Bordetella pertussis, o agente etiológico da coqueluche ou tosse comprida, que estabelece a infecção através da fixação bacteriana no epitélio do trato respiratório superior. Os principais mediadores de adesão da bactéria são a toxina pertússica (PT) e a hemaglutinina filamentosa (FHA). A FHA é a adesina majoritária e contém pelo menos 4 domínios: porção Nterminal, domínio de reconhecimento de carboidrato (CRD) (FHA1141-1279), trinca de aminoácidos Arginina-Glicina-Ácido aspártico (RGD) (FHA1097-1099) e o sítio de ligação a heparina (domínio Hep) (FHA442-863). Neste trabalho, foi realizado a amplificação de duas regiões do domínio de ligação à heparina, as regiões MAL80 (FHA299-873) e HEP (FHA430-873). As regiões foram amplificadas, clonadas no vetor de expressão pAE, expressas utilizando a cepa BL21 SI de E. coli e purificadas. A proteína HEP purificada de E. coli possui baixa afinidade por heparina e não é capaz de aglutinar hemácias. A proteína recombinante HEP purificada foi utilizada para a produção de anticorpos. Através do experimento de ELISA foi demonstrado que o anti-soro anti-HEP é capaz de reconhecer além da região HEP, a região MAL80 e a proteína FHA. Estes resultados foram confirmados por experimentos de Western. Neste período também foi realizada a amplificação do domínio HEP de FHA e da subunidade S1 da toxina pertússica (PT) de B. pertussis através do método de TAP Express. Este método envolve duas reações de PCR e no final do processo é obtido um fragmento que contém uma região promotora (CMV), uma seqüência codificadora e uma região terminadora (SV40), que está pronto para ser introduzido e expresso em animais. De posse deste material e da proteína recombinante HEP, foi realizado o desafio intracerebral com Bordetella pertussis e através do monitoramento dos camundongos foi observado que nenhum dos candidatos testados foi capaz de proteger os animais contra B. pertussis. Foi realizado também a expressão do domínio Hep em lactobacilos, visando um possível sistema de imunizações de mucosas. Os anticorpos produzidos nos camundongos imunizados com a proteína HEP expressa em E. coli, lactobacilos e por vacina de DNA foram capazes de inibir a hemaglutinação promovida pela proteína FHA. / Bordetelfa pertussis, the agent of whopping cough, establishes infection by attaching to the ciliated epithelial cells of the respiratory tract. The bacterial adherence is mediated by pertussis toxin and filamentous hemagglutinin (FHA). FHA is the major adhesin of B. pertussis and displays multipie adherence activities. FHA contains four distin\'ct domains that exhibit specific affinities for different ligands or receptor, the amino-terminal end, the RGD triplet (FHA1097-1099), the lectin domain (FHA1141-1279) and the heparin-binding domain (FHA442-863). In this study, two overlapping regions of the heparin-binding domain, Mal80 (FHA299-873) and Hep (FHA442-873), were amplified by peR and subcloned in pAE expression vectors for E. coli. The fusion proteins in pAE were transformed in E. coli BL21 SI, induced with NaCI 0,3 M and purified using a nickel-charged metal chelating resin. The purified protein has low heparin affinity and does not have hemagglutination activity. The purified protein HEP was used to produce polyclonal antibodies in mouse. The anti-HEP antibodies are able to recognize the HEP, MAL80 and FHA proteins in ELISA and western assays, but anti-FHA only recognized the FHA protein. The genetically detoxified S1 subunit of pertussis toxin and Hep domain were amplified by the TAP Express method. There are two PCR reactions involved in the TAP processo At the end of the process the fragment of interest will carry a CMV promoter and a SV40 terminator and is ready to be introduced into animals or cell by transfection. Groups were immunized with proteins and/or DNA, challenged i.c. with a lethal dose of live Bordetelfa pertussis and the survival was monitored. No groups were protected against the challenge. The recombinant protein HEP were also expressed in Lactobacilfus aiming the development of potential mucosal vaccines. The polyclonal antibodies produce in mouse immunized with DNA and protein Hep expressed in E. coli and Lactobacillus were able to inhibition the FHA hemagglutination activity.

