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Increasing the effectiveness of vaccines against respiratory viruses is particularly relevant for the elderly, since they are prone to develop serious infections due to comorbidities and the senescence of the immune system. The addition of saponin-based adjuvants is an interesting strategy to increase the effectiveness of vaccines. We have previously shown that ISCOM matrices from Q. brasiliensis (IMXQB) are a safe and potent adjuvant. In this study, we evaluated the use of IMXQB as an adjuvant for the seasonal trivalent influenza vaccine (TIV) in an aged mice model. Herein, we show that subcutaneous injection of the adjuvanted vaccine promoted higher titers of IgM, IgG (and isotypes), and serum hemagglutination inhibition titers (HAI). Notably, aged mice immunized by intranasal route also produced higher IgG (and isotypes) and IgA titers up to 120 days after priming, as well as demonstrating an improvement in the HAI antibodies against the TIV. Further, experimental infected aged mice treated once with sera from adult naïve mice previously immunized with TIV-IMXQB subcutaneously successfully controlled the infection. Overall, TIV-IMXQB improved the immunogenicity compared to TIV by enhancing systemic and mucosal immunity in old mice conferring a faster recovery after the H1N1pdm09-like virus challenge. Thus, IMXQB nanoparticles may be a promising platform for next-generation viral vaccines.
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Salmonella enterica serovar Derby causes foodborne disease (FBD) outbreaks worldwide, mainly from contaminated pork but also from chickens. During a major epidemic of FBD in Uruguay due to S. enteritidis from poultry, we conducted a large survey of commercially available eggs, where we isolated many S. enteritidis strains but surprisingly also a much larger number (ratio 5:1) of S. Derby strains. No single case of S. Derby infection was detected in that period, suggesting that the S. Derby egg strains were impaired for human infection. We sequenced fourteen of these egg isolates, as well as fifteen isolates from pork or human infection that were isolated in Uruguay before and after that period, and all sequenced strains had the same sequence type (ST40). Phylogenomic analysis was conducted using more than 3,500 genomes from the same sequence type (ST), revealing that Uruguayan isolates clustered into four distantly related lineages. Population structure analysis (BAPS) suggested the division of the analyzed genomes into nine different BAPS1 groups, with Uruguayan strains clustering within four of them. All egg isolates clustered together as a monophyletic group and showed differences in gene content with the strains in the other clusters. Differences included variations in the composition of mobile genetic elements, such as plasmids, insertion sequences, transposons, and phages, between egg isolates and human/pork isolates. Egg isolates showed an acid susceptibility phenotype, reduced ability to reach the intestine after oral inoculation of mice, and reduced induction of SPI-2 ssaG gene, compared to human isolates from other monophyletic groups. Mice challenge experiments showed that mice infected intraperitoneally with human/pork isolates died between 1-7 days p.i., while all animals infected with the egg strain survived the challenge. Altogether, our results suggest that loss of genes functions, the insertion of phages and the absence of plasmids in egg isolates may explain why these S. Derby were not capable of producing human infection despite being at that time, the main serovar recovered from eggs countrywide.
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Colistin remains one of the last-resort therapies for combating infections caused by multidrug-resistant (MDR) Enterobacterales, despite its adverse nephro- and neuro-toxic effects. This study elucidates the mechanism of action of a non-antibiotic 4-anilinoquinazoline-based compound that synergistically enhances the effectiveness of colistin against Salmonella enterica. The quinazoline sensitizes Salmonella by deactivating intrinsic, mutational, and transferable resistance mechanisms that enable Salmonella to counteract the antibiotic impact colistin, together with an induced disruption to the electrochemical balance of the bacterial membrane. The attenuation of colistin resistance via the combined treatment approach also proves efficacious against E. coli, Klebsiella, and Acinetobacter strains. The dual therapy reduces the mortality of Galleria mellonella larvae undergoing a systemic Salmonella infection when compared to individual drug treatments. Overall, our findings unveil the potential of the quinazoline-colistin combined therapy as an innovative strategy against MDR bacteria.
