Rozprawy doktorskie na temat „Vaccination”

Bovine tuberculosis (TB), caused by Mycobacterium bovis, is increasing in incidence in the United Kingdom and detailed knowledge of host-pathogen interactions in the natural host is essential to facilitate disease control. Vaccination of neonatal calves with Bacille Calmette Guerin (BCG) induces a significant level of protection from infection with M. bovis. Since neonatal vaccination of humans with BCG induces activation of natural killer (NK) cells, and neonatal calves have high circulating numbers of these cells, it is proposed that NK cells are important in the response to BCG. Furthermore, NK cells play an important role in shaping adaptive immune responses through interactions with dendritic cells (DCs). The overall hypothesis of this project was that the enhanced efficacy of BCG in neonates is due to the increased number of NK cells, which through interactions with DCs can polarise Th1-type CD4+ and CD8+ T cell responses, both of which are involved in protection against M. bovis infection. Initially, the frequency and phenotype of NK cells across the blood, afferent lymph and the lymph nodes in steady-state conditions were compared. CD2- NK cells were the principal subset of NK cells migrating from the skin to the draining lymph node and were highly activated in afferent lymph and lymph nodes, compared with peripheral blood. It was also demonstrated that CD2- NK cells were the main subset of NK cells egressing from the lymph node via the efferent lymphatic vessel to return to circulation. Since many vaccines including BCG are delivered subcutaneously, NK cell responses in the blood and the skin draining afferent lymphatic vessel, lymph nodes and efferent lymphatic vessel were determined after BCG vaccination. Alterations in the frequency and receptor repertoire were evident following vaccination, supporting a role for NK cells during BCG vaccination of neonatal calves. To investigate the interactions of NK cells and BCG-infected DCs, in vitro co-cultures were established. CD2- NK cells were preferentially activated following culture with BCG-infected DCs and secreted high levels of IFN-γ. Overall, this thesis provides novel evidence that NK cells may re-circulate in steady-state conditions, play a role in BCG vaccination of neonatal calves, and that through interactions with BCG-infected DCs, may be involved in driving protective Th1-type adaptive immune responses.

Infectious disease is an important issue for animal breeders, farmers and governments. Solutions to control infectious disease are needed and research focused on the genetic loci determining variation in immune-related traits has the potential to deliver solutions. The primary aim of this thesis is to discover regions of the bovine genome which influence the immune response post immunisation. To accomplish this two types of immunising agents, a Foot-and-Mouth Disease Virus (FMDV) peptide (FMDV15) and a commercial vaccine for Bovine Respiratory Syncytial Virus (BRSV), were used to immunise the second generation (F2 and backcrosses) of the Roslin Bovine Genome (RoBoGen) herd, a Charolais Holstein cross population. The FMDV15 peptide consisted of two sections of the VP1 protein located on the FMDV capsid, together encompassing the major neutralising antibody sites that are known to be immunogenic. Protection against FMDV is generally believed to relate to the levels of neutralising antibody and has been correlated with IgG1 and IgG2 levels as well as interferon- . In addition it has been shown that T cell responses also play a role in protection against FMDV. Thus all of these were used as phenotypic measurements post immunisation to the FMDV15 peptide. The BRSV vaccine used was an attenuated live vaccine. Protective mechanisms against BRSV infection include IgA, IgG1, IgG2 and IgM BRSV-specific antibodies and antibody titres particularly those of the IgG isotypes are considered to be correlates of protection. Thus, IgG1 and IgG2 antibody levels were measured post vaccination with the BRSV vaccine. All phenotypes were measured across time, and allowed analysis of the primary and secondary adaptive immune responses. Both agents caused considerable variation in the phenotypes measured post immunisation, with significant responses detected two weeks post immunisation. REstricted Maximum Likelihood (REML) analysis attributed much of this variation to sire, highlighting the heritable component, and environmental effects. Significant positive correlations were detected across time within each trait for both the FMDV and BRSV responses. The FMDV and BRSV antibody levels also correlated with each other at later time points, suggesting that there may be animals which are genetically predisposed to be high or low responders in general. Initially a linkage mappingapproach was followed using 165 microsatellite markers, which detected 77 QTL in response to the FMDV peptide and 27 QTL in response to the BRSV vaccine. There were some overlapping QTL, for example QTL which spanned the Major Histocompatibility Complex. Further analysis was conducted by developing a Perl scripted program which genotyped the RoBoGen herd in two ways; 1) Single Nucleotide Polymorphism(s) (SNP) were genotyped within the confidence intervals of the previously discovered QTL and 2) SNP were genotyped via a candidate gene approach. Association study methodology, accounting for relationship stratification via principal components of the genetic relationship matrix, was used to detect significant SNP, in response to both the FMDV peptide and the BRSV vaccine. Twenty significant SNP associations were discovered across 19 traits, with some SNP located in genes with known biological relevance to an immune response, such as the Toll-Like Receptors (TLR), TLR4 and TLR8. This thesis has detected regions of the genome which are significantly associated with the immune responses elicited by two different agents, suggesting similar pathway(s)/gene(s) may be used in defence of multiple pathogens. Once regions of significance were detected, further analysis using SNP markers identified significant, non-synonymous SNP that were associated with the immunising agents. The novel markers discovered in this study may aid breeding for resistance to disease via marker assisted selection. In addition, they may also have highlighted new targets for vaccinologists to develop ‘next generation’ vaccines.

This thesis investigated the effects of acute eccentric exercise on the immune response to vaccination in young humans. Study one investigated whether the efficacy of the eccentric exercise intervention was affected by manipulating the timings of exercise prior to influenza vaccination. Three exercise groups were vaccinated immediately, 6 hr or 48 hr after exercise and antibody responses at 28 days post-vaccination were compared to those from a resting control group. All participants exhibited robust antibody responses to the vaccine and no effect of exercise was observed; therefore, it was not possible to determine the effects of exercise timing on vaccine responses. Study two investigated whether the antibody response to influenza vaccination was influenced by the intensity of eccentric exercise. Three groups exercised at an intensity eliciting 60%, 85% or 110% of one repetition maximum, and the antibody responses at 28 days post-vaccination were compared to those from a resting control group. In the exercise groups, both men and women showed enhanced antibody responses against the B/Florida strain, and men had enhanced responses against A/Uruguay, in comparison to resting controls. In both cases, the control group exhibited poorer responses against these strains, but no effect of exercise intensity was observed. Study three investigated whether the site of vaccine administration affected the efficacy of the immune response to hepatitis B vaccination following eccentric exercise. The antibody seroconversion rate to the vaccine was low (approx. 5 %), and thus, further analysis between exercise and control participants was not feasible. In sum, supporting previous research, it appears that acute eccentric exercise can enhance the immune response to poorly immunogenic strains of influenza, but research is needed to establish if exercise can enhance other poorly immunogenic vaccines, or vaccine responses in the immuno-compromised.