Vaccines play a vital role in public health by preventing infectious disease across the globe. Vaccine formulations represent a weakened form of a microbe or toxin that is injected into the human body to elicit an immune response, generating antibodies to protect against a future infection. To this day, it is a challenge to identify and engineer important antigens and epitopes to focus this immune response in a safe and effective manner. The example of Bordetella pertussis is used to highlight the problems and lessons learned in designing a vaccine for this global epidemic. In particular, this review will focus on the advantages and disadvantages of chemical versus genetic detoxification and whole cell versus acellular vaccines in the context of pertussis. The latter part of this review will provide a summary of general strategies, such as epitope mapping and manipulation, synthesis of truncated variants, reverse vaccinology, and structural vaccinology, that have been successful in addressing increasingly complex diseases. Collectively, these techniques provide an invaluable set of tools to focus the immune response by finding and engineering specific antigens and epitopes. / text

Introduction: Bordetella pertussis is an important human pathogen that causes whooping cough (pertussis), an endemic illness responsible of significant morbidity and mortality, especially in infants and children. Worldwide, there are an estimated of 16 million cases of pertussis, resulting in about 195,000 child deaths per year. In Peru, pertussis is a major health problem that has been on the increase despite immunization efforts. The objective of this study was to determine the prevalence of B. pertussis among children under five years of age suspected to have whopping cough in Cajamarca, Peru. Methodology: Children diagnosed with whooping cough admitted to the Hospital Regional de Cajamarca from August 2010 to July 2013 were included. Nasopharyngeal samples were obtained for B. pertussis culture and polymerase chain reaction (PCR) detection. Results: In 133 children, the pertussis toxin and IS481 gene were detected in 38.35% (51/133) of the cases by PCR, while only 9.02% (12/133) of the Bordetella cultures were positive. The most frequent symptoms in patients with positive B. pertussis were paroxysm of coughing 68.63% (35/51), cyanosis 56.86% (29/51), respiratory distress 43.14% (22/51), and fever 39.22% (20/51). Pneumonia and acute bronchial obstructive syndrome were present in 17.65% (9/51) and 13.72% (7/51) of the cases, respectively. Conclusions: B. pertussis is responsible for an important proportion of whooping cough in hospitalized children in Cajamarca. Epidemiologic surveillance programs for B. pertussis are essential in Peru, especially in children who could most benefit from the vaccine.

Das Tex Protein aus Bordetella pertussis definiert eine neue Familie hoch konservierter Proteine in Eubakterien. Ursprünglich wurde das tex Gen aufgrund seines Einflusses auf die Toxinexpression in bestimmten regulatorischen Mutanten von B. pertussis gefunden (Fuchs et al. (1996), J Bacteriol 178, 4445-52). Wie hier gezeigt wird, sind Leserahmen für entsprechende Proteine bei den Eubakterien ubiquitär und mehrheitlich zu über 69 Prozent konserviert. Eine Ausnahme bilden einige wenige Taxa mit bekanntermaßen reduzierten Genomen, bei denen das Gen wahrscheinlich verloren gegangen ist, wie zum Beispiel verschiedene Mycoplasma spp. oder der obligate Blattlaus- (Aphiden-) Symbiont Buchnera aphidicola. In Eukaryonten und Archaeen konnte ein zu tex homologes Gen bisher nicht gefunden werden. Die Funktion