Conventional pertussis vaccine prepared from killed whole cell B. pertussis organisms has been in widespread use since the early 1950's. Despite marked reductions in the incidence of pertussis, the use of the vaccine has caused concern because of questions of significant adverse reactions. Whooping cough is not notifiable in South Africa, and there is consequently a paucity of hard data on efficacy; in addition few cases are proven. Incidence, prevalence, severity and transmission of the disease hence remain a matter of conjecture. In order to provide background information and determine baseline data for undertaking further studies, available clinical and epidemiological data on whooping cough (pertussis) in South Africa was collated. It was intended to compare the pattern of disease seen in this country with that known in other parts of the world. Clinical and epidemiological findings from 1525 whooping cough admissions (diagnosed on the basis of clinical criteria) obtained from 6 major infectious disease hospitals around the country for the period 1980-1987/1988 are reported. The data from Durban were available in some detail. Incidence, morbidity and mortality of the disease in South Africa, as elsewhere, was higher in infancy. Infants and young children were predominantly affected, with 31.3% of cases under 12 months of age and 7.2% less than 2 months. Mortality was disproportionately higher in infancy; 53 .2% of deaths were in those younger than 12 months of age. There was a slight female preponderance, both in respect of prevalence and mortality. Patients were admitted with pertussis throughout the year, although there was a peak during the winter months (May to October). The typical whoop accompanied the cough in 55.6% of patients . A raised white cell count was recorded in 66% of patients. The most commonly detected The average duration of hospital stay was between 1 0-13 days. Of those with vaccination records, 26% were unvaccinated, 44% had 1 or 2 doses, and 27% had been fully vaccinated with a whole-cell pertussis vaccine combined with diphtheria and tetanus toxoids. The picture, incomplete though it is, reveals a pattern of pertussis similar to that described in other developing countries. The study reveals huge gaps in our knowledge of this subject in South Africa. More research needs to be done, particularly with respect to improved diagnosis, prevention and treatment; further pertussis should be made a notifiable disease in South Africa. The whole-cell pertussis vaccine currently used in South Africa has not been adequately evaluated for post-vaccination events and immunogenicity. The development of new purified component pertussis vaccines, which appear to be safer than conventional whole-cell preparations and of equal or almost equal efficacy (although optimal vaccine composition remains to be defined), requires that the concept of early vaccination with this vaccine compared with conventional whole-cell vaccine be examined in order to optimise the immune response to these vaccines in infants; more especially since neonates do not appear to benefit from passive immunity. Acellular pertussis vaccine has not been evaluated previously in neonates. In order to address the problem of high morbidity and mortality from pertussis in early infancy; and the incorporation of the vaccine into routine vaccination schedules, a phase 11 trial of acellular and whole-cell vaccines was undertaken in very young infants. The effect of neonatal vaccination with acellular pertussis vaccine on subsequent immunity; and the immunogenicity and shortterm safety and reactogenicity of routine primary vaccination with acellular vaccine compared with conventional whole-cell preparations was investigated. Three hundred and forty-five healthy, full-term African babies were, enrolled in the study at birth; 58% of whom were successfully followed for 9 months. Infants were assigned to 1 of 3 vaccine groups in sequence at birth and received either acellular or whole-cell pertussis vaccine , combined with 0 and T (A-OTP or W-OTP) at 2, 4 and 6 months of age. Groups I and 11 received A-OTP and Group III W-OTP. In addition, at birth, Group I received an additional dose of A-OTP and Group 11, a saline placebo injection. No unvaccinated controls were studied for ethical reasons. Serologic IgG responses to 3 major protective antigens of B. pertussis, filamentous haemagglutinin (FHA), pertussis toxin (PT) and fimbrial agglutinogens 2,3 (AGG2,3), were measured by ELlSA in sera obtained at birth, and before vaccination at 2, 4 and 6 months and at 9 months of age. The incidence and nature of post-vaccination events were recorded for 14 days after each dose. A-OTP induced serum IgG responses to PT and FHA comparable with those reported in other studies, with peak PT titres occurring at 6 months of age after 2 doses in babies vaccinated according to the routine schedule (Group 11). Surprisingly, response to W-OTP was found merely to restore levels of antibody to PT and FHA to those found in cord blood after 3 doses of vaccine, which questions the immunogenicity of the South African product. Four doses of AOTP (Group I) did not produce a better antibody response than the 3-dose schedule. Incidence of all post-vaccination events to both acellular and whole-cell vaccines was low (85.96/1000 doses, 