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Mariposas , Infecções por Salmonella , Animais , Colistina/farmacologia , Colistina/uso terapêutico , Escherichia coli , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Farmacorresistência Bacteriana Múltipla , Infecções por Salmonella/tratamento farmacológico , Testes de Sensibilidade MicrobianaRESUMO
Innate immune cells can undergo long-term functional reprogramming after certain infections, a process called trained immunity (TI). Here, we focus on antigens of Leishmania braziliensis, which induced anti-tumor effects via trained immunity in human monocytes. We reveal that monocytes exposed to promastigote antigens of L. braziliensis develop an enhanced response to subsequent exposure to Toll-like receptor (TLR)2 or TLR4 ligands. Mechanistically, the induction of TI in monocytes by L. braziliensis is mediated by multiple pattern recognition receptors, changes in metabolism, and increased deposition of H3K4me3 at the promoter regions of immune genes. The administration of L. braziliensis exerts potent anti-tumor capabilities by delaying tumor growth and prolonging survival of mice with non-Hodgkin lymphoma. Our work reveals mechanisms of TI induced by L. braziliensis in vitro and identifies its potential for cancer immunotherapy.
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Leishmania braziliensis , Leishmaniose Cutânea , Neoplasias , Humanos , Camundongos , Animais , MonócitosRESUMO
The threat of viral influenza infections has sparked research efforts to develop vaccines that can induce broadly protective immunity with safe adjuvants that trigger robust immune responses. Here, we demonstrate that subcutaneous or intranasal delivery of a seasonal trivalent influenza vaccine (TIV) adjuvanted with the Quillaja brasiliensis saponin-based nanoparticle (IMXQB) increases the potency of TIV. The adjuvanted vaccine (TIV-IMXQB) elicited high levels of IgG2a and IgG1 antibodies with virus-neutralizing capacity and improved serum hemagglutination inhibition titers. The cellular immune response induced by TIV-IMXQB suggests the presence of a mixed Th1/Th2 cytokine profile, antibody-secreting cells (ASCs) skewed toward an IgG2a phenotype, a positive delayed-type hypersensitivity (DTH) response, and effector CD4+ and CD8+ T cells. After challenge, viral titers in the lungs were significantly lower in animals receiving TIV-IMXQB than in those inoculated with TIV alone. Most notably, mice vaccinated intranasally with TIV-IMXQB and challenged with a lethal dose of influenza virus were fully protected against weight loss and lung virus replication, with no mortality, whereas, among animals vaccinated with TIV alone, the mortality rate was 75%. These findings demonstrate that TIV-IMXQB improved the immune responses to TIV, and, unlike the commercial vaccine, conferred full protection against influenza challenge.
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Vacinas contra Influenza , Influenza Humana , Nanopartículas , Animais , Camundongos , Humanos , Influenza Humana/prevenção & controle , Quillaja , Linfócitos T CD8-Positivos , Adjuvantes Imunológicos , Adjuvantes Farmacêuticos , Saponinas de Quilaia , Imunoglobulina GRESUMO
Localized melanoma is easy to remove by surgery, resulting in a high five-year relative survival rate. However, when disseminated the disease management is challenging. The use of immunotherapies, such as anti-checkpoint monoclonal antibodies, has improved treatment options but still only a small percentage of patients responds to these expensive treatments. In this work, we apply a bacteria-based immunotherapy using LVR01, an attenuated Salmonella enterica serovar Typhimurium, as neoadjuvant therapy one week before surgery in a preclinical disseminated murine melanoma model. LVR01 administration resulted in tumor growth retardation prior to tumor resection, due to a rapid upregulation of inflammatory genes in the tumor microenvironment. As a consequence, cell infiltration increased, particularly neutrophils, macrophages and NK cells, being the latter involved in Salmonella anti-tumor activity. Besides, tumor-draining lymph node infiltration is characterized by reinvigorated CD4+ and CD8+ lymphocytes. Induced immune response could account for the prevention or delay of tumor recurrence and appearance of metastasis, resulting in a prolonged overall survival after surgery. Furthermore, upon rechallenge mice show partial protection, suggesting the existence of specific memory against melanoma. We propose that neoadjuvant LVR01 treatment could represent an interesting inexpensive alternative that may ease tumor resection, while preventing tumor recurrence in patients with melanoma.