Human subjects maintain long-term immunological memory against infective organisms but the mechanism is unclear. CD4+ T helper memory cells (Thmem) are pivotal in controlling humoral and cellular responses, therefore their longevity and response to vaccination are critical for maintenance of protective immunity. To probe the dynamics of the Thmem response to antigenic challenge, we investigated subjects following a booster injection with tetanus toxoid (TT). Expansion of TT-specific Thmem cells, and cytokine production, showed complex kinetics. Strikingly, parallel expansion and cytokine production occurred in pre-existing Thmem cells specific for two other common antigens, Purified Protein Derivative of tuberculin (PPD), and Candida albicans (C.alb). Bystander expansion occurred in Thmem but not in Thnaive cells. Antibody production against TT peaked ~2 weeks post-vaccination and gradually declined. However, pre-existing antibody against the other antigens did not change. It appears that, although all Thmem cells are readily stimulated to expand, antibody responses are controlled by antigen availability. These human findings which relate to the maintenance of memory and have consequences for assessments of specific T-cell responses to vaccination, have been further investigated in a mouse model. A transgenic model (OT-II) where CD4+ T cells express a TCR specific for an ovalbumin peptide (peptide 323-339, OVAp) was used first to ask the question as to whether naïve or antigen-activated T cells were influenced in a bystander manner during a secondary immune response directed against a protein antigen that was unrelated to their cognate one. For this, carboxyfluorescein diacetate succunimidyl ester (CFSE)-labeled OT-II cells were adoptively transferred, either as naïve or 4 following in vitro activation with OVAp, into C57/BL6 wild type recipient mice which were immune to TT. Recipient mice were then challenged with TT antigen and susceptibility of OT-II cells to bystander activation and proliferation was tested. Naïve T cells were found not to be influenced, but antigen-activated cells were responsive and underwent further activation and bystander proliferation, with accompanying phenotypic changes. Interestingly bystander proliferation appeared to be proportional to the strength of TT-specific cellular immune response. The second question was whether the bystander influence on activated T cells was also evident during a primary immune response to TT. To address this question, antigen-activated OT-II cells and control naïve cells were adoptively transferred into wild type naïve recipient mice and their activation and proliferation was assessed after challenge with TT. In this case no bystander activation or proliferation of OT-II cells was observed. These results underline the susceptibility to bystander activation and proliferation as a unique feature of antigen-activated OT-II cells as opposed to naïve OT-II cells. They mirror those obtained in our study on human subjects and add formal proof of bystander proliferation occurring in vivo. Furthermore this well defined mouse model paves the way for further investigations aimed at addressing the mechanisms responsible for the observed phenomenon.

Syften med uppsatsen var att ta reda på hur folk som är en del av anti-vaccinationsrörelsen kommunicerar i sociala medier. Uppsatsen undersöker hur de kommunicerar i inlägg och kommentarer i en sluten grupp på Facebook. Med hjälp av ett kodschema så blev 97 inlägg analyserade och placerad i olika kategorier med olika variabler. Utifrån kodschemat blev flera tabeller skapade som visade den mest relevanta faktan. Utifrån den information, och med flera detaljerat beskriva exempel-inlägg, beskrivs den generella stämningen i gruppen och hur de talar med varandra. Resultatet blev att de kommunicerar med varandra på ett vänligt och stöttande sätt, bidrar med relevant information när det frågas efter, men med lite fientlighet visas mot de som kommer med motsatta åsikter. Gruppen var sluten, vilket innebär att bara individer som delar samma åsikter som dem är medverkande på plattformen, vilket också betyder att åsikter blir så gott som aldrig utmanade.
The purpose of this essay has been to figure out how people that are a part of the anti-vaccination movement communicate in social media. The essay explores how they communicate in posts and comments in a closed group on Facebook. With the help of a coding scheme, 97 posts became analysed and placed in different categories with different variables. A couple of tables were created from the code scheme that demonstrated the most relevant facts. From that information, and with several detailed descriptions of example posts, describes the general mode in the group and how they speak with each other. The result was that they communicate with each other in a nice and supporting way, contribute with relevant information when it was asked for, but with some hostility shown towards those who come with opposite views. The group was closed, which means that only individuals who share the same views as them are involved with the platform, which also means that opinions are almost never challenged.

A Thesis submitted to The University of Arizona College of Medicine - Phoenix in partial fulfillment of the requirements for the Degree of Doctor of Medicine.
Introduction: Thousands of people die from influenza or its complications each year despite the fact that it is one of the few vaccine preventable diseases. Immunocompromised cancer patients are among the most vulnerable to this infection and flu‐related complications, and therefore vaccination is highly recommended in these patients; however, current vaccination rates and attitudes towards vaccination remain unknown. We hypothesize that immunization rates are lower than the 100% recommendation rate, and hope to understand the reasoning behind the discrepancy. The purpose of this study is to assess cancer patient attitudes towards influenza vaccination in an effort to minimize barriers to vaccination and eventually increase vaccination rates in this immunocompromised population. Methods: Cancer patients enrolled in phase I clinical oncology trials at the Virginia G Piper Cancer Center at Scottsdale Healthcare were invited to participate in a voluntary survey. The 15‐item survey consisted of demographic information, knowledge regarding the flu vaccine, vaccination status after cancer diagnosis and while on treatment, and general attitudes towards vaccination. A total of 84 cancer patients completed the survey. Results were stratified by age, gender, education level, and vaccination status. As this was a descriptive study, no statistical analyses were performed. Results: A total of 84 (n=84) advanced cancer patients enrolled in phase I clinical oncology trials completed the survey. Results indicate that although 71% of patients received the vaccine prior to cancer diagnosis, only 58% of patients have received the vaccine since their cancer diagnosis, and only 48% have been vaccinated while on cancer treatment. Of those vaccinated since cancer diagnosis, 94% reported doctor recommendation of the vaccine and most vaccinate to protect themselves from the virus. Of those not vaccinated since cancer diagnosis, only 37% report their doctor recommends the vaccine and the majority avoid vaccination because they believe the vaccine can cause the flu, they do not feel at risk of infection, and they do not believe the vaccine is effective. Conclusion: Our findings suggest that although the CDC strongly recommends influenza vaccination in cancer patients due to the risk of secondary complications and even death in these immunocompromised individuals, vaccination rates remain low. Our data demonstrates that patients who receive a doctor recommendation for the vaccine are more likely to be vaccinated, but not all doctors recommend the vaccine. Furthermore, false information regarding the vaccine, its efficacy, and its ability to cause infection continues to deter patients from vaccination. Together, this information offers profound insight into the cancer patient population and suggests the need for increased physician and patient education regarding the benefits of annual influenza vaccination to improve vaccination rates and decrease influenza infection and complications in the future.

