Neonatal BCG Immunization Followed by DNAhsp65 Boosters: Highly Immunogenic but not Protective Against Tuberculosis - a Paradoxical Effect of the Vector?

A new tuberculosis vaccine is urgently needed. Prime-boost strategies are considered very promising and the inclusion of BCG is highly desirable. In this investigation, we tested the protective efficacy of BCG delivered in the neonatal period followed by boosters in the adult phase with a DNA vaccine containing the hsp65 gene from Mycobacterium leprae (pVAXhsp65). Immune responses were characterized by serum anti-hsp65 antibody levels and IFN-gamma and IL-5 production by the spleen. Amounts of these cytokines were also determined in lung homogenates. Protective efficacy was established by the number of colony-forming units (CFU) and histopathological analysis of the lungs after challenge with Mycobacterium tuberculosis. Immunization with BCG alone triggered a significant reduction of CFU in the lungs and also clearly preserved the pulmonary parenchyma. BCG priming also increased the immunogenicity of pVAXhsp65. However, boosters with pVAXhsp65 or the empty vector abolished the protective efficacy of BCG. Also, higher IL-5 levels were produced by spleen and lungs after DNA boosters. These results demonstrated that neonatal BCG immunization followed by DNAhsp65 boosters is highly immunogenic but is not protective against tuberculosis.

Characterization and in vitro activities of cell-free antigens from Histoplasma capsulatum-loaded biodegradable microspheres

In the last decades, the incidence of histoplasmosis, a pulmonary fungal disease caused by Histoplasma capsulatum, has increased worldwide. In this context, vaccines for the prevention of this infection or therapies are necessary. Cell-free antigens (CFAgs) from H. capsulatum when administered for murine immunization purposes are able to confer protection and control of the infection, since they activate cellular immunity. However the most of vaccination procedures need several anti, gens administrations and immunoadjuvants, which are not approved for use in humans. The aim of this study was to develop and characterize a vaccination approach using biodegradable PLGA microspheres (MS) that could allow the controlled and/or sustained release of the encapsulated antigens from H. capsulatum. CFAgs-loaded MS presented a size less than 10 mu m, were marked engulfed by bone marrow-derived macrophages (BMDM phi) and induced the nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) production by these cells. Our data show that CFAgs-loaded MS induce cell activation, suggesting an immunostimulant effect to be further investigated during immunization procedures. CFAgs-loaded MS present potential to be used as vaccine in order to confer protection against H. capsulatum infection. (C) 2009 Elsevier B.V. All rights reserved.

Protective efficacy of different strategies employing Mycobacterium leprae heat-shock protein 65 against tuberculosis

Background: Tuberculosis is a major threat to human health. The high disease burden remains unaffected and the appearance of extremely drug-resistant strains in different parts of the world argues in favor of the urgent need for a new effective vaccine. One of the promising candidates is heat-shock protein 65 when used as a genetic vaccine (DNAhsp65). Nonetheless, there are substantial data indicating that BCG, the only available anti-TB vaccine for clinical use, provides other important beneficial effects in immunized infants. Methods: We compared the protective efficacy of BCG and Hsp65 antigens in mice using different strategies: i) BCG, single dose subcutaneously; ii) naked DNAhsp65, four doses, intramuscularly; iii) liposomes containing DNAhsp65, single dose, intranasally; iv) microspheres containing DNAhsp65 or rHsp65, single dose, intramuscularly; and v) prime-boost with subcutaneous BCG and intramuscular DNAhsp65. Results: All the immunization protocols were able to protect mice against infection, with special benefits provided by DNAhsp65 in liposomes and prime-boost strategies. Conclusion: Among the immunization protocols tested, liposomes containing DNAhsp65 represent the most promising strategy for the development of a new anti-TB vaccine.

HMGB1-derived peptide acts as adjuvant inducing immune responses to peptide and protein antigen

There is a need for new adjuvants that will induce immune responses to subunit vaccines. We show that a short peptide, named Hp91, whose sequence corresponds to an area within the endogenous molecule high mobility group box (HMGB1) protein 1 potentiates cellular immune responses to peptide antigen and cellular and humoral immune responses to protein antigen in vivo. Hp91 promoted the in vivo production of the immunomodulatory cytokines, IFN-gamma, TNF-alpha, IL-6, and IL-12 (p70), as well as antigen-specific activation of CD8+ T cells. These results demonstrate the ability of a short immunostimulatory peptide to serve as an adjuvant for subunit vaccines. (C) 2010 Elsevier Ltd. All rights reserved.

A Dynamical Modeling to Study the Adaptive Immune System and the Influence of Antibodies in the Immune Memory

Immunological systems have been an abundant inspiration to contemporary computer scientists. Problem solving strategies, stemming from known immune system phenomena, have been successfully applied to chall enging problems of modem computing. Simulation systems and mathematical modeling are also beginning use to answer more complex immunological questions as immune memory process and duration of vaccines, where the regulation mechanisms are not still known sufficiently (Lundegaard, Lund, Kesmir, Brunak, Nielsen, 2007). In this article we studied in machina a approach to simulate the process of antigenic mutation and its implications for the process of memory. Our results have suggested that the durability of the immune memory is affected by the process of antigenic mutation.and by populations of soluble antibodies in the blood. The results also strongly suggest that the decrease of the production of antibodies favors the global maintenance of immune memory.

Evaluation of the vaccination status in pediatric renal transplant recipients

To assess the immunization status of pediatric renal transplant patients followed at a single center in Brazil, vaccination charts of all patients aged between one and 18 yr were analyzed both pre- and post-transplantation. Appropriate immunization was defined according to the National Immunization Program (routine vaccines) - for all Brazilian children - and the Special Immunobiological Agents Program that also includes special vaccines for immunodeficient or other high-risk children. A total of 46 patients was evaluated (mean age 13.7 yr; range 4-17 yr). Vaccination charts were found to be up to date in only two patients (4.3%) pretransplant and in two (4.3%) post-transplant. Although 36 patients (62.2%) in the pretransplant phase and 24 (52.1%) in the post-transplant phase had been vaccinated according to the National Immunization Program, they had not received the special vaccines indicated for their immunocompromised condition. Therefore, despite being followed at a referral center, almost all patients presented an incomplete immunization status pre- and post-transplant. This probably reflects missed opportunities and medical/parental apprehension related to vaccination of patients with chronic renal insufficiency, dialysis or kidney transplantation. Efforts should be made to ensure adequate vaccination in children with kidney diseases, especially before kidney transplantation.

Rubella Vaccination of Unknowingly Pregnant Women: The Sao Paulo Experience, 2001

Background. Rubella vaccination is contraindicated during pregnancy. During mass immunization of women of childbearing age against rubella, women unknowingly pregnant may be vaccinated. To evaluate the effects of rubella vaccination during pregnancy, the Brazilian state of Sao Paulo conducted a follow-up study of pregnant women vaccinated during a rubella campaign in 2001. Methods. Women vaccinated during pregnancy were reported to a national surveillance system. In the state of Sao Paulo, follow-up of vaccinated women included household interviews. Serum samples from vaccinated women were tested for antirubella antibodies to classify susceptibility to rubella infection. Children born to susceptible mothers were tested for evidence of congenital rubella infection and evaluated for signs of congenital rubella syndrome. Results. The Sao Paulo State Health Department received 6473 notifications of women vaccinated during pregnancy. Serology performed for 5580 women identified 811 (15%) that were previously susceptible. Incidence of spontaneous abortion or stillbirth among previously susceptible vaccinated women was similar to women with prior immunity. Twenty-seven (4.7%) of 580 newborns tested had evidence of congenital rubella infection; none had congenital rubella syndrome. Conclusions. Mass rubella vaccination of women of childbearing age was not associated with adverse birth outcomes or congenital rubella syndrome among children born to women vaccinated during pregnancy.

Epidemiology of meningococcal disease in Latin America: current situation and opportunities for prevention

Objective: Meningococcal disease continues to be a serious public health concern, being associated with high morbidity and mortality rates in many countries from Latin America. In addition to discussing recent changes in the epidemiology of meningococcal disease in the region, we also analyse the development and potential impact of new vaccines on the prevention of meningococcal disease. Methods: MEDLINE, SciELO, LILACS and websites of the national Ministries of Health databases were searched using the terms meningococcal disease, meningococcal epidemiology, Neisseria meningitidis, meningococcal vaccines and the name of Latin America countries, from 1998 to 2008, with emphasis on review articles, clinical trials and epidemiological studies. Results: Epidemiology of meningococcal disease in Latin America is characterized by marked differences from country to country. The overall incidence of meningococcal disease per year varied from less than 0.1 cases per 100,000 inhabitants in countries like Mexico to two cases per 100,000 inhabitants in Brazil. The highest age-specific incidence of meningococcal disease occurred in infants less than 1 year of age. Serogroups B and C were responsible for the majority of cases reported, but the emergence of serogroups W135 and Y was reported in some countries. Serogroup A disease is now rare in Latin America. Discussion: Although a few countries have established meningitis surveillance programs, the information is not uniform, and the quality of the reported data is poor in the majority of the region. The availability of new effective meningococcal conjugate vaccines and promising protein-based vaccine candidates against meningococcus B highlights the importance of a better understanding of the true burden of meningococcal disease in Latin America and also the need for cost-effectiveness studies before incorporating the new meningococcal vaccines to national immunization programs.

Clinical and Immunological Insights on Severe, Adverse Neurotropic and Viscerotropic Disease following 17D Yellow Fever Vaccination

Yellow fever (YF) vaccines (17D-204 and 17DD) are well tolerated and cause very low rates of severe adverse events (YEL-SAE), such as serious allergic reactions, neurotropic adverse diseases (YEL-AND), and viscerotropic diseases (YEL-AVD). Viral and host factors have been postulated to explain the basis of YEL-SAE. However, the mechanisms underlying the occurrence of YEL-SAE remain unknown. The present report provides a detailed immunological analysis of a 23-year-old female patient. The patient developed a suspected case of severe YEL-AVD with encephalitis, as well as with pancreatitis and myositis, following receipt of a 17D-204 YF vaccination. The patient exhibited a decreased level of expression of Fc-gamma R in monocytes (CD16, CD32, and CD64), along with increased levels of NK T cells (an increased CD3(+) CD16(+/-) CD56(+/-)/CD3(+) ratio), activated T cells (CD4(+) and CD8(+) cells), and B lymphocytes. Enhanced levels of plasmatic cytokines (interleukin-6 [IL-6], IL-17, IL-4, IL-5, and IL-10) as well as an exacerbated ex vivo intracytoplasmic cytokine pattern, mainly observed within NK cells (gamma interferon positive [IFN-gamma(+)], tumor necrosis factor alpha positive [TNF-alpha(+)], and IL-4 positive [IL-4(+)]), CD8(+) T cells (IL-4(+) and IL-5(+)), and B lymphocytes (TNF-alpha(+), IL-4(+), and IL-10(+)). The analysis of CD4(+) T cells revealed a complex profile that consisted of an increased frequency of IL-12(+) and IFN-gamma(+) cells and a decreased percentage of TNF-alpha(+), IL-4(+), and IL-5+ cells. Depressed cytokine synthesis was observed in monocytes (TNF-alpha(+)) following the provision of antigenic stimuli in vitro. These results support the hypothesis that a strong adaptive response and abnormalities in the innate immune system may be involved in the establishment of YEL-AND and YEL-AVD.

NO EVIDENCE OF PATHOLOGICAL AUTOIMMUNITY FOLLOWING MYCOBACTERIUM LEPRAE HEAT-SHOCK PROTEIN 65-DNA VACCINATION IN MICE

Heat-shock proteins (HSPs) are currently one of the most promising targets for the development of immunotherapy against tumours and autoimmune disorders. This protein family has the capacity to activate or modulate the function of different immune system cells. They induce the activation of monocytes, macrophages and dendritic cells, and contribute to cross-priming, an important mechanism of presentation of exogenous antigen in the context of MHC class I molecules, These various immunological properties of HSP have encouraged their use in several clinical trials. Nevertheless, an important issue regarding these proteins is whether the high homology among HSPs across different species may trigger the breakdown of immune tolerance and induce autoimmune diseases. We have developed a DNA vaccine codifying the Mycobacterium leprae Hsp65 (DNAhsp65), which showed to be highly immunogenic and protective against experimental tuberculosis. Here, we address the question of whether DNAhsp65 immunization could induce pathological autoimmunity in mice. Our results show that DNAhsp65 vaccination induced antibodies that can recognize the human Hsp60 but did not induce harmful effects in 16 different organs analysed by histopathology up to 210 days after vaccination. We also showed that anti-DNA antibodies were not elicited after DNA vaccination. The results are important for the development of both HSP and DNA-based immunomodulatory agents.

Current developments on Schistosoma proteomics

Several papers on Schistosoma mansoni and Schistosoma japonicum proteomes have been published worldwide in the past few years, representing an emerging field of study. Knowledge of the schistosome proteome may greatly enhance our understanding of schistosome physiological processes at the molecular level, and may provide new models for development of vaccines or drugs. Despite the importance of this approach. schistosome proteomic research in Brazil is still incipient. Here we review the development of schistosome proteomic research around the world and provide an appreciation on the future opportunities in Brazil. (c) 2008 Elsevier B.V. All rights reserved

Cytokine Signatures of Innate and Adaptive Immunity in 17DD Yellow Fever Vaccinated Children and Its Association With the Level of Neutralizing Antibody

Background. The live attenuated yellow fever (YF) vaccines have been available for decades and are considered highly effective and one of the safest vaccines worldwide. Methods. The impact of YF-17DD-antigens recall on cytokine profiles of YF-17DD-vaccinated children were characterized using short-term cultures of whole blood samples and single-cell flow cytometry. This study enrolled seroconverters and nonseroconverters after primovaccination (PV-PRNT(+) and PV-PRNT(-)), seroconverters after revaccination (RV-PRNT(+)), and unvaccinated volunteers (UV-PRNT(-)). Results. The analysis demonstrated in the PV-PRNT(+) group a balanced involvement of pro-inflammatory/regulatory adaptive immunity with a prominent participation of innate immunity pro-inflammatory events (IL-12(+) and TNF-alpha(+) NEU and MON). Using the PV-PRNT(+) cytokine signature as a reference profile, PV-PRNT(+) presented a striking lack of innate immunity proinflammatory response along with an increased adaptive regulatory profile (IL-4(+) CD4(+) T cells and IL-10(+) and IL-5(+) CD8(+) T cells). Conversely, the RV-PRNT(+) shifted the overall cytokine signatures toward an innate immunity pro-inflammatory profile and restored the adaptive regulatory response. Conclusions. The data demonstrated that the overall cytokine signature was associated with the levels of PRNT antibodies with a balanced innate/adaptive immunity with proinflammatory/regulatory profile as the hallmark of PV-PRNT(MEDIUM+), whereas a polarized regulatory response was observed in PV-PRNT(-) and a prominent proinflammatory signature was the characteristic of PV-PRNT(HIGH+).

Expression and characterization of HPV-16 L1 capsid protein in Pichia pastoris

Human papillomaviruses (HPVs) are responsible for the most common human sexually transmitted viral infections. Infection with high-risk HPVs, particularly HPV16, is associated with the development of cervical cancer. The papillomavirus L1 major capsid protein, the basis of the currently marketed vaccines, self-assembles into virus-like particles (VLPs). Here, we describe the expression, purification and characterization of recombinant HPV16 L1 produced by a methylotrophic yeast. A codon-optimized HPV16 L1 gene was cloned into a non-integrative expression vector under the regulation of a methanol-inducible promoter and used to transform competent Pichia pastoris cells. Purification of L1 protein from yeast extracts was performed using heparin-sepharose chromatography, followed by a disassembly/reassembly step. VLPs could be assembled from the purified L1 protein, as demonstrated by electron microscopy. The display of conformational epitopes on the VLPs surface was confirmed by hemagglutination and hemagglutination inhibition assays and by immuno-electron microscopy. This study has implications for the development of an alternative platform for the production of a papillomavirus vaccine that could be provided by public health programs, especially in resource-poor areas, where there is a great demand for low-cost vaccines.

Feline injection-site sarcoma

The feline injection-site sarcoma (FIS) is a challenge for the veterinarian and the affected cat`s owner. The injectable applications (vaccines, medications) seems to be the reason for that neoplasia, more specifically, the inflammation caused by injury of given drugs or antigens to the health tissue. Generally the FIS presents a more aggressive behavior when compared to sarcoma not associated to application. The most effective treatment his not been established yet, but it is believed that a multimodality of therapies, surgery, radiotherapy, and chemotherapy would be the most indicated option. The knowledge of the illness in all of its aspects will Supply to professionals colleges subsidies in relation to the best way to approach its diagnosis and treatment.

Molecular Epidemiology of Avian Infectious Bronchitis in Brazil from 2007 to 2008 in Breeders, Broilers, and Layers

Multiple lineages of Brazilian strains from 2007 to 2008 of avian infectious bronchitis virus (IBV) were detected in flocks of breeders, broilers, and layers. Organs samples from 20 IBV-positive flocks with variable clinical signs were submitted to the partial amplification of S gene (nucleotides 726-1071) of IBV. Fifteen of the 20 sequenced strains segregated in a unique Brazilian cluster subdivided in three subclusters (Brazil 01, 02, and 03). Whereas three strains could be classified as Massachusetts (Mass) genotype, the remaining two strains, originating from flocks with reproductive and respiratory disorders, grouped within the 4/91-793B genotype, a genotype that has not been detected before in Brazil. The potential relevance of the findings to the poultry industry is discussed because the low level of identity of the sequenced part of the S gene from 17 of 20 detected field strains and the vaccines of the Massachusetts serotype used suggest that the level of cross-protection by the Massachusetts vaccines might be low.