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Salmonella-based cancer therapies show great potential in preclinical models, but for most cases the observed antitumor effect is transient. Understanding the basis of the antitumor efficacy might guide the design of improved strains that elicit long-lasting effects, paving the wave for clinical use. Here, we deepened into the role of macrophages and inflammasome activation in the context of Salmonella anti-melanoma effect. We showed inflammasome activation in melanoma cells upon infection, which correlated with cell surface exposure of gasdermin-D (GSDM-D) and calreticulin (CRT) and High mobility group box 1 protein (HMGB-1) release, suggesting immunogenic cell death, particularly pyroptosis. Salmonella infection upregulated levels of Caspase-11 (Casp11) mRNA, but not Nlrp3 or Nlrc4 mRNA, the only described inflammasome receptors engaged by Salmonella, suggesting that non-canonical inflammasome activation could be occurring in melanoma cells. Intratumoral administration of Salmonella to melanoma-bearing mice elicited local inflammasome activation and interleukin-1ß (IL-1ß) production together with tumor growth retardation and extended survival in wild type but not Caspase-1/11 (Casp1/11) knockout mice despite similar levels of intratumoral IL-1ß in the later. Salmonella antitumor activity was also suppressed in melanoma bearing Nlrp3 knockout mice. Salmonella induced macrophage recruitment to the tumor site and infiltrating cells exhibited inflammasome activation. Depletion experiments confirmed that macrophages are also essential for Salmonella anti-melanoma effect. Intratumoral macrophages showed a marked M2/M1 shift soon after treatment but this inflammatory profile is then lost, which could explain the transient effect of therapy. All in all, our results highlight CASP-1/11 axis and macrophages as essential players in Salmonella-based cancer immunotherapy and suggest a possible target for future interventions.
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Inflamassomos , Macrófagos , Neoplasias , Salmonella , Animais , Caspase 1/metabolismo , Inflamassomos/imunologia , Interleucina-1beta/metabolismo , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Neoplasias/imunologia , Neoplasias/terapia , RNA Mensageiro/metabolismo , Microambiente TumoralRESUMO
Treatment of malignant melanoma has improved in the last few years owing to early detection and new therapeutic options. Still, management of advanced disease remains a challenge because it requires systemic treatment. In such cases, dacarbazine-based chemotherapy has been widely used, despite low efficacy. Neoadjuvant therapies emerge as alternative options that could help chemotherapy to achieve increased benefit. In this work, we evaluate LVR01, an attenuated Salmonella enterica serovar typhimurium, as neoadjuvant intralesional therapy in combination with dacarbazine in a preclinical melanoma model. B16F1 melanomaâbearing mice received intraperitoneal administration of dacarbazine for 3 consecutive days. LVR01 treatment, consisting of one single intratumoral injection, was applied 1 day before chemotherapy began. This therapeutic approach retarded tumor growth and prolonged overall survival, revealing a strong synergistic antitumor effect. Dacarbazine induced a drastic reduction of secondary lymphoid organ cellularity, which was partially restored by Salmonella, particularly potentiating activated cytotoxic cell compartments. Systemic immune reactivation could be a consequence of the intense inflammatory tumor microenvironment induced by LVR01. We propose that the use of LVR01 as neoadjuvant intralesional therapy could be considered as an interesting strategy with close clinical application to boost chemotherapy effect in patients with melanoma.