von Tex in der Bakterienzelle ist unklar. Während das Gen in B. pertussis essenziell ist und auch nicht überexprimiert werden kann, sind Deletionsmutanten in Neisseria meningitidis und Escherichia coli phänotypisch nicht von den entsprechenden Wildtypen unterscheidbar. Ausgiebige Wachstumsstudien mit einer E. coli tex-Mutante unter verschiedenen Wachstums- und Stressbedingungen ergaben ebenfalls keinen Hinweis auf eine Bedeutung von Tex, die die außerordentliche Konservierung des Proteins erklären könnte. Das Protein zeigt in seinem N-Terminus ausgeprägte Ähnlichkeit mit dem Mannitol-Repressor (MtlR) von Escherichia coli und besitzt eine C-terminale S1-Domäne. Da die meisten der Proteine mit S1-Domänen als RNA-bindende Proteine gelten, wurde die Fähigkeit von Tex untersucht, mit Nukleinsäuren zu interagieren. In Festphasen-Bindeassays mit an Magnetkügelchen immobilisiertem Tex Protein aus E. coli konnte eine spezifische Bindung an RNA-Gesamtpräparationen gezeigt werden. DNA wurde hingegen nicht gebunden. Durch Verkürzung des N-Terminus geht die präferenzielle Bindung an RNA jedoch verloren und die Bindung von DNA erfolgt mit der gleichen Effizienz wie die von RNA. Festphasen-Bindeassays wurden weiterhin dazu benutzt, mögliche spezifische Interaktionspartner von Tex aus RNA-Gesamtpräparationen zu finden. Tatsächlich konnten über diesen Ansatz die regulatorische RNA CsrB und die ribosomale 16S RNA als spezifische Liganden isoliert werden. Über die biologische Relevanz dieser Interaktion kann zum gegenwärtigen Zeitpunkt allerdings noch keine Aussage gemacht werden. / The Bordetella pertussis Tex protein defines a new family of highly conserved eubacterial proteins. The tex gene was originally found due to its negative influence on toxin expression when over-expressed slightly in certain regulatory mutants of B. pertussis (Fuchs et al. (1996), J Bacteriol 178, 4445-52). As shown in this work apparently homologous reading frames are ubiquitous within the Eubacteria. The majority of these mostly putative proteins are more than 69 per cent conserved. Only some of the genomes of some obligate intracellular pathogens, e. g. various Mycoplasma ssp., or the aphid symbiont Buchnera aphidicola known for their reduced gene numbers were found to lack a tex gene and probably have lost it during phylogeny. The actual function of Tex within the bacterial cell is as yet mysterious as the gene can neither be deleted nor can it be highly over-expressed in B. pertussis. However, deletion mutants in Neisseria gonorrhoeae and Escherichia coli do not show any phenotype different to the respective wild type. Extensive growth studies on an E. coli tex-mutant did not reveal any clue to the reason for the extraordinary conservation of the protein. The N-terminus of Tex shows extensive similarity to the mannitol repressor (MtlR) of E. coli whereas the C-terminus contains an S1 domain. Because most S1 domain containing proteins have been shown to bind RNA, the capacity of Tex to interact with nucleic acids was investigated. In solid phase binding assays with purified Tex protein from E. coli immobilized to magnetic beads the protein showed specific binding of RNA but not DNA. NÓterminal deletions of different length abolished this preference for RNA and both types of nucleic acid bound equally well. Again a solid phase binding assay was used to enrich for specific ligands of Tex from total cellular RNA preparations. This approach identified the regulatory CsrB RNA and ribosomal 16S RNA as preferential binding targets. However, it is not yet possible to comment on the biological relevance of these findings.