Vaccination is the most effective method of preventing the spread of infectious diseases. In this thesis, we develop and apply mathematical models to study vaccination. The thesis consists of three main parts. Firstly, in deciding whether to be vaccinated before the outbreak of the epidemic, people need to consider the risk from vaccination and the probability of being infected. The decision of one individual is indirectly influenced by the decisions of all other individuals. Because the vaccine coverage levels are determined by all decisions of all individuals. We apply game theory to this scenario, to predict the expected vaccine coverage level. We construct and analyze four vaccination games. The novelty of this part of work is we introducing the replicator equations to describe the evolutionary processes. For all games, we are able to predict the vaccine coverage levels accurately. Secondly, In order to prove the uniqueness of Nash equilibrium in games, it is crucial to prove the attack ratios are decreasing functions of the vaccine coverage levels. We are able to obtain complete results for the cases of homogeneous mixing population. Only partial results are obtained for the case of heterogeneous mixing population. Thirdly, some insights into the dynamics of malaria infection are obtained. We propose two new malaria models. For delayed malaria transmission models, we calculate the basic reproduction number ℛ₀, the disease-free equilibrium and the possible endemic equilibrium. We also analyze the stabilities of the equilibria. For malaria vaccination model, we calculate the control reproduction number ℛc and the disease-free equilibrium. The threshold of eradication is discussed. We find that, under certain circumstances, the disease of malaria can be eradicated.
Science, Faculty of
Mathematics, Department of
Graduate

Invasive amebiasis, caused by the protozoan parasite Entamoeba histolytica, is one of the leading parasitic causes of mortality worldwide, and there are no vaccines available to control the disease. The heavy subunit of the E. histolytica Gal-lectin is regarded as a potential subunit vaccine candidate. A Th1 (cell-mediated) immune response is protective against invasive amebiasis, and DNA vaccination is a strategy to induce such a response against specific antigens. The objective of this study was to construct and test a Gal-lectin-based DNA vaccine against E. histolytica. DNA encoding as 894--1081 of the Gal-lectin heavy subunit was resynthesized using a gerbil codon frequency bias and inserted in a mammalian expression vector to generate the DNA vaccine pCISToGL6. Balb/c mice vaccinated intradermally developed a Gal-lectin-specific cellular immune response, as well as an anti-Gal-lectin humoral immune response. Serum antibodies recognized a recombinant portion of the Gal-lectin heavy subunit by immunoblot and ELISA, and bound to native Gal-lectin on the surface of live trophozoites, inhibiting adherence to target cells. The Gal-lectin-specific serum antibodies were of the IgG2a isotype, indicating that a Th1 response was stimulated by the vaccine. We were also interested in using DNA encoding IL-12, IL-18 or GM-CSF as genetic adjuvants co-injected with pCISToGL6 to potentiate the immune response. Since the DNA vaccine was destined to confer protection in the gerbil model of invasive amebiasis, we cloned gerbil IL-12 (p35 and p40), IL-18 and its convertase caspase-1, and GM-CSF. The proteins were expressed in mammalian cells and showed bioactivity in vitro. Taken together, these results have laid the foundation to optimize and test a working Gal-lectin with co-stimulatory molecules to elicit a Th1 immune response for protective immunity against invasive amebiasis.

Invasive meningococcal disease (IMD) is caused by Neisseria meningitidis; a Gram-negative, aerobic encapsulated diplococcus that can cause large scale meningitis outbreaks. The risk of meningococcal carriage and disease is higher among particular age groups, certain community settings and travellers. The prevalence of invasive meningococcal disease peaks during infancy due to lack of immunity and again, to a lesser extent, in late adolescence/early adulthood due to increased social mixing. Closed and semi-closed community settings are also associated with high rates of meningococcal transmission and carriage acquisition and have experienced recurrent outbreaks of IMD. Travelling, including pilgrimage to Hajj, is yet another vulnerable circumstance. Despite advanced intensive care support, about 20% of survivors are left with long-term sequelae following IMD, thus prevention by vaccination is the most practical and effective measure of reducing the morbidity and mortality associated with IMD. The use of polysaccharide and then conjugate vaccines against specific meningococcal serogroups causing outbreaks led to a dramatic decline in IMD incidence. However, the observation of waning of immune responses over time following early childhood vaccination remains a concern for resurgence of disease in adolescence. Additionally, issues such as quantifying the effect of various meningococcal vaccines on carriage and the immune interaction between the carrier protein components of meningococcal conjugate vaccines and other vaccines containing the same proteins as antigens remain outstanding. To this end, the purpose of this thesis is to explore ways to better protect vulnerable populations, focusing on vaccination coverage in settings where a mandatory vaccination policy is in place, understanding the effect of vaccination on meningococcal carriage, and interpretation of the immune interactions between vaccines and changes to the immune response over time following vaccination. This thesis considers the annual Hajj pilgrimage as an exceptional context to achieve its aims as it comprises two groups of at-risk individuals: travellers and those within a closed population, and pilgrims often receive multiple vaccines during their preparation to the Hajj journey including meningococcal vaccine. Meningococcal vaccination is mandatory for domestic Hajj pilgrims and healthcare workers; however, their uptake has not been thoughtfully assessed. Hence, we evaluated the meningococcal vaccine coverage among these two groups and explored possible influencing factors. The uptake was suboptimal among both groups; gender, education, employment, receiving pre-Hajj health advice and distance of travel were important influencing factors and lack of awareness was the main barrier. One possible way to compensate for the suboptimal vaccination coverage is to assess and, if possible, limit meningococcal carriage. Therefore, a randomised controlled trial was conducted primarily to explore if meningococcal conjugate vaccines are better than their polysaccharide counterparts in reducing nasopharyngeal carriage among Hajj pilgrims. Actually, among the 1146 participants, the carriage of meningococci was almost non-existent. This may be suggestive of a successful vaccination policy or, more likely, a low season of meningococcal carriage. Hajj pilgrims, as travellers, are often required to receive multiple vaccinations within a short time period. These vaccines may include conjugate vaccines that could interact with other diphtheria-tetanus containing vaccines. Thus, the immune response of Neisseria meningitidis serogroup W (MenW) to the quadrivalent meningococcal conjugate vaccine (conjugated to cross-reacting material 197 (CRM197)) one month after being administered concurrently with, or 3-4 weeks prior to, or following combination tetanus, diphtheria and acellular pertussis vaccine (Tdap) was evaluated in a randomised controlled trial among Hajj pilgrims. The trial found that concurrent or sequential administration of Tdap and Neisseria meningitidis serogroups A, C, W and Y (MenACWY) CRM197 conjugate vaccine did not have a significant effect on the MenW immune response. While gradual waning of immune response, particularly following early childhood immunisations have been observed extensively and investigated, however, an exceptional phenomenon of a natural rise in immune responses has also been noted but not previously investigated. Thus, a secondary analysis of available data on changes over several years to the immune response to meningococcal serogroup C conjugate vaccines among children vaccinated in early childhood was undertaken. This analysis found that a substantial minority (~15%) of children had a rise in their bactericidal antibody titers in the absence of a booster dose of vaccine. This may be attributed to a potential carriage-induced booster response and hence raises concerns that herd immunity is not as well-maintained as previously thought.