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Melanoma , Neoplasias Cutâneas , Animais , Dacarbazina/uso terapêutico , Humanos , Melanoma/patologia , Camundongos , Terapia Neoadjuvante , Salmonella typhimurium , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Microambiente TumoralRESUMO
Polyvalent bacterial lysates have been in use for decades for prevention and treatment of respiratory infections with reported clinical benefits. However, besides claims of broad immune activation, the mode of action is still a matter of debate. The lysates, formulated with the main bacterial species involved in respiratory infections, are commonly prepared by chemical or mechanical disruption of bacterial cells, what is believed influences the biological activity of the product. Here, we prepared two polyvalent lysates with the same composition but different method of bacterial cell disruption and evaluated their biological activity in a comparative fashion. We found that both bacterial lysates induce NF-kB activation in a MyD88 dependent manner, suggesting they work as TLR agonists. Further, we found that a single intranasal dose of any of the two lysates, is sufficient to protect against pneumococcal pneumonia, suggesting that they exert similar biological activity. We have previously shown that protection against pneumococcal pneumonia can also be induced by prior S. pneumoniae sub lethal infection or therapeutic treatment with a TLR5 agonist. Protection in those cases depends on neutrophil recruitment to the lungs, and can be associated with increased local expression of IL-17A. Here, we show that bacterial lysates exert protection against pneumococcal pneumonia independently of neutrophils, IL-17A or Caspase-1/11 activation, suggesting the existence of redundant mechanisms of protection. Trypsin-treated lysates afford protection to the same extent, suggesting that just small peptides suffice to exert the protective effect or that the molecules responsible for the protective effect are not proteins. Understanding the mechanism of action of bacterial lysates and deciphering the active components shall allow redesigning them with more precisely defined formulations and expanding their range of action.
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Bactérias/química , Fatores Biológicos/farmacologia , Caspase 1/metabolismo , Interleucina-17/metabolismo , Neutrófilos/metabolismo , Pneumonia Pneumocócica/prevenção & controle , Streptococcus pneumoniae/efeitos dos fármacos , Células A549 , Animais , Fatores Biológicos/química , Ativação Enzimática , Humanos , Camundongos , Pneumonia Pneumocócica/metabolismo , Pneumonia Pneumocócica/microbiologia , Substâncias Protetoras/química , Substâncias Protetoras/farmacologia , Streptococcus pneumoniae/fisiologia , Análise de Sobrevida , Células THP-1RESUMO
Salmonella comprises two species and more than 2500 serovars with marked differences in host specificity, and is responsible for a wide spectrum of diseases, ranging from localized gastroenteritis to severe life-threatening invasive disease. The initiation of the host inflammatory response, triggered by many Pathogen-Associated Molecular Patterns (PAMPs) that Salmonella possesses, recruits innate immune cells in order to restrain the infection at the local site. Neutrophils are known for killing bacteria through oxidative burst, amid other mechanisms. Amongst those mechanisms for controlling bacteria, the release of Extracellular Traps (ETs) represents a newly described pathway of programmed cell death known as ETosis. Particularly, Neutrophil Extracellular Traps (NETs) were first described in 2004 and since then, a number of reports have demonstrated their role as a novel defense mechanism against different pathogens. This released net-like material is composed of cellular DNA decorated with histones and cellular proteins. These structures have shown ability to trap, neutralize and kill different kinds of microorganisms, ranging from viruses and bacteria to fungi and parasites. Salmonella was one of the first microorganisms that were reported to be killed by NETs and several studies have confirmed the observation and deepened into its variants. Nevertheless, much less is known about their counterparts in other immune cells, e.g. Macrophage Extracellular Traps (METs) and Salmonella-induced MET release has never been reported so far. In this work, we observed the production of METs induced by Salmonella enterica serovar Typhimurium and recorded their effect on bacteria, showing for the first time that macrophages can also release extracellular DNA traps upon encounter with Salmonella Typhimurium. Additionally we show that METs effectively immobilize and reduce Salmonella survival in a few minutes, suggesting METs as a novel immune-mediated defense mechanism against Salmonella infection. Of note, this phenomenon was confirmed in primary macrophages, since MET release was also observed in bone marrow-derived macrophages infected with Salmonella. The evidence of this peculiar mechanism provides new incipient insights into macrophages´ role against Salmonella infection and can help to design new strategies for the clinical control of this transcendental pathogen.