A coqueluche é uma doença contagiosa, causada por Bordetella pertussis e B. parapertussis e as vacinas de células inteiras (WCPs) contra pertussis, embora eficazes, foram associadas a efeitos indesejáveis. Já as vacinas pertussis acelulares são menos reatogênicas, mas caras, o que as torna inviáveis para países em desenvolvimento. Nesse estudo, avaliamos a resposta imune induzida por uma vacina pertussis celular com menor teor de LOS (Plow), desenvolvida pelo Instituto Butantan. Para isso, camundongos C57BL-6 foram imunizados com WCP tradicional ou Plow, formulada com MPL-A de B. pertussis ou com Hidróxido de Alumínio e vacinas tríplice bacterianas com componente pertussis acelular, Pertacel ou Adacel. Após o esquema de imunização, foi avaliada a resposta imune humoral e celular contra a B. pertussis, e B. parapertussis, além de resposta inata a um antígeno não relacionado, BCG. Ensaio de transmissão também foi realizado, após desafio com B. pertussis ou B. parapertussis. Nossos resultados consolidam a avaliação da resposta imune humoral e celular induzida pela Plow, além de apresentar resultados bastante interessantes relativos à atividade da Plow na diminuição da transmissão bacteriana, tanto de pertussis quanto de parapertussis. / Bordetella pertussis and B. parapertussis are the causative agents of whooping cough. Whole cell pertussis (wPs) vaccines, although effective, were associated with undesirable effects. On the other hand, aP vaccines are less reactogenic, but expensive, which made their use unable for developing countries. In this study, we assessed the immunological response, induced by a whole cell pertussis vaccine with low LOS content (Plow), developed by Instituto Butantan. To this, C57BL-6 mice were immunized with traditional whole cell pertussis vaccine or Plow, administrated with Monophosporil lipid A from B.pertussis or Aluminum hydroxide, and diphtheria-tetanus-acellular pertussis vaccines, Pertacel or Adacel. After the immunization scheme, were evaluated humoral and cellular immune responses against B. pertussis and B. parapertussis, in addition to innate response to an unrelated antigen, BCG. Transmission assay was also performed, after B. pertussis or B. parapertussis challenge. Our results consolidated the immune humoral and cellular responses induced by Plow, besides interesting results with regards the efficacy of the vaccine in decreasing the transmission of B. pertussis and B. parapertussis in mice.

Objectives To report the incidence, epidemiology, and clinical features of Bordetella pertussis in Peruvian infants under 1 year old. Patients and methods A prospective cross-sectional study was conducted in five hospitals in Peru from January 2010 to July 2012. A total of 392 infants under 1 year old were admitted with a clinical diagnosis of whooping cough and tested for B. pertussis by PCR. Results The pertussis toxin and IS481 genes were detected in 39.54% (155/392) of the cases. Infants aged less than 3 months were the most affected, with a prevalence of 73.55% (114/155). The most common household contact was the mother, identified in 20% (31/155) of cases. Paroxysm of coughing (89.03%, 138/155), cyanosis (68.39%, 106/155), respiratory distress (67.09%, 104/155), and breastfeeding difficulties (39.35%, 61/155) were the most frequent symptoms reported. Conclusion An increase in pertussis cases has been reported in recent years in Peru, despite national immunization efforts. Surveillance with PCR for B. pertussis is essential, especially in infants less than 1 year old, in whom a higher rate of disease-related complications and higher mortality have been reported. / This 312 work was supported by Sanofi Aventis del Peru. Conflict 313 of interest: On behalf of all authors, the corresponding author 314 states that there are no conflicts of interest or funding related 315 to this study