136.36/1000 doses and 76.30/1000 doses in Groups I, 11 and III respectively). Minor symptoms were more common in recipients of A-OTP, although no significant differences in rates were demonstrated. Neurologic post-vaccination events (without sequelae) occurred more frequently hi recipients of W-OTP. No infant vaccinated with A-OTP from 2 months of age (Group 11) experienced a neurologic symptom. The risks of major post-vaccination events cannot however be fully quantified In a study of this small size and diseases. A-DTP vaccination commencing at birth produced final antibody titres to PT and FHA which were superior to those of South African whole-cell vaccine but were considerably lower than when the vaccine was incorporated into routine schedules commencing at 2 months of age. The study findings suggest that acellular pertussis vaccines, whether given from birth or from the age of 2 months, appeared safer and produced a better serologic response than the South African whole-cell product which may have impaired immunogenicity. During the course of the above study, 11 full-term infants with pertussis infection (10 subclinical) were retrospectively diagnosed on the basis of ser~logic evidence. Of the infants with subclinical disease, all 10 had a ~ 4-fold rise in at least 2 different pertussis IgG antibodies and nine had ~ 4-fold rise in all 3 IgG antibodies measured. Seven infants had raised IgA antibodies to PT and FHA and 4 infants had raised IgA antibodies to AGG2,3. Subclinical infection provoked antibody production to multiple antigens to differing degrees. The role of various factors which may have contributed to asymptomatic infection were analysed, viz - household contacts; type of antibody response (clinical vs. subclinical; vaccinated vs. subclinical); maternally acquired antibody levels; vaccination status (number of vaccine doses received); age and gender; and nutritional status. SpeCial features of the study which require emphasis are: pertussis remained subclinical in unvaccinated babies (most of the subjects were unvaccinated). Subclinical infection followed incomplete primary vaccination with either acellular or whole-cell vaccines commencing at 2 months of age. Subjects were 8 months of age or younger and only 1 had completed primary vaccination. Other infections of infancy were commonly detected; 4 infants had upper Likelihood of subclinical infection was related to significantly lower levels of maternallyacquired pertussis IgG-AGG2,3 antibodies but not associated with infants' nutritional status. Subclinical pertussis is described in very young African babies at an age when the disease is most severe, and therefore has implications for infant morbidity and mortality; it is also relevant to disease surveillance and vaccine-efficacy studies . Some perinatal factors influencing vaccination were also explored . In this context we looked at: i. The acquisition of maternal antibodies to B. pertussis in paired mother-infant sera from both well -nourished and SGA full-term and pre-term infants, and infants who subsequently developed pertussis infection, and effect of these maternally-acquired antibodies on subsequent antibody responses to acellular pertussis vaccine administered soon after birth, and to acellular and whole-cell vaccines administered from the age of 2 months. if. The acquisition of maternal antibodies to diphtheria and tetanus (OT) in paired motherinfant sera from full-term and pre-term infants, and the effect of these maternallyacquired antibodies on serologic responses to neonatal OT vaccination followed by whole-cell OTP vaccination at 2, 4 and 6 months. Maternal antibodies were measured since little is known about materno-fetal transfer of pertussis antibodies, especially in African countries where inhibition of placental transfer might occur for a variety of reasons. Furthermore, the effect of peri-natal malnutrition and prematurity on transplacental transfer of diphtheria, tetanus and pertussis antibodies has not been conclusively established in these areas. sera with levels in the latter frequently being higher, Indicating active transplacental transfer of antibodies. The significant pertussis antibody levels in maternal sera were unlikely to be due to the currently used South African whole-cell vaccine (given the poor antibody response to PT and FHA shown in this study). It is assumed that the presence of these antibodies are the end result of natural infection and therefore that pertussis is widespread in the African community. Maternal and cord IgG-PT and AGG2,3 titres were significantly lower (p <0.05) and maternal IgG-FHA marginally lower (p ... O.05) in SGA infants compared to cohorts, although placental transfer was equally efficient in both groups~ This study has demonstrated that the high titres of maternally derived circulating B. pertussis antibodies do not have an inhibitory effect on the subsequent serologic response to acellular vaccine administered in early infancy (2 months) or with the first dose given soon after birth. Protective levels of diphtheria and tetanus antibodies were detected in 100% and 76% of cord sera in pre-term infants and in 85% and 67% of cord sera in full-term infants. Although the number of infants studied was too small to make a definite comment, there did not appear to be 'tolerance' due