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Armadilhas Extracelulares , Animais , Armadilhas Extracelulares/metabolismo , Macrófagos , Camundongos , Neutrófilos , Explosão Respiratória , Salmonella typhimuriumRESUMO
Bacterial lysates, prepared from the microorganisms most frequently involved in human Respiratory Tract Infections (RTIs) have been in the market for several decades, and at present, several different brands are available in many countries worldwide. They all claimed to exert local and systemic immunomodulatory effects but different clinical trials show disparate results between them. The lack of consistency of predicted therapeutic effects has undermined their clinical use and hampered licensing in several countries. One explanation for such lack of consistency in the results is that their methods of preparation are also very different. Here, we review the available literature describing methods of preparation of bacterial lysates, including patent disclosure documents. We found a great variety of methodologies of preparation and a lack of standardized procedures among them. The main conclusion of our study is that there is a clear need for standardized protocols of production to obtain comparable results in clinical trials worldwide.
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An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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The Salmonella enterica PhoP/PhoQ two-component signaling system coordinates the spatiotemporal expression of key virulence factors that confer pathogenic traits. Through biochemical and structural analyses, we found that the sensor histidine kinase PhoQ acted as a receptor for long-chain unsaturated fatty acids (LCUFAs), which induced a conformational change in the periplasmic domain of the PhoQ protein. This resulted in the repression of PhoQ autokinase activity, leading to inhibition of the expression of PhoP/PhoQ-dependent genes. Recognition of the LCUFA linoleic acid (LA) by PhoQ was not stereospecific because positional and geometrical isomers of LA equally inhibited PhoQ autophosphorylation, which was conserved in multiple S. enterica serovars. Because orally acquired Salmonella encounters conjugated LA (CLA), a product of the metabolic conversion of LA by microbiota, in the human intestine, we tested how short-term oral administration of CLA affected gut colonization and systemic dissemination in a mouse model of Salmonella-induced colitis. Compared to untreated mice, CLA-treated mice showed increased gut colonization by wild-type Salmonella, as well as increased dissemination to the spleen. In contrast, the inability of the phoP strain to disseminate systemically remained unchanged by CLA treatment. Together, our results reveal that, by inhibiting PhoQ, environmental LCUFAs fine-tune the fate of Salmonella during infection. These findings may aid in the design of new anti-Salmonella therapies.
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Proteínas de Bactérias/metabolismo , Histidina Quinase/metabolismo , Ácido Linoleico/metabolismo , Infecções por Salmonella/metabolismo , Salmonella typhimurium/metabolismo , Transdução de Sinais , Animais , Proteínas de Bactérias/genética , Feminino , Histidina Quinase/genética , Ácido Linoleico/genética , Camundongos , Fosforilação , Infecções por Salmonella/genética , Salmonella typhimurium/genética , Salmonella typhimurium/patogenicidadeRESUMO
Salmonella enterica serovar Enteritidis is a major cause of foodborne disease in Uruguay since 1995. We used a genomic approach to study a set of isolates from different sources and years. Whole genome phylogeny showed that most of the strains are distributed in two major lineages (E1 and E2), both belonging to MLST sequence type 11 the major ST among serovar Enteritidis. Strikingly, E2 isolates are over-represented in periods of outbreak abundance in Uruguay, while E1 span all epidemic periods. Both lineages circulate in neighbor countries at the same timescale as in Uruguay, and are present in minor numbers in distant countries. We identified allelic variants associated with each lineage. Three genes, ycdX, pduD and hsdM, have distinctive variants in E1 that may result in defective products. Another four genes (ybiO, yiaN, aas, aceA) present variants specific for the E2 lineage. Overall this work shows that S. enterica serovar Enteritidis strains circulating in Uruguay have the same phylogenetic profile than strains circulating in the region, as well as in more distant countries. Based on these results we hypothesize that the E2 lineage, which is more prevalent during epidemics, exhibits a combination of allelic variants that could be associated with its epidemic ability.