Thesis (M.A.)--Boston University PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you.
Human papillomavirus (HPV) is the most common sexually transmitted infection in the United States, and in women, infection by the virus is a major risk factor for developing cervical cancer. In 2006 the U.S. Food and Drug Administration licensed the first prophylactic HPV vaccine for use, a quadrivalent formula aimed at the prevention of cervical cancer and its precursors, vaginal and vulvar cancer and its precursors and anogenital warts. Large-scale studies have shown the vaccine to be both safe and effective. Subsequently, many countries adopted a nationwide vaccine program to ensure all girls, usually around the age of 11 or 12, are vaccinated. Such programs have been largely successful and coverage rates of up to 77% have been reported for the three-dose vaccine series. However, HPV vaccine uptake rates in the United States are meager at best. This study aims to elucidate some of the reasons why adolescent girls in the United States are not receiving the HPV vaccine. Pediatric providers at an urban community health center were asked to fill out a survey that assessed their general vaccine knowledge, current HPV vaccination practices, barriers and attitudes toward vaccination and office practices. Due to the strong influence of provider recommendation of the HPV vaccine on uptake rates, and the association between provider knowledge and likelihood of recommending the vaccine, providers should become more familiar with current guidelines for administering the HPV vaccine. Furthermore, it is important that providers educate patients and their parents about the proven safety and efficacy of the vaccine. With time, these measures alone may increase HPV immunization rates in the United States without the need for an HPV vaccine mandate.

Influenza is a potentially serious disease, causing hospitalisations and deaths, particularly in children and the elderly. Traditionally immunisation efforts targeted the elderly and those at high risk from the disease. As awareness of the paediatric burden of influenza disease and potential population benefits of vaccinating children grew, immunisation efforts expanded, but coverage among children remains low. Expansion of influenza vaccine funding to include all Australian children is being considered, but there is little information about whether parents accept or support this. This thesis examines preventive health beliefs, knowledge, attitudes and practices of parents, both generally and in relation to influenza and influenza vaccination of children, and explores which information sources parents value, trust and use. Methods include a systematic review, qualitative interviews and a quantitative survey. The timing of two influenza-related health scares – the 2009 influenza A H1N1 pandemic, and the 2010 suspension of influenza vaccination for children under 5 due to high numbers of adverse events – allowed examination of the impact of health scares on parents’ views and information needs. The findings revealed that parental beliefs about health and illness, and decision-making about vaccination risk, are complex, incorporating both scientific and ‘folk’ beliefs and interacting with contextual variables. Parents trust healthcare workers as information sources, and are positively influenced by vaccination recommendations from them. Pre-existing beliefs about influenza and influenza vaccine influence parents’ interpretations of influenza-related health scares. Health scares affect parents’ trust and information needs. This thesis provides practical recommendations for public health practitioners and policymakers to assist in developing appropriate influenza vaccination campaigns and information materials that meet the needs of parents.

Despite the existence of effective vaccines for both diseases, measles and poliomyelitis still cause significant worldwide morbidity and mortality. The live-attenuated measles and inactivated polio vaccines are both given using a standard needle and syringe injection. This method of delivery poses many problems for large-scale vaccination campaigns. Microneedles are micron-scale needles which have the potential to overcome many of these hurdles. In the first study, we showed that the measles vaccine could be successfully incorporated into a solid, metal microneedle system which induced potent neutralizing antibody titers after administration into cotton rats. This response was statistically identical to the same dose delivered using a subcutaneous injection. The second study focused on enhancing the stability of the measles vaccine after drying and long-term storage. Using a new assay developed from a measles virus variant engineered to encode for green fluorescent protein, it was determined that a combination of sucrose and threonine provided the highest stabilizing effect. Vaccine mixed with this solution retained more than 90% of its activity after 6 months of storage at 4°C and 25°C. The third study involved the incorporation of the measles vaccine into a dissolving microneedle patch. These patches were used to vaccinate rhesus macaques and the immune response was found to be statistically identical to the same dose delivered by syringe injection. Furthermore, after creation and storage, these patches retained 100% of their infectivity after 2 months at 4°C and 25°C. The final study attempted to create a dissolving microneedle patch containing a full dose of the inactivated polio vaccine. These patches were then used to deliver a full dose of IPV into the skin of a rhesus macaque. This delivery method produced neutralizing antibody titers to IPV type 1 and 2 that were statistically identical to the same dose delivered using a needle and syringe. Overall, these studies show that the microneedle patch was a safe, simple and effective method for measles and polio vaccination. This delivery platform has the potential to overcome many of the hurdles that currently stand in the way of measles elimination and polio eradication.

Background: Following the publication of Andrew Wakefield’s article claiming a link between Autism and the MMR vaccine in 1998, the U.K. and U.S. experienced a decline in vaccination rates. Combating the anti-vaccine messages highlighted by the media are the medical providers, who are consistently reported as an influential source of information for parental vaccine decision making. Despite efforts of the medical and public health community, some developed countries have seen a resurgence of vaccine preventable diseases. Purpose: This study seeks to examine parental vaccination concern in a private pediatric practice in metropolitan Atlanta. Methods: A questionnaire was created by the PI to assess parental vaccination concerns, including items to assess parental feelings toward the providers and nurses regarding preventative care. Data was analyzed in SPSS version 19.0. The study was approved by the IRB at Georgia State University. Results: A total of 283 participant responses were included in the sample. Overall vaccine adherence was 96.1% (272). However, a large minority of participants who were considered to have vaccine concerns were identified: 40.3% (114) of participants responded yes to at least one vaccine hesitation item. Conclusion: Vaccine adherence in a private pediatric practice remains high. However some parents continue to have vaccination concerns and may be at risk for deviating from the vaccine schedule. Using qualitative methods to obtain parental beliefs may provide a deeper understanding of parental decisions to aid in the development of public health education programs. The feasibility of collecting data at a private pediatric practice is discussed.