to neonatal diphtheria-tetanus vaccination of pre-term infants, or to high levels of maternally-acquired antibody. During analysis of the data from the phase 11 study of acellular pertussis vacclnes, 25 infants with protein-energy-malnutritlon (PEM) were detected on the basis of anthropometric indices. Seventeen infants were smaIJ-for-gestatlonal-age (SGA). of whom 9 developed PEM by the age of 9 months. Eight other infants developed post-natal PEM before the completion of the primary vaccination course. and extent of immunological impairment, if any, on the serologic responses to acellular vaccine and to conventional whole-cell OTP in SGA infants, and in infants who developed PEM before the completion of the primary vaccination course. The following indices were evaluated in malnourished infants; (i) anthropometric indices of nutritional status, (ii) intercurrent illnesses including pertussis infection, (iii) post-vaccination events, (iv) serologic responses to vaccination. Results were compared with those obtained in well-nourished (WN) age- and vaccine-matched cohorts. Overall, peak titres and seroconversion data of all 3 antibodies were not significantly different in malnourished infants though final anti-AGG2,3 titres (at 9 months of age) in Group III were significantly lower (p = 0.035). Although peak and final PT and FHA antibody titres in many SGA and malnourished infants were lower than in WN infants and peak responses were attained at a later age, malnutrition did not significantly affect the response to A-OTP. Peak and final AGG2,3 antibody titres were similar in SGA, malnourished and WN infants. Overall malnourished infants responded no less well to pertussis vaccination than did other vaccinees. Incidence of minor local and systemic post-vaccination symptoms was not significantly different in PEM and WN groups although induration at injection site and irritability were more frequently reported in the latter. No major neurologic post-vaccination symptoms to either vaccine were reported in SGA infants or infants with PEM at the time of vaccination. No significant differences was noted in the incidence of major symptoms in PEM as compared with WN infants. One male infant (Group I) who was malnourished at birth and who had been given 2 doses of AOTP, developed clinical signs of pertussis infection between 2 and 4 months of age. Pertussis antibody levels immediately prior to infection were not significantly different from those of un infected age-matched cohorts. The percentage of infants afflicted with common childhood illnesses were similar in PEM and WN groups (46 vs. 43.2%) although the former group incurred significantly more illnesses at an earlier age' (s6 months) (p=O.05, chi square). These findings, albeit preliminary given the small numbers of subjects studied, suggest that acellular pertussis vaccine may be incorporated into routine vaccination schedules followed in developing countries with the expectation that adequate antibody responses will be provoked in SGA infants and in infants who develop post-natal PEM and that the incidence of vaccinerelated adverse effects will be no higher than in WN infants. Further and more extensive studies are indicated before the use of acellular pertussis vaccines can be recommended for routine primary vaccination of infants in preference to whole-cell preparations in developing countries. / Thesis (Ph.D.)-University of Natal, Durban, 1991.

En este trabajo de tesis doctoral y bajo la hipótesis de la utilización por parte la bacteria de Bordetella pertussis del desarrollo en biofilm como un posible mecanismo de persistencia en el hospedador se evaluó la capacidad de formación de biofilm por parte de aislados clínicos de Bordetella pertussis obtenidos de pacientes infectados y tratados en el Hospital Sor María Ludovica de la ciudad de La Plata. Todos los aislados clínicos estudiados, incluyendo cepas aisladas entre los años 2001 y 2007 presentaron mayor capacidad de formación de biofilm en superficies abióticas cuando se las comparó con la cepa de referencia Bordetella pertussis Tohama I, cepa adaptada al laboratorio y utilizada en la producción de vacunas contra la enfermedad. De los ocho aislados clínicos estudiados se seleccionó uno, debido a una exacerbada producción de biofilm (produjo 5 veces más biomasa en las mismas condiciones de cultivo cuando se la comparó con la cepa de referencia) y se realizó un estudio de proteómica comparativa para establecer un posible fenotipo adaptado a la condición de cultivo en estudio. Los experimentos se llevaron a cabo en colaboración con el grupo de la Dra. Monika Elingh Schulz de la Universidad de Veterinaria de Viena, Austria y se utilizó la tecnología 2D-DIGE para evaluar la expresión diferencial de proteínas de la fracción soluble entre ambas cepas en condiciones de cultivo líquido y biofilm. Un análisis de componente principal indicó que las diferencias más significativas entre los fenotipos expresados es consecuencia de la diferencia de cepas y posteriormente consecuencia de la condición de cultivo. Se identificaron 32 proteínas diferencialmente expresadas (mayor de 3 veces de diferencia) entre ambas cepas, las cuales fueron asignadas a las categorías: metabolismo y obtención de energía, síntesis de aminoácidos y proteínas, transporte y virulencia (Bcrh2, OmpQ, Vag8 