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Proteínas de Bactérias/genética , Surtos de Doenças , Filogenia , Infecções por Salmonella , Salmonella enteritidis/genética , Humanos , Tipagem de Sequências Multilocus , Infecções por Salmonella/epidemiologia , Infecções por Salmonella/genética , Salmonella enteritidis/isolamento & purificação , Uruguai/epidemiologiaAssuntos
Farmacorresistência Bacteriana Múltipla , Infecções por HIV/microbiologia , Plasmídeos/genética , Salmonella typhimurium/genética , Proteínas de Bactérias/genética , Biologia Computacional/métodos , Transferência Genética Horizontal , Humanos , Salmonella typhimurium/classificação , Salmonella typhimurium/isolamento & purificação , UruguaiRESUMO
The rapid emergence of multidrug resistance among bacterial pathogens has become a significant challenge to human health in our century. Therefore, development of next-generation antibacterial compounds is an urgent need. Two-component signal transduction systems (TCS) are stimulus-response coupling devices that allow bacteria to sense and elaborate adaptive responses to changing environmental conditions, including the challenges that pathogenic bacteria face inside the host. The differential presence of TCS, present in bacteria but absent in the animal kingdom, makes them attractive targets in the search for new antibacterial compounds. In Salmonella enterica, the PhoP/PhoQ two-component system controls the expression of crucial phenotypes that define the ability of the pathogen to establish infection in the host. We now report the screening of 686 compounds from a GlaxoSmithKline published kinase inhibitor set in a high-throughput whole-cell assay that targets Salmonella enterica serovar Typhimurium PhoP/PhoQ. We identified a series of quinazoline compounds that showed selective and potent downregulation of PhoP/PhoQ-activated genes and define structural attributes required for their efficacy. We demonstrate that their bioactivity is due to repression of the PhoQ sensor autokinase activity mediated by interaction with its catalytic domain, acting as competitive inhibitors of ATP binding. While noncytotoxic, the hit molecules exhibit antivirulence effect by blockage of S Typhimurium intramacrophage replication. Together, these features make these quinazoline compounds stand out as exciting leads to develop a therapeutic intervention to fight salmonellosis.
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Quinazolinas/farmacologia , Salmonella typhimurium/efeitos dos fármacos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica/genética , Salmonella typhimurium/metabolismo , Transdução de Sinais , Relação Estrutura-Atividade , Virulência/genéticaRESUMO
Multidrug-resistant Salmonella enterica isolates are an increasing problem worldwide; nevertheless, the mechanisms responsible for such resistance are rarely well defined. Multidrug-resistant S. enterica serovar Typhimurium isolates ST3224 and ST827 were collected from two patients. The characteristics of both genomes and antimicrobial resistance genes were determined using next-generation sequencing.
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Commercially available saponins are extracted from Quillaja saponaria barks, being Quil A® the most widely used. Nanoparticulate immunostimulating complexes (ISCOMs or ISCOMATRIX) formulated with these, are able to stimulate strong humoral and cellular immune responses. Recently, we formulated novel ISCOMs replacing QuilA® by QB-90 (IQB-90), a Quillaja brasiliensis leaf-extracted saponin fraction, and reported that IQB-90 improved antigen uptake, and induced systemic and mucosal antibody production, and T-cell responses. However, its mechanism of action remains unclear. In this study we provide a deeper insight into the immune stimulatory properties of QB-90 and ISCOMATRIX-like based on this fraction (IMXQB-90). We show herein that, when used as a viral vaccine adjuvant, QB-90 promotes an "immunocompetent environment". In addition, QB-90 and IMXQB-90 induce immune-cells recruitment at draining-lymph nodes and spleen. Subsequently, we prove that QB-90 or IMXQB-90 stimulated dendritic cells secret IL-1ß by mechanisms involving Caspase-1/11 and MyD88 pathways, implying canonical inflammasome activation. Finally, both formulations induce a change in the expression of cytokines and chemokines coding genes, many of which are up-regulated. Findings reported here provide important insights into the molecular and cellular mechanisms underlying the adjuvant activity of Q. brasiliensis leaf-saponins and its respective nanoparticles.