DNA vaccination is based on in vivo delivery of plasmids encoding an antigen, leading to antigen synthesis and specific immunity. This technology is still in its infancy but has advantages over conventional vaccination, such as the stimulation of all arms of the immune response, i.e., humoral immunity, T-helper (17h) cells and cytotoxic T lymphocytes (CTLs). We studied DNA vaccination against human carcinoembryonic antigen (CEA) in mice. Our hypothesis was that codelivery of cytokine- and CEA-encoding plasmids could regulate responses, and allow polarization of Th responses to either type 1 (Th1) or type 2 (Th2). We found that intramuscular injection of the CEA plasmid alone induced antibodies, Th1 cells and CTLs reactive to CEA. These mice had increased immunity against transplanted syngeneic CEA+ stably transfected tumor cell lines, but always developed lethal tumors. Coinjection of the CEA plasmid with a vector encoding either IL-12 or interferon gamma (IFNgamma) markedly enhanced IgG2a production (IFNgamma-dependent), IFNgamma secretion by spleen cells (a Th1 cytokine) and CTL-mediated tumor cell lysis, in a CEA-specific way. Moreover, resistance to a tumor challenge was greatly improved, such that up to 80% of mice survived tumor free. In contrast, coinjection of CEA and IL-4 genes increased CEA-specific IgG1 levels (IL-4-dependent) and IL-4 secretion by lymphocytes (a Th2 cytokine), but decreased both CTL activity and tumor resistance. Thus, we could readily enhance or polarize immunity. The IL-12 cDNA had the strongest adjuvant effect, which was only observed when it was injected at the same site as the CEA gene. To analyze effector components, we studied IL-12-plasmid-enhanced DNA vaccination in gene knockout mice, lacking either CD3, CD4, CD8. IFNgamma, perforin or Fas ligand (FasL). Only mice expressing all of CD3, CD4, IFNgamma, CD8 and perforin, and inoculated with both the CEA and IL-12 genes, could fully resist a tumor challenge. The Fas/FasL lytic

Thesis (Ph. D.)--Massachusetts Institute of Technology, Sloan School of Management, Operations Research Center, 2008.
Includes bibliographical references (p. 125-129).
Annual influenza outbreaks incur great expenses in both human and monetary terms, and billions of dollars are being allocated for influenza pandemic preparedness in an attempt to avert even greater potential losses. Vaccination is a primary weapon for fighting influenza outbreaks. The influenza vaccine supply chain has characteristics that resemble the Newsvendor problem, but possesses several characteristics that distinguish it from many other supply chains. Differences include a nonlinear value of sales (caused by the nonlinear health benefits of vaccination that are due to infection dynamics) and vaccine production yield issues. In this thesis we present two models in the interface of operations and supply chain management and public health policy. In the first model, we focus on a supply chain with a government and a manufacturer. We show that production risks, taken currently by the vaccine manufacturer, lead to an insufficient supply of vaccine. Several supply contracts that coordinate buyer (governmental public health service) and supplier (vaccine manufacturer) incentives in many other industrial supply chains can not fully coordinate the influenza vaccine supply chain. We design a variant of the cost sharing contract and show that it provides incentives to both parties so that the supply chain achieves global optimization and hence improves the supply of vaccines. In the second mode, we consider the influenza vaccine supply chain with multiple countries.
(cont.) Each government purchases and administers vaccines in order to achieve an efficient cost-benefit tradeoff. Typically different countries have different economics sensitivities to public outcomes of infection and vaccination. It turns out that the initiating country, while having a significant role in the spread of the disease, does not receive enough vaccine stockpiles. Our model indicates that lack of coordination results in vaccine shortfalls in the most needed countries and vaccine excess in the regions where are not as effective, if the governments in the model act rationally. We show the role of contracts to modify monetary flows that purchase vaccination programs, and therefore modify infectious disease flows.
by Hamed Mamani.
Ph.D.

Staphylococcus aureus est une bactérie qui possède un potentiel virulent très élevé. Elle cause plusieurs types d’infections telle que des septicémies, des endocardites, et des mammites. La mammite bovine à Staphylococcus aureus est une infection très affligeante pour les animaux et les producteurs laitiers touchés. Ses nombreux facteurs de virulence permettent à la bactérie d’infecter les glandes mammaires des animaux en lactation. Il n’existe toujours pas de vaccins efficaces pour combattre ou prévenir les infections à Staphylococcus aureus. Il est donc primordial de trouver de nouvelles approches de vaccination. La vaccination à l’ADN apporte un nouvel espoir de vaccination bénéfique pour le combat de ces infections. Ce projet avait pour objectif de tester, dans un modèle murin, l’efficacité de certains antigènes à induire chez la souris une réponse immunitaire efficace et protectrice contre Staphylococcus aureus. Nous avons choisi des antigènes impliqués dans la virulence de la bactérie en supposant que ces derniers nous permettent d’avoir un vaccin génique efficace contre Staphylococcus aureus. Nous avons donc construit des plasmides pour les vaccins contenant nos gènes d’intérêt soit la séquence codant pour la Sortase et le peptide auto-inducteur, deux protéines impliquées dans la virulence de la bactérie. Deux autres antigènes ont été inclus dans nos expérimentations. Ces antigènes sont le « dumping factor A » (Clfa) et la « fibronectin-binding-protein A », deux adhésines liant le fibrinogène et la fibronectine respectivement. Nous avons pu obtenir une réponse humorale significative contre les antigènes Clfa, la Sortase et le peptide autoinducteur et une réponse cellulaire significative contre l’antigène Clfa. Un effet de protection statistiquement significatif contre une infection expérimentale avec un vaccin contenant quatre plasmides codant pour chacun des antigènes mentionnés plus haut a également été observé. Ces résultats suggèrent de poursuivre les recherches avec ces antigènes.