y BrkA). Todos los factores de virulencia identificados corresponden a proteínas reguladas por el sistema de dos componentes BVGAS. Sabiendo que dicho sistema regula la expresión de importantes adhesinas en Bordetella pertussis y que su inactividad provoca la incapacidad de formación de biofilm es que se decidió estudiar a través de la técnica de PCR en tiempo real la expresión del ARN mensajero de un conjunto mayor de factores de virulencia regulados por este sistema incluyendo adhesinas tales como FHA, PRN, Fim, BipA y la proteína de secreción del sistema tipo III Bsp22 que ha sido reportada en la bibliografía como participante junto a Fim en la producción de filamentos que brindarían estructura a la matriz extracelular del biofilm de B. pertussis. Los resultados obtenidos indicaron un aumento en la expresión de ARN mensajero para la totalidad de los genes estudiados indicando que posiblemente el aumento en la expresión de adhesinas reguladas por el sistema BvgAS, sumado a la expresión de enzimas involucradas en la producción de energía que podrían explicar la alta velocidad de duplicación de la cepa clínica Bp2723 le conferirían la capacidad de formar un biofilm más robusto comparado con la cepa de referencia en las condiciones de cultivo estudiadas.

Made available in DSpace on 2014-12-05T18:34:10Z (GMT). No. of bitstreams: 2 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) 140.pdf: 1388646 bytes, checksum: 577741ef3784956408ba81f9fa41b743 (MD5) Previous issue date: 2006 / Fundação Oswaldo Cruz. Instituto Nacional de Controle de Qualidade em Saúde / Em nosso trabalho, investigamos a resposta inflamatória produzida em camundongos por duas vacinas DTP de diferentes produtores (DTP-1 e DTP-2) em um estudo comparativo com as vacinas DT, DTP Acelular e uma vacina de Referência. Utilizando-se da metodologia da medida do edema de pata em camundongos que constitui-se de um modelo que permite a investigação de reações locais e do estudo das população de leucócitos, que constitui-se de um modelo de investigação da reação sistêmica, procuramos investigar a resposta inflamatória aguda induzida pela vacina DTP. Esta medida do edema foi tomada através de um pletismômetro e a contagem de células foi feita em câmaras de Neubauer. Paralelamente, foram também realizados: um estudo histopatológico, onde analisou-se o envolvimento dos tecidos epitelial e muscular; a dosagem do conteúdo de citocinas próinflamatórias (IL-1β, IL-6 e TNF-α) pelo método de ELISA; a quantificação do conteúdo de LPS através do LAL para todas as vacinas e a quantificação do conteúdo da PT pelo método do efeito clustering em células CHO para as vacinas que contêm o componente pertussis. Observamos que o edema de pata foi mais intenso nas vacinas onde o conteúdo de PT foi maior como na DTP-2 e Referência, não descartando um possível envolvimento do LPS. A vacina de Referência destacou-se pela alta concentração de LPS. Os leucócitos sofreram alterações após o tratamento com as vacinas DTP-1, DTP-2 e Referência, com promoção de acentuada neutrofilia, sendo também observado, em alguns casos discreto aumento dos linfócitos e dos monócitos. A análise histopatológica revelou a formação de intenso exsudato purulento e acúmulo de polimorfonucleares para todas as vacinas. Com relacão a produção de citocinas pró-inflamatórias, foi evidenciada somente a citocina IL-6, citocina característica de processos inflamatórios causados por bactérias. A presença desta interleucina foi detectada somente para as vacinas DTP-1, DTP-2 e Referência. / DTP vaccine has been used for more than 70 years and still represents a majorprophylactic measure in the control of pertussis, tetanus and diphtheria infectionsdespite being associated with severe adverse reactions such as neurologicalderangements, seizures and shock. Sparse information can be obtained regarding thecorrespondence between toxicity test in animal models and the occurrence of adversereactions upon vaccination of humans. The mechanisms implicated in the adversereactions associated with DPT vaccine is still not fully understood, but several evidencepoint to LPS and active residues of pertussis toxin (PT) as the main culprits for local andsystemic complications. In this work we investigated the inflammatory response inducedby two different DTP vaccines (originated from two different producers – DPT-1 andDPT-2) in mice and compared this with DT vaccine, acellular DPT vaccine, and aReference vaccine. We applied the paw edema model using a pletismograph, followedby systemic analysis of the leukogram, analysis of the blood levels of pro-inflammatorycytokines (IL-1 ,IL-6eTNF- ) and histopathological analysis of the paw tissue. LPSand PT contents were determined by the LAL and CHO cells clustering test,respectively. We observed that paw edema was more intense in those vaccines with ahigher content of PT, although the contribution of LPS to the reaction could not be ruledout. A marked neutrophilia and a discrete increase in lymphocyte and monocytenumbers were observed with DPT-1, DPT-2 and the Reference vaccine. Histologicalexaminations revealed an intense exsudative process with neutrophilia and pusformation in all vaccines tested. Cytokine analysis showed that IL-6 was the onlycytokine with increased levels detected after DTP-1, DPT-1 and Reference vaccinechallenge.