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Adjuvantes Imunológicos , Nanopartículas/química , Quillaja/química , Saponinas , Vacinas Virais , Adjuvantes Imunológicos/química , Adjuvantes Imunológicos/farmacologia , Animais , Caspase 1/imunologia , Caspases/imunologia , Caspases Iniciadoras , Células Dendríticas/imunologia , Cães , Feminino , Interleucina-1beta/imunologia , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos BALB C , Fator 88 de Diferenciação Mieloide/imunologia , Saponinas/química , Saponinas/farmacologia , Vacinas Virais/química , Vacinas Virais/imunologia , Vacinas Virais/farmacologiaRESUMO
Polyvalent clostridial vaccines, composed of a complex mixture of toxoids from up to 9 different species, are highly effective in controlling clostridial diseases in cattle and sheep. Commercially available vaccines usually state that in normal field conditions two doses administered 4 to 6 weeks apart elicit protective antibody levels that will last for one year. However, studies on the development and duration of the antibody response against the different Clostridium species in target animals are scarce and only partial. Evaluating the temporal evolution of the antibody responses upon vaccination in target species is relevant to understand the bases of protective immunity induced by these vaccines and to develop new optimized vaccines. Here, we assessed the antibody response in sheep against each Clostridium component of two different 9-valent Clostridial vaccines over the period of one year. One vaccine was a commercially available vaccine and the other was an experimental vaccine prepared by us with the same antigens that we used to set up a specific ELISA for each Clostridium species. Both vaccines showed similar results, irrespectively of the origin of the antigens used for the ELISAs, with antibody titers that peaked at day 36 after vaccination and large inter individual variations in the magnitude of the response. Antibody titers were maintained up to 90 days and then markedly decreased, becoming even undetectable in some animals 6 months after vaccination. Given that the current scheme of yearly revaccination has largely shown to be effective at controlling the burden of disease, our results strongly suggest that circulating antibody levels cannot completely explain the protective immunity elicited by these vaccines, and prompt for further studies into the correlates of protection of clostridial vaccines.
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Vacinas Bacterianas/imunologia , Infecções por Clostridium/veterinária , Doenças dos Ovinos/imunologia , Ovinos/imunologia , Tétano/veterinária , Animais , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/imunologia , Clostridium , Infecções por Clostridium/imunologia , Infecções por Clostridium/prevenção & controle , Clostridium tetani , Injeções Subcutâneas , Análise de Componente Principal , Doenças dos Ovinos/microbiologia , Doenças dos Ovinos/prevenção & controle , Tétano/imunologia , Tétano/prevenção & controle , Fatores de Tempo , Vacinação/veterináriaRESUMO
Salmonella enterica serovar Enteritidis is a major agent of foodborne diseases worldwide. In Uruguay, this serovar was almost negligible until the mid 1990s but since then it has become the most prevalent. Previously, we characterized a collection of strains isolated from 1988 to 2005 and found that the two oldest strains were the most genetically divergent. In order to further characterize these strains, we sequenced and annotated eight genomes including those of the two oldest isolates. We report on the identification and characterization of a novel 44 kbp Salmonella prophage found exclusively in these two genomes. Sequence analysis reveals that the prophage is a mosaic, with homologous regions in different Salmonella prophages. It contains 60 coding sequences, including two genes, gogB and sseK3, involved in virulence and modulation of host immune response. Analysis of serovar Enteritidis genomes available in public databases confirmed that this prophage is absent in most of them, with the exception of a group of 154 genomes. All 154 strains carrying this prophage belong to the same sequence type (ST-1974), suggesting that its acquisition occurred in a common ancestor. We tested this by phylogenetic analysis of 203 genomes representative of the intraserovar diversity. The ST-1974 forms a distinctive monophyletic lineage, and the newly described prophage is a phylogenetic signature of this lineage that could be used as a molecular marker. The phylogenetic analysis also shows that the major ST (ST-11) is polyphyletic and might have given rise to almost all other STs, including ST-1974.