Childhood immunisation effectively protects personal and public health, but a sizeable minority of parents actively reject vaccines for their children. This Thesis explores how parents decide whether to have their children immunised, in order to inform efforts to improve immunisation uptake. A consistent profile of beliefs relating to vaccine rejection emerged across a systematic review of existing evidence, a semi-structured interview study (n=24), and two evidence-based questionnaire studies (n=900), which all focused on the measles, mumps and rubella (MMR) vaccine. Vaccine-rejecting parents doubted vaccine safety and efficacy, mistrusted health professionals and immunisation policy, perceived ‘pro-vaccine’ bias in most available information, believed most vaccine-preventable diseases are mild and uncommon, and were not motivated by the potential social benefits of MMR uptake. The review also indicated several pervasive methodological flaws in the evidence – including retrospective designs, parent-reported outcomes, and lack of multifactorial models – which were remedied in the new empirical work. Three behavioural experiments (n=703) were then used to explore the influence of this belief profile on immunisation decision-making under controlled conditions. These experiments indicated that the belief profile was less influential when decision-makers mistrusted vaccine providers and policy, were generally anxious, or sought to assimilate multiple belief profile factors on the basis of limited information. They also showed that information processing limitations and biases may influence decisions independently of information content, and accordingly written risk communication method was found in the final study (n=42) to impact on outcomes even after adjusting for information content. These findings indicate that parents’ immunisation decisions are typically not borne of simple vaccine versus disease risk-benefit analyses. Interventions to increase trust in the immunisation system and to communicate the social desirability (and normality) of vaccine acceptance may improve immunisation uptake and support informed, satisfying decision-making.

Schools are group settings where vaccine-preventable diseases can spread quickly, especially if vaccination rates are suboptimal. Vaccination of school children has been the subject of many studies; however, data are lacking regarding the vaccination status, vaccination perceptions, and potential barriers to vaccination for school employees. Method: A questionnaire was developed to measure school employees' perceptions,awareness of current vaccination status, and potential barriers to vaccinations. This study included a convenience sample of 277 employees from a small urban school district located in central Utah. Results: Adult vaccination knowledge is lacking in the school employee population, with over half believing they were fully vaccinated even though 57.8% had not had an influenza vaccination this season. Many school employees were unaware of their vaccination status for highly virulent diseases such as measles and pertussis. In addition, most subjects believed vaccinations were safe and effective, although they believed vaccinations were more important for children than adults. Almost half of respondents believed vaccine mandates should exist for school employees. Conclusion: Knowledge gaps regarding adult vaccines can be positively influenced by nurses, especially school nurses. These knowledge gaps may be especially important to bridge concerning adults working in the school setting, an environment ideal for the spreading of communicable diseases.

Background Influenza causes annual, worldwide epidemics of respiratory disease that affects all segments of the population. Mass vaccination of healthy children, who are playing an important role in the transmission of influenza, is promoted to be a complementary approach in prevention and control of influenza. However, lack of published systemic review evidencing the indirect protection of vaccinating healthy children makes the implementation under uncertainty. Method A systemic review was conducted by computerized bibliographic searches in PubMed and the Cochrane Library identifying the published studies on the effectiveness and cost-effectiveness of vaccinating healthy children to control influenza epidemics by reducing transmission in the community. Any study design with vaccinating healthy children as the intervention versus control group with no influenza vaccine was included. Only outcomes measured on the contacts of children, either the community or household members were considered. Result Twenty-two articles were selected to be reviewed in this project, in which 17 of them covered the public health benefit of vaccinating healthy children to protect others in the community against influenza, and five of them were economic studies. Overall the result suggested that vaccinating health children produces a public health benefit in protecting others in the community against influenza and that it is a cost-effective measure. Discussion Targeting vaccines to healthy children should be promoted for optimal vaccine allocation, maximizing the vaccination effectiveness. Community planning on vaccine delivery infrastructure as well as educational and communicational strategies is necessary to improve influenza vaccine coverage. Further well-designed studies such as RCT with larger sample sizes, as well as studies in Hong Kong or other sub-tropical regions should be carried out and included. Moreover, large and population-based studies should be conducted to examine the overall impact of universal childhood influenza immunization.
published_or_final_version
Public Health
Master
Master of Public Health

As key members of the school environment, it is important for school employees to be vaccinated. Employees are in direct contact with children in close quarters for long periods of time and such an environment can easily serve as an outbreak center for vaccine-preventable communicable diseases such as measles. Despite the fact that most school employees believe vaccines are safe and effective and many school employees report they are up-to-date with their vaccines, a closer examination reveals discrepancy between belief and behavior. As a vaccine advocate, the school nurse can be influential in providing adult vaccination education for school employees, thus increasing awareness of the importance of adult vaccines and knowing one's vaccination status. Additionally, school nurses might need to meet with school district policymakers to promote vaccine mandates for school employees and to assist in the creation of containment plans in the event of a measles outbreak at school.

Includes abstract.
Includes bibliographical references.
Measles virus is known to be one of the most contagious of infectious agents and despite considerable progress towards elimination, a number of Sub-Saharan African countries experienced epidemics in 2009-2011, including South Africa, in which there were over 18 000 confirmed cases. The South African measles vaccination programme started in 1975 with 1 dose schedule, and from 1996-8 has followed the World Health Organization-United Nations Children’s Fund strategy. This includes a 2 dose routine vaccination schedule for children at 9 and 18 months of age, supplementary mass vaccination campaigns (MVCs) for children conducted 4 yearly, improved case management and casebased laboratory surveillance. Administrative monitoring of routine vaccination coverage is problematic, and often overestimated, because of denominator and numerator inaccuracies. The potential for a significant outbreak in the Western Cape Province was therefore not recognized. Over 2000 cases were confirmed in the Western Cape epidemic which began in September 2009 and peaked in March 2010. The Metropole district was mainly affected and over 60% of the cases were under 5 years of age, with 29% aged 6 to 11 months. A MVC, against measles had already been planned; however as a result of the epidemic the targeted age group for measles vaccination was extended from 9 to 59 months, to include children from 6 months to 15 years. This was conducted nationally from 12 to 23 April 2010.