Submitted by Lívia Faria Orso (livia_orso@hotmail.com) on 2018-04-19T13:08:51Z No. of bitstreams: 1 dissertação_mestrado_orso_faria_livia.pdf: 2240731 bytes, checksum: 33859f25eb32033ff60f35a5cfe6e9ab (MD5) / Approved for entry into archive by Luciana Pizzani null (luciana@btu.unesp.br) on 2018-04-19T15:03:41Z (GMT) No. of bitstreams: 1 orso_lf_me_bot.pdf: 2240731 bytes, checksum: 33859f25eb32033ff60f35a5cfe6e9ab (MD5) / Made available in DSpace on 2018-04-19T15:03:41Z (GMT). No. of bitstreams: 1 orso_lf_me_bot.pdf: 2240731 bytes, checksum: 33859f25eb32033ff60f35a5cfe6e9ab (MD5) Previous issue date: 2018-03-01 / Introdução: A coqueluche é uma doença que compõe a Lista Nacional de Doenças de Notificação Compulsória, e várias hipóteses têm sido sugeridas para a reemergência da coqueluche no Brasil e no mundo. Sabe-se que a faixa etária mais acometida é constituída por crianças menores de um ano e que o monitoramento rigoroso da segurança vacinal é a principal estratégia para adesão aos programas de imunização. Além disso, é imprescindível que os serviços de vigilância no Brasil se mantenham alertas sobre a situação epidemiológica da coqueluche. Objetivo: Analisar o perfil epidemiológico dos casos de coqueluche em crianças do nascimento há quatro anos de idade, retrospectivos à implantação da vacina dTpa em gestantes, nos municípios de Botucatu e Marília do Estado de São Paulo. Método: Trata-se de um estudo descritivo, comparativo e de série histórica no município de Botucatu e Marília. Os dados foram coletados do Sistema Nacional de Agravos de Notificação (SINAN), de crianças na faixa etária de zero a quatro anos, confirmadas com coqueluche, no período de primeiro de janeiro de 2008 a 31 de outubro de 2014 do município de Botucatu e Marília. Resultados: Observa-se um aumento gradativo dos casos a partir de 2014, nos dois municípios. A sazonalidade dos casos confirmados em Botucatu ocorreu no inverno, com 42,86% de ocorrências, e em Marília, no verão, com 35,71%. A faixa etária mais acometida pela doença foram crianças menores de um ano em ambos os municípios. O exame da distribuição dos casos confirmados por região geográfica mostrou que, em Botucatu, o maior registro de casos foi na zona norte, com 57,14%, e para o município de Marília, na zona sul, com 42,86%. Quanto ao esquema vacinal, Botucatu apresentou vacinação em 57,14% das crianças, e Marília, 52,38%. Em Botucatu, 35,71% das crianças apresentaram complicações da doença, e em Marília, 16,66%, sendo a pneumonia a complicação mais frequente. Em Botucatu e Marília 100% e 92,85% das crianças evoluíram para a cura respectivamente. Conclusão: A vacina pentavalente que contém o componente pertussis, continua sendo uma medida de prevenção importante, porém não está sendo efetiva devido acometimento da faixa etária inferior ao recomendado pela vacina nos dois municípios. Diante disso este estudo sugere estudos que avaliem o perfil epidemiológico da coqueluche após a implementações de medidas de prevenção pelo Ministério da Saúde, como a vacina dTpa na gestante. / Introduction: Pertussis is a disease that belongs to the National List of Compulsory Notification Diseases, and several hypotheses have been suggested for the re-emergence of pertussis in Brazil and worldwide. It is known that the most affected age group is children under one year old and strict monitoring of vaccine safety is the main strategy for adherence to immunization programs. In addition, it is imperative that surveillance services in Brazil be alert about the epidemiological situation of pertussis. Objective: Analyze the epidemiological profile of pertussis cases in children born four years of age, retrospective to the implantation of the dTpa vacine in pregnant women, in the municipalities of Botucatu and Marília, State of São Paulo. Method: This is a descriptive, comparative and historical series study conducted in the cities of Botucatu and Marília. The data were collected from the National