Bordetellen sind Gram-negative Kokkobazillen, die phylogenetisch zu den β-Proteobakterien zählen und in der Familie der Alcaligenaceae eingeordnet sind. Der bedeutendste Vertreter der Gattung, die nach heutigem Kenntnisstand neun Arten umfasst, ist Bordetella pertussis, der Erreger des Keuchhustens. Der Keim ist obligat humanpathogen und besitzt zahlreiche Virulenzfaktoren, um die Epithelzellen des Respirationstraktes zu besiedeln und zu zerstören, wodurch es zu dem charakteristischen Krankheitsverlauf kommt. Neben B. pertussis werden noch B. bronchiseptica und B. parapertussis dem sogenannten B. bronchiseptica-Cluster zugeteilt. Alle Vertreter des B. bronchiseptica-Clusters sind in der Lage, bei verschiedenen Wirtsspezies respiratorische Erkrankungen mit unterschiedlichem Schweregrad auszulösen. Dabei weist B. bronchiseptica ein breiteres Wirtsspektrum auf und kann Atemwegserkrankungen in einer Vielzahl von Säugetieren auslösen, wohingegen B. parapertussis vornehmlich Schafe und Menschen infiziert und bei letzteren eine schwächere Form des Keuchhustens bewirkt. Das Hfq-Protein wurde ursprünglich als Wirtsfaktor identifiziert, welcher für die Replikation des RNA-Phagen Qβ in Escherichia coli benötigt wird (host factor for Qβ oder HF-1). Es ist in Struktur und Funktion homolog zu den Sm-Proteinen aus Eukaryoten, die am Splicing von mRNAs involviert sind. Die Beteiligung des Hfq-Proteins an regulatorischen Vorgängen, die durch kleine nicht-kodierende RNAs (sRNAs) vermittelt werden, wurde erstmals in einer Studie zum Mechanismus der rpoS-Regulation durch die kleine regulatorische RNA OxyS ersichtlich. Seitdem konnte für eine Vielzahl an sRNAs gezeigt werden, dass sie an Hfq gebunden vorliegen und die Hilfe des Proteins bei der post-transkriptionellen Kontrolle ihrer Ziel-mRNAs benötigen. In dieser Hinsicht übernimmt Hfq die Rolle eines RNA-Chaperons, indem es trans-kodierte sRNAs stabilisiert und die Basenpaarung mit ihren Ziel-mRNAs fördert. Dabei beeinflusst die Bindung der sRNA-Regulatoren an ihre Ziel-mRNAs deren Translation, sowohl aktivierend als auch inhibierend. Bislang wurden Hfq-Homologe in der Hälfte aller sequenzierten Gram-positiven und Gram-negativen Bakterienarten gefunden. Eine BLAST-Analyse ergab, dass B. pertussis und B. bronchiseptica Homologe zum Hfq-Protein aufweisen und diese in der veröffentlichten Genomsequenz bereits als Hfq-Protein annotiert sind. Fokus dieser Arbeit war weitestgehend, die Funktion des Hfq-Proteins in B. pertussis und vergleichend in B. bronchiseptica zu charakterisieren. Mittels Primer Extension-Analyse konnte zunächst der Startpunkt des hfq-Transkripts in B. pertussis und B. bronchiseptica unter logarithmischen Wachstumsbedingungen bestimmt werden. Dieser Startpunkt war zudem unter stationären Wachstumsbedingungen und nach Hitzestress aktiv, was in Diskrepanz zur Beobachtung in E. coli steht. Ferner konnte festgestellt werden, dass die hfq-Transkription nach Induktion verschiedener Stressformen in beiden Organismen erhöht war. Nach Generierung der jeweiligen Δhfq-Mutanten in beiden Organismen wurden diese charakterisiert. Die B. pertussis Δhfq-Mutante zeigte ein deutliches Wachstumsdefizit gegenüber dem Wildtyp, im Gegensatz zu B. bronchiseptica Δhfq, die sich im Wachstum wie der Wildtyp verhielt. Beide Mutanten zeigten sich sensitiver gegenüber H2O2-Stress als der Wildtyp, nicht jedoch gegenüber weiteren oxidativen Stressbedingungen oder Membranstress induzierenden Substanzen. Die Δhfq-Mutante in B. pertussis war zudem in ihrer Fähigkeit zur Biofilmbildung beeinträchtigt, was jedoch nicht für B. bronchiseptica Δhfq galt. Da Hfq an sRNA-mRNA-Interaktionen, welche die Translation der mRNAs beeinflussen, beteiligt ist, sollte über 2D-Gelelektrophorese das Hfq-regulierte Proteom in B. pertussis und B. bronchiseptica bestimmt werden. Auffällig war, dass viele periplasmatische Transport-bindeproteine von der Δhfq-Mutation betroffen waren. Es zeigten sich aber auch Stoffwechselenzyme und wichtige Housekeeping-Faktoren, wie z. B. der Elongationsfaktor EF-Tu und das Chaperon GroEL, in der Δhfq-Mutante dereguliert. Generell scheint das Hfq-regulierte Proteom in B. pertussis und B. bronchiseptica nur einen kleinen Teil des gesamten Proteoms auszumachen. Zudem ist das Hfq-regulierte Proteom variabel zwischen verschiedenen Wachstumsbedingungen, aber auch zwischen den beiden Organismen trotz der engen Verwandtschaft. Die Expression ausgewählter Virulenzfaktoren zeigte keinen Unterschied zwischen