Els virus de la influença tipus A (VIA) són patògens zoonòtics que poden infectar un ampli nombre d’hostes incloent-hi les aus, els porcs i els homes, entre altres. Anualment es documenten milions d’infeccions en humans causades per virus de la influença estacionals. Les pandemies causades pel virus influença també tenen una elevada repercussió pel que fa a la sanitat i l’economia. Tot i que determinats subtipus de VIA s’adapten millor en espècies d’aus que en humans, hi ha hagut casos d’infeccions en humans per virus de la influença de tipus aviars. La susceptibilitat dels porcs per infectar-se amb virus de la influença tant d’origen aviar com humà és també important pel que fa a la salut pública. El genoma del virus influença és segmentat in consta de vuit molècules de ARN de cadena senzilla i sentit negatiu que codifiquen per 11 o 12 proteïnes. Per tant, si una cèl·lula s’infecta simultàniament per dos VIA diferents, pot succeir un reagrupament amb la conseqüent generació d’una nova soca de virus. A més, mutacions a les glicoproteïnes de superfícies (sobretot a l’hemaglutinina, HA) són les responsables de l’elevada variabilitat de VIA. Tot i que les vacunes front a les epidèmies estacionals són eficaces, no produeixen resposta immunològica front una amplia varietat de VIA. És a dir, les vacunes estacionals només protegeixen front a les soques virals circulants durant una determinada estació. Aquest fet, junt amb el risc de possibles pandèmies, han fet encara més important i urgent el desenvolupament d’una vacuna universal capaç de produir immunitat front a múltiples subtipus virals. En la present tesis s’ha estudiat la resposta immunitària front a la infecció i vacunació del VIA en el context de VIA d’alta patogenicitat (vIAAP) A/H5N1 i A/H7N1 i el virus pandèmic A/H1N1 (pH1N1). El treball s’ha dividit en tres parts i cada part s’ha subdividit en capítols. Part I (capítols 1 i 2), conté la introducció general i els objectius de la tesi doctoral. L’objectiu d’aquesta primera part és donar una visió global i introduir informació per entendre (i) la infecció pel virus de la influença, (ii) la resposta immunològica provocada després de la infecció per VIA i (iii) un breu resum de les vacunes actuals front a influença. A continuació, s’exposen els objectius a aconseguir. Part II, és el cos de la tesis i conté els quatre treballs (del capítol 3 al 6) duts a terme durant els quatre anys que ha durat el programa de doctorat. Tots els capítols presentats han estat publicats o sotmesos a publicació en revistes indexades internacionals. Per tant, cada estudi manté l’estructura estàndard de: resum, introducció específica, materials i mètodes, resultats i breu discussió. Estudiar el paper dels determinants virals i caracteritzar la infecció pel VIA en diversos hostes pot ser de gran interès a l’hora de dissenyar vacunes òptimes. S’ha descrit la proteïna NS1 com a un dels principals determinants de virulència en mamífers, però no s’ha estudiat gaire el paper d’aquesta en aus. En el capítol 3 es va avaluar la implicació de la proteïna NS1 en la patogenicitat viral en pollets. Es van infectar pollets amb vIAAP H7N1 que contenien el segment NS de vIAAP H5N1. Les manifestacions patològiques i la resposta immunològica conseqüència de la infecció amb cada un dels virus van ser avaluades. També és molt important el paper de la immunitat prèvia durant un brot perquè pot ser determinant de la mort o supervivència de l’animal. En el capítol 4 es van exposar pollets a un virus H7N2 de baixa patogenicitat (vIABP) i a continuació es van infectar amb un vIAAP H7N1. Posteriorment es van infectar amb un vIAAP H5N1. Els animals que havien estat infectats prèviament amb vIABP quedaven protegits a la posterior infecció letal amb el vIAAP H7N1. No obstant, la resposta immunitària produïda no era suficient per a protegir els pollets front a la infecció amb un virus heterosubtípic (vIAAP H5N1). La presència o absència d’anticossos inhibitoris front a H7- i H5- correlacionaven amb la presència o absència de protecció, respectivament. Conèixer els programes de vacunació actuals i la seva eficàcia és útil per a planificar i dissenyar futures estratègies de vacunació. Les aus aquàtiques són el reservori dels VIA; per tant, són extremadament importants pel que fa a l’ecologia del virus. Aprofitant els programes de vacunació es va testar el sèrum de diverses espècies d’aus de zoològics i centres de recuperació d’Espanya (capítol 5). Els sèrums es van utilitzar per a l’avaluació de la resposta humoral deguda a la vacuna. El principal objectiu del treball era determinar l’eficàcia de vacunes disponibles (inactivades en suspensió oliosa) en diverses espècies d’aus i comparar la variabilitat inter- i intra-espècie. Finalment, i tenint en compte el potencial risc del VIA, els esforços es van focalitzar en desenvolupar una vacuna capaç de protegir a un ampli nombre de subtipus de VIA. La pandèmia de 2009 amb el virus H1N1 (pH1N1) és un clar exemple que els porcs poden actuar com a “coctelera” i generar nous virus. En el capítol 6 es van immunitzar porcs amb pèptids derivats de l’HA i a continuació es van infectar amb el virus pH1N1. Tot i que els pèptids-HA produïen una molt bona resposta humoral i cel·lular, no es va detectar activitat neutralitzant i només es va obtenir un efecte parcial en l’eliminació del virus. Part III (Capítols 7 i 8), és la secció on es discuteixen les implicacions dels resultats obtinguts en els diferents estudis i on s’enumeren les conclusions principals. En una secció a part, s’han inclòs totes les referències bibliogràfiques utilitzades per a l’elaboració de la tesi. S’ha inclòs també un apèndix per afegir informació addicional.
Influenza A viruses (IAV) are zoonotic pathogens that can replicate in a wide range of hosts, including birds, pigs and humans, among others. Millions of human infections caused by seasonal influenza virus are reported annually. Influenza pandemics have also a significant health and economic repercussions. Although certain subtypes of IAV are better selected in avian species than in humans, there are reports that evidence cases of human infections with avian influenza viruses (AIV). The susceptibility of pigs to infection with influenza viruses of both avian and human origins is also important for public health. The genome of influenza virus is segmented and consists of eight single-stranded negative-sense ribonucleic acid (RNA) molecules encoding 11 or 12 proteins. Thus, if a single cell is simultaneously infected by two distinct influenza viruses, a reassortment can occur resulting in the generation of a novel virus strain. Moreover, mutations in the surface glycoproteins (mainly in the hemagglutinin, HA) are the responsible of the high variability of IAV. Influenza vaccines against seasonal epidemics, although have good efficacy do not elicit immune response against a wide variety of IAV. Thus, seasonal vaccines only confer protection against the circulating viral strains. This, together with the risk of potential pandemics, has highlighted the importance of developing a universal vaccine able to elicit heterosubtypic immunity against multiple viral subtypes. In this thesis the immune response to IAV infection and vaccination was evaluated in the light of the risk of highly pathogenic AIV (HPAIV) A/H5N1 and A/H7N1, and the pandemic IAV A/H1N1. The work is divided into three parts and each one is further divided into chapters. Part I (chapters 1 and 2) contains the general introduction and the objectives of the thesis. The aim of this first part is to give a global overview and to introduce information to understand (i) the influenza infection, (ii) the immune responses elicited after IAV infection and (iii) a brief summary of current vaccines against influenza. Afterwards, the initial objectives to be achieved are exposed. Part II is the body of the thesis and it contains four studies (from chapter 3 to 6) developed during the four-year period comprising the PhD program. All the chapters are published or submitted to publish in international peer-reviewed journals. Thus, each study contains an abstract, a specific introduction, the materials and methods section, the obtained results and a discussion. To study the role of IAV determinants and to characterize the influenza infection in different hosts could be of great importance to direct the efforts to the formulation of more efficient vaccines. The non structural 1 (NS1) protein is known to be a major determinant of virulence in mammals but little is known about its role in avian species. In chapter 3, the involvement of NS1 in viral pathogenicity was evaluated in chickens. Birds were challenged with two reassortant AIV carrying the NS-segment of H5N1 HPAIV in the genetic background of an H7N1 HPAIV. The pathological manifestations, together with the immunological outcome were evaluated. The role of pre-existing immunity during an outbreak is also important and can determine whether the animals succumbed to infection or not. In chapter 4, chickens pre-exposed to H7N2 low pathogenic AIV (LPAIV) were challenged with H7N1 HPAIV and subsequently infected with H5N1 HPAIV. Pre-exposed animals were protected against the lethal H7N1-challenge whereas naïve animals succumbed. However, pre-existing immunity did not provide protection against HA-heterosubtypic virus (H5N1 HPAIV). The presence or absence of H7- and H5-inhibitory antibodies correlate with the protection (or lack of it) afforded. The control of current vaccination programs and their efficacy is useful to plan and design better vaccines. It is well known that wildfowl are the reservoirs of IAV; thus they are extremely important concerning the ecology of the virus. Sera from several avian species from Spanish zoos and wildlife centers were collected during two successive vaccination programs and were tested to evaluate the vaccine-elicited humoral response (chapter 5). The main objective of this work was to determine the efficacy of current vaccines (inactivated water-in-oil) in several avian species and to compare the differences inter- and intra-specie. Finally, and taking into account the potential risk that IAV represent to our society, the efforts were focused on developing a broadly protective influenza vaccine. The 2009 human H1N1 pandemic (pH1N1) is a clear example that pigs can act as a vehicle for mixing and generating new assortments of viruses. In chapter 6 pigs were immunized with HA-derived peptides and subsequently infected with pH1N1 virus. Although the HA-peptides induced broad humoral and cellular responses no neutralization activity was detected and only a partial effect on virus clearance was observed. Part III (chapters 7 and 8) is where the implications of all the findings from the studies are discussed and the major conclusions are listed. A list of all the references used to develop the thesis is listed after the three parts, in an independent section. An appendix section is also included to give further information.