System of Notifiable Diseases (SINAN), in children aged 0 to 4 years old, confirmed with pertussis, in the period from January 1, 2008 to October 31, 2014 in the cities of Botucatu and Marília. Results: It is observed a gradual increase in cases from 2014 on in both cities. The seasonality of confirmed cases in Botucatu occurred in winter, with 42.86% occurrences, and in Marília, in the summer, with a percentage of 35.71%. The age group most affected by the disease was formed by children under one year old in both cities. The analysis of the distribution of confirmed cases by geographic region showed that, in Botucatu, the largest case register occurred in the northern area, with 57.14%, and in the city of Marília, it happened in the south area, with 42.86%. Regarding the vaccination scheme, Botucatu presented vaccination in 57.14% of the children, and in Marília the percentage was 52.38%. In Botucatu, 35.71% of the children presented complications of the disease, and in Marília it was 16.66%, with pneumonia being the most frequent complication. In Botucatu and Marília, 100% and 92,85% of the children evolved to cure respectively.Conclusion: The pentavalent vaccine, which contains the pertussis component, still is an important preventive measure, but is not being effective because of the lower age range than the one recommended by the vaccine in the two cities. Therefore, this study suggests studies that evaluate the epidemiological profile of pertussis after the implementation of preventive measures by the Ministry of Health, such as the dTpa vaccine in pregnant women.

La toxine de pertussis (PTX) est une A-B protéine considérée comme l’un des principaux facteurs de virulence de Bordetella pertussis, l’agent bactérien responsable de la coqueluche. Aujourd’hui, cette maladie représente encore un réel danger pour les nouveaux-nés et les<p>nourrissons non ou partiellement immunisés. Actuellement, la coqueluche provoque encore la<p>mort d’environ 350.000 individus par an. La toxicité de la PTX est liée à l’activité<p>enzymatique de sa sous-unité A capable d’inhiber les voies de signalisation associées aux<p>protéines Gi. La partie B, quant à elle, permet l’entrée de cette sous-unité A dans le<p>cytoplasme des cellules cibles en se liant spécifiquement à son ou ses récepteurs<p>membranaires toujours inconnus de nos jours.<p><p>Des études réalisées chez la souris et chez l’homme ont montré que les vaccins anticoquelucheux combinés à différents antigènes vaccinaux étaient capables de moduler<p>leurs réponses humorales spécifiques. Par ailleurs, la PTX est couramment qualifiée d’agent<p>immunostimulant. En effet, des modèles murins de vaccination permirent d’identifier des<p>propriétés adjuvantes de la PTX coadministrée avec des antigènes non relevants.<p><p>Le travail développé dans ce manuscrit étudie les effets de la PTX sur 2 types cellulaires<p>primordiaux sollicités lors d’une vaccination :la cellule dendritique (DC) et le lymphocyte T<p>CD4+ naïf.<p><p>Les DC sont les seules cellules présentatrices d’antigènes aptes à initier une réponse immune<p>primaire. Dans un premier temps, nous avons montré que la PTX était capable d’activer des<p>DC générées in vitro à partir de monocytes. En effet, elles acquièrent un phénotype mature<p>caractérisé par une augmentation de l’expression membranaire des molécules costimulatrices<p>et du CMH de classe II, démontrant un effet direct et spécifique de la PTX sur les DC<p>myéloïdes. Parallèlement, ces DC produisent du TNF-a, de l’IL-12p40 et de l’IL-12p70 et<p>activent NF-kappaB, un facteur de transcription essentiel au processus de maturation. Nous<p>avons obtenu des résultats similaires avec une toxine génétiquement modifiée qui est<p>enzymatiquement inactive. A partir de sang total incubé avec la PTX, nous avons par ailleurs<p>observé que les DC circulantes du nouveau-né étaient déficientes dans leur maturation et leur<p>sécrétion d’IL-12p70 comparées aux DC de l’adulte.