Δhfq-Mutante und B. pertussis-Wildtyp. / Bordetellae are Gram-negative coccobacilli phylogenetically belonging to the β-group of proteobacteria and therein to the family of Alcaligenaceae. The most prominent member of the genus comprising nine species so far is Bordetella pertussis, the etiological agent of whooping cough. This organism is an obligatory human pathogen and expresses a variety of virulence factors in order to colonize and destroy the epithelial cells of the respiratory tract causing the characteristic symptoms of the disease. In addition to B. pertussis, B. bronchiseptica and B. parapertussis are assigned to the so-called B. bronchiseptica-cluster. All members of the B. bronchiseptica-cluster have the ability to cause respiratory symptoms with varying severity. B. bronchiseptica exhibits a broad host range causing respiratory symptoms in a variety of mammals, whereas B. parapertussis infects sheep and humans causing a milder form of whooping cough in the latter. The Hfq protein was originally identified as a host factor necessary for the replication of the RNA-phage Qβ in Escherichia coli (host factor for Qβ or HF-1). It is functionally and structurally homologous to Sm-proteins involved in splicing of mRNAs in eukaryotes. The involvement of Hfq in regulatory processes caused by small non-coding RNAs (sRNAs) was first recognized in a study on the mechanism of rpoS-regulation by the small regulatory RNA OxyS. Since then a variety of sRNAs were shown to be bound to Hfq and require its help for post-transcriptional control of their target-mRNAs. In this regard, Hfq functions as an RNA-chaperone by stabilizing trans-encoded sRNAs and their basepairing to target-mRNAs. Binding of the sRNA-regulators to their target-mRNAs thereby either activates or inhibits their translation. To date Hfq homologues were identified in half of all sequenced Gram-positive and Gram-negative bacterial species. BLAST analysis revealed that B. pertussis and B. bronchiseptica possess an Hfq homologue which has already been annotated as such in the published genome sequence. The main focus of this work was to characterize the function of the Hfq protein in B. pertussis as well as in B. bronchiseptica. By primer extension analysis we could identify the start of the hfq-transcript in B. pertussis and B. bronchiseptica under logarithmic growth conditions. This transcriptional start site was also active under stationary growth conditions and after heat shock which is discrepant from the observations in E. coli. Furthermore, it could be shown that the hfq-transcription was elevated in both B. pertussis and B. bronchiseptica under various stress conditions. Δhfq-mutants were established and characterized in both organisms. The Δhfq-mutant of B. pertussis exhibited a pronounced growth deficit in comparison to the wildtype whereas the Δhfq-mutant of B. bronchiseptica showed the same growth properties as the wildtype. Both Δhfq-mutants expressed a higher sensitivity to stress caused by H2O2 compared to the wildtype. However, there was no increased sensitivity of the Δhfq-mutants to other oxidative stress agents or membrane stress inducing agents. Furthermore, the Δhfq-mutant of B. pertussis but not the Δhfq-mutant of B. bronchiseptica was impaired in its ability to form biofilms. Since Hfq is involved in sRNA-mRNA-interactions affecting the efficient translation of mRNAs, the Hfq-regulated proteome of B. pertussis and B. bronchiseptica was determined by 2D-gelelectrophoresis. Strikingly, a variety of periplasmic binding proteins involved in transport were affected by the Δhfq-mutation. In addition, enzymes of various metabolic pathways and important housekeeping factors, such as elongation factor EF-Tu and the protein chaperone GroEL, were deregulated in the Δhfq-mutant. The Hfq-regulated proteome comprises generally only a small part of the complete proteome in B. pertussis and B. bronchiseptica. Furthermore, this Hfq-regulated proteome differs between certain growth conditions as well as between the two closely related organisms. No difference could be observed in the expression of selected virulence factors between B. pertussis Δhfq and wildtype.