Background: Infection with Human Papillomavirus, HPV, is one of the world's most common sexual transmitted infections. The virus causes genital warts (condyloma) but certain virus types can cause cancer. The most common cancer type caused by the virus is cancer of the cervix. Nowadays there is a screening program for women between 23 and 60 years of age where Pap smears of the cervix is taken to discover abnormalities at an early stage. Cervical cancer is very common all over the world and this screening program saves many lives, but not in the less developed countries since the screening program is too expensive. A vaccine has been developed against the four most common HPV- types 6, 11, 16 and 18, and the vaccine costs less than the screening program, so it is now possible to save lives worldwide. The vaccine contains virus-like particles, VLP's, synthesized by L1-proteins from HPV types 6, 11, 16 and 18. HPV 6 and 11 are associated with the sexual transmitted disease condyloma, and HPV 16 and 18 can cause cancer in the cervix among women, but also other kinds of anogenital cancer types. Today, the HPV vaccine is included in the general vaccination program for girls in 5th and 6th grade in Sweden, but not for boys. The risk to get infected with the virus is equal between girls and boys and this has raised the question whether or not boys should be included in the general HPV-vaccination program as well. Purpose: The aim of this study was to evaluate whether or not there is evidence that suggest vaccinating other populations than young girls. Methods: The study is a literature study of clinical trials collected from the database PubMed. Results: Many clinical trials, where the vaccine efficacy among young initially HPV-naive girls have been evaluated, have been performed. They show that the vaccine is effective in preventing HPV-infection in this population. In a population with older participants, that have been sexually active and, thereby, are likely to have been infected with the virus, the vaccine's efficacy is more unknown. Clinical trials have been made in populations like this, with participants with initially unknown HPV-status, and the vaccine shows efficacy against infection with the HPV type, or types, that the individual is not already infected with. Clinical trials have also been made among boys and young men and the vaccine's efficacy is noninferior among men compared with the vaccine efficacy among girls and young women. Conclusions: Those findings support the inclusion of boys in the general vaccination program. They also show that already infected individuals can take benefit from the vaccine since it is not very likely that they are infected with all four vaccine types. But, before a gender neutral vaccination can be recommended, and before vaccination of already infected individuals is suggested, more and larger studies must be made. The new trials should also have longer follow up-times to determine whether a refill vaccination in the future will be needed.

This thesis attempts to apply the theory of rational disease dynamics to the human papillomavirus by testing whether HPV vaccinations are prevalence elastic. A prevalence elastic relationship suggests that HPV vaccination rates respond positively to increasing prevalence rates of cervical cancer. Prevalence rates are measured both by incidence rates of cervical cancer and by mortality rates of cervical cancer. Data from the NIS-Teen Survey as well as data from the United States Cancer Statistics branch of the Centers for Disease Controls are used to construct a linear regression that controls for income, education levels, proxies for social culture, and proxies for physician access. Incidence was found to have no statistically significant effect on vaccination rates, while mortality rates were found to have a negative relationship with vaccination rates, suggesting that the rational disease dynamics theory does not apply to human papillomavirus and that vaccination rates for HPV are not prevalence elastic.

Vaccination, one of public health’s greatest disease prevention tools, is broadening to focus on adolescents. Now that there are more vaccines targeted specifically for adolescents, it is time to give more focus to vaccine delivery in this population. This research will increase the knowledge base to support informed changes in adolescent vaccine delivery by identifying knowledge and attitudes of adolescents toward vaccination within the context of barriers and solutions. Perceived susceptibility to disease, benefits and barriers to vaccination and other constructs were collected through a survey to 1368 high school students. In this population, a scheduled adolescent healthