipients. Since DTaP did provide a significant boost of anti-pertussis antibodies and a significantly higher anti-diphtheria response than conventional Td vaccine without an increase of side effects, it might be an appropriate candidate for use in adults.

CD40 stimulation promotes human secondary immunoglobulin responses in HuPBL-SCID chimeras.

Antibodies to CD40 have been demonstrated to promote B-cell growth and differentiation in vitro. In order to determine if CD40 stimulation could promote antigen-specific human immunoglobulin (Ig) production in vivo, we examined the effects of anti-human CD40 MoAb in an in vivo system where human peripheral blood lymphocytes (huPBL) were engrafted into mice with severe combined immune deficiency (SCID). The huPBL-SCID mice were then given various doses of diphtheria-tetanus toxoid (DT) vaccine and were examined for the presence of human DT-specific antibodies by ELISA. Surprisingly, treatment with anti-CD40 significantly lowered background DT responses versus untreated chimeras in unimmunized huPBL-SCID mice. However, after immunization, huPBL-SCID mice treated with anti-CD40 MoAb responded to a significantly greater extent in response to the vaccine compared with control huPBL-SCID mice, although total Ig levels were sometimes lower in anti-CD40-treated mice. The predominant Ig isotype induced after immunization was IgG. Thus, CD40 stimulation promotes human secondary IgG responses in huPBL-SCID mice. These data demonstrate that CD40 stimulation is capable of promoting antigen-specific human B-cell responses in vivo.

Monokine production following in vitro stimulation of the THP-1 human monocytic cell line with pertussis vaccine components.

Whole-cell pertussis found in diphtheria-tetanus-pertussis (DTP) vaccine can produce symptoms reminiscent of biological responses to circulating proinflammatory monokines such as IL-6, IL-1beta, and TNFalpha. Therefore the ability of pertussis-containing vaccines and several heat-killed Bordetella pertussis preparations to stimulate cytokine production in a human monocytic cell line, THP-1, were examined. The whole-cell pertussis vaccine induced significantly more IL-6, IL-1beta, and TNFalpha production than did the acellular pertussis or diphtheria-tetanus-only vaccine. Polymyxin B was able to inhibit most of the IL-6 induced by pertussis endotoxin and a heat-killed preparation of B. pertussis containing a null mutation in bvgAS, a regulatory locus required for expression of all known protein virulence factors synthesized by this organism. However, it only partially inhibited IL-6 production induced by other pertussis-containing preparations, including DTP vaccine. These results indicate that in vitro whole-cell vaccine is a potent stimulator of IL-6, IL-1beta, and TNFalpha. They also suggest that although endotoxin is a major inducer of IL-6, other components of B. pertussis also contribute to IL-6 production by monocytes.