In this study, a suite of single antigen DNA vaccines, combination DNA vaccines and dual modality vaccines, were developed and evaluated for their potential to induce humoral and cell-mediated immune responses, and protective efficacy against Bordetella pertussis, the aetiological agent of whooping cough, using the mouse respiratory challenge model.

Background: Pertussis, also known as whooping cough, is an infectious disease of the respiratory tract caused by the bacterium bordatella pertussis. In 2008, the Centers for Disease Control and Prevention (CDC) recommended that all post-partum women not previously vaccinated receive the Tdap (tetanus, diphtheria and acellular pertussis) vaccine in the immediate post-partum period in an effort to protect their newborns from this serious infection. In response, some birthing units in the state of Georgia have introduced programs to administer the Tdap vaccination to their post-partum patients. The purpose of this study is to examine the availability, design and effectiveness of these programs. Methodology: Surveys were sent to 72 birthing units in the state of Georgia. The survey was designed to illicit insight into each birthing unit’s policies and procedures regarding the administration and dissemination of the vaccine to post-partum women. In addition, the data collection instrument provided information on which centers offer the vaccination to their post-partum patients, how successful their program has been in reaching the target population and what barriers may need to be addressed to increase vaccination compliance to this particular population. Results: The results of this study suggest that Tdap programs in Georgia birthing centers can be successful in reaching the intended population and educating patients and hospital staff members on the importance of this vaccination. The majority of birthing centers in middle and southern Georgia do not have Tdap programs. In addition, barriers to Tdap vaccination programs have been identified such as lack of funding, lack of available education resources for both patient and hospital staff and absence of available staff members to administer vaccination. Conclusion: The examination of Tdap vaccination programs in Georgia birthing units can assist policy makers and public health agencies understand how to best allocate resources in an effort to increase vaccination compliance. Further research needs to be conducted to better understand how to improve program quality and availability state-wide and to correlate birthing center Tdap programs with increased vaccination compliance in postpartum women.

Objetivos generales de la tesis: Con el desarrollo de esta propuesta se espera contribuir al conocimiento que sirva de base para el diseño de una vacuna más efectiva contra pertussis, no sólo en términos generales, sino en lo que se refiere a su efectividad en Argentina, determinando la definición de la/s cepas / componentes a incluir en una nueva formulación. Objetivos específicos de la tesis: En este marco conceptual y tomando como hipótesis la divergencia de la población bacteriana circulante respecto a las cepas vacunales hoy en uso, se proponen los siguientes objetivos: 1-Caracterizar mediante la aplicación de técnicas de genómica funcional los aislamientos de B. pertussis locales. Identificar de nuevos inmunógenos. Los aislamientos clínicos de nuestra colección han sido previamente agrupados en base a sus divergencias a nivel de las secuencias que codifican para antígenos específicos, así como en su genoma completo a través de los ensayos de PFGE y los años en que fueron aislados. Proponemos abordar la búsqueda de diferencias específicas, a nivel de expresión génica, entre los aislamientos circulantes y las cepas que hoy se usan en la producción de vacunas. Para ello hemos emplearemos herramientas de genómica funcional, proteómica e inmunoblot; para identificar potenciales candidatos vacunales a incorporar en una formulación acelular. 2-Analizar la relevancia de la divergencia genética entre la Población Bacteriana Circulante (PBC) y las cepas vacunales respecto a la protección contra pertussis. Para abordar este punto emplearemos el modelo animal de desafío intranasal. Consideramos que la información obtenida marcará la necesidad o no de incluir determinadas variantes polimórficas para obtener una nueva vacuna más efectiva. El abordaje de este aspecto se espera también contribuya al conocimiento general de la adaptación y evolución bacteriana bajo la presión de selección ejercida por las diferentes estrategias de vacunación (celular/acelular). / Tesis digitalizada en SEDICI gracias a la Biblioteca Central de la Facultad de Ciencias Exactas (UNLP).