<p><p>D’autre part, il a été décrit précédemment que la PTX exerçait des effets mitogènes sur les<p>lymphocytes T humains et murins. Cependant, le rôle qu’elle joue sur la population des<p>lymphocytes T CD4 naïfs reste peu connu. A l’issue de notre second travail, nous pouvons<p>dès lors affirmer que la PTX est également capable d’activer des lymphocytes T<p>CD4+CD45RA+ naïfs isolés à partir des cellules mononuclées du sang périphérique, et ce<p>indépendamment de son activité enzymatique. En effet, ces lymphocytes T CD4+ naïfs stimulés par la PTX prolifèrent, synthétisent des quantités non négligeables d'ARN messagers<p>codant pour l’IL-2 et le TNF-a, augmentent l’expression membranaire des molécules CD40L,<p>CD69 et CD25 et expriment la protéine Foxp3. Cette activation s’accompagne de la translocation nucléaire de NF-kappaB et NFAT. Parallèlement à l’adulte, la PTX active les lymphocytes T CD4 néonataux. Néanmoins, ceux-ci prolifèrent moins bien et expriment plus faiblement le CD40L à leur surface.<p><p>Enfin, la PTX induit la sécrétion de taux importants d’IFN-g par des T CD4+CD45RA+ naïfs<p>adultes mis en présence de DC autologues.<p><p>Nous terminerons en proposant l’hypothèse suivante :La PTX pourrait exercer ses propriétés<p>adjuvantes par l’intermédiaire de différents mécanismes comprenant notamment la maturation<p>des DC d’origine myéloïde et l’activation des lymphocytes T CD4+CD45RA+ naïfs. Ces 2 populations cellulaires sont en effet les principaux protagonistes impliqués dans la réponse<p>immune primaire. / Doctorat en sciences pharmaceutiques / info:eu-repo/semantics/nonPublished

: Filamentous hemagglutinin (FHA) is adhesive protein molecule that is secreted by Gram- negative bacterium Bordetella pertusis, the causative agent of whooping cough (pertussis). The C-terminal segment of FHA plays a crucial role in host-pathogen interaction, however, the structural features are still unknown. Here, we identified the C-terminal residue of FHA and processed form of FHA (FHA*) as alanine residues in position 2304 and 2228, respectively. Circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy demonstrated that the C-terminal segment of FHA(FHA 1995-2228) is characterized by alpha-helical contribution without any compact protein fold. Moreover, suppression of transcription of small regulatory RNA pairing to the 5'-end of fhaB transcript resulted in two- fold increase of FHA production. These data suggested that the C-terminal segment of FHA appear to be an unstructured protein and FHA secretion is negatively regulated by small regulatory RNA. (In Czech) Keywords: Bordetella pertussis, filamentous hemagglutinin, small RNA

The Gram-negative pathogen bacterium Bordetella pertussis is the infectious agent causing pertussis or whooping cough. The infection is dangerous to infants, often being deadly if untreated. Since whole-cell pertussis vaccines have been replaced by acellular pertussis vaccines, pertussis has become the most prevalent vaccine-preventable disease in developed countries. Therefore, the development of a new generation of pertussis vaccines has become a high priority. Opsonophagocytic assays are one method used to assess the efficacy of new vaccines. The main objective of the thesis is to develop opsonophagocytic killing and uptake assays for the measurement of functional antibody activity against Bordetella pertussis. Neutrophils from mice and humans were isolated by three different methods and used for the assessment of different human and mouse sera in opsonophagocytic killing and uptake assays. Different experimental conditions were tested, including multiplicity of infection and serum dilutions. The opsonophagocytic uptake assay proved to discriminate between naïve and immune sera. Serum from mice vaccinated with the whole-cell pertussis vaccine enhanced opsonophagocytic uptake of B. pertussis cells into neutrophils, while serum from mice immunized with the acellular pertussis vaccine did not....

Pertussis is an infectious disease caused by the Gram-negative bacterium Bordetella pertussis. In Peru, actual public health programs indicate that vaccination against B. pertussis must be mandatory and generalized, besides all detected cases must be reported. The objective of this study was to determine the prevalence of B. pertussis among children under five years of age with a presumptive diagnosis of whopping cough in Cajamarca, a region located in northern Peru. / Background and Objectives: Pertussis is an infectious disease caused by the Gram-negative bacterium Bordetella pertussis. In Peru, actual public health programs indicate that vaccination against B. pertussis must be mandatory and generalized, be-sides all detected cases must be reported. The objective of this study was to determine the prevalence of B. pertussis among children under five years of age with a presumptive diagnosis of whopping cough in Cajamarca, a region located in northern Peru. Materials and Methods: The population of this cross-sectional study were children under 5 years old hospitalized as presumptive cases of pertussis during December 2017 to December 2018. The nasopharyngeal samples were analyzed by real-time PCR for the detection of B. pertussis. Results: B. pertussis was identified as PCR + in 42.3% of our sample (33/78). The clinical presentation that was observed most frequently includes paroxysmal coughing (97%), difficulty breathing (69.7%), cyanosis (72.7%) and post-tussive em-esis (60.6%). Additionally, pneumonia was the most observed complication (33.3%). Four of the patients with PCR+ for B. pertussis presented only lymphocytosis, five only leukocytosis, two patients with decreased leukocytosis and lymphocytes and only one patient with leukopenia and relative lymphocytosis. There was a percentage of 84.8% of unvaccinated children in the PCR+ group. Finally, the mother was the most frequent symptom carrier (18.2%). Conclusion: In conclusion, in the studied population there is a high rate of PCR+ cases for B. pertussis. Laboratory values may show leukopenia or lymphopenia in patients with pertussis. It is necessary to use appropriate laboratory diagnostic tests in all infants with respiratory symptoms for B. pertussis. Since, the clinical diagnosis overestimates the diagnosis of pertussis. / Revisión por pares

Background: Acute respiratory infections (ARIs) represent an important cause of morbidity and mortality in children, remaining a major public health concern, especially affecting children under 5 years old from low-income countries. Unfortunately, information regarding their epidemiology is still limited in Peru. Methods: A secondary data analysis was performed from a previous cross-sectional study conducted in children with a probable diagnosis of Pertussis from January 2010 to July 2012. All samples were analyzed via Polymerase Chain Reaction (PCR) for the following etiologies: Influenza-A, Influenza-B, RSV-A, RSV-B, Adenovirus, Parainfluenza 1 virus, Parainfluenza 2 virus, Parainfluenza 3 virus, Mycoplasma pneumoniae and Chlamydia pneumoniae. Results: A total of 288 patients were included. The most common pathogen isolated was Adenovirus (49%), followed by Bordetella pertussis (41%) from our previous investigation, the most prevelant microorganisms were Mycoplasma pneumonia (26%) and Influenza-B (19.8%). Coinfections were reported in 58% of samples and the most common association was found between B. pertussis and Adenovirus (12.2%). Conclusions: There was a high prevalence of Adenovirus, Mycoplasma pneumoniae and other etiologies in patients with a probable diagnosis of pertussis. Despite the presence of persistent cough lasting at least two weeks and other clinical characteristics highly suspicious of pertussis, secondary etiologies should be considered in children under 5 years-old in order to give a proper treatment. / Revisión por pares

Objective: This study aimed to determine the prevalence of A. baumannii in children aged less than 1 year admitted with a clinical diagnosis of whooping cough. Results: A total of 225 nasopharyngeal samples from children under 1 year old hospitalized with clinical diagnosis of whooping cough were studied from January 2010 to July 2012. The presence of A. baumannii was detected in 20.89% (47/225) of the nasopharyngeal swab samples. Among the 47 patients with A. baumannii: 5 were diagnosed with A. baumannii monoinfection, 17 co-infection with bacteria, 7 co-infection with virus and 18 co-infection with bacteria + virus. It was observed that 51.6% (116/225) were children between 29 days and 3 months old, this same group had the highest overall prevalence with 53.3%. The most common co-infecting pathogens were Bordetella pertussis in 55.3%, Adenovirus in 42.6% and Mycoplasma pneumoniae in 23.4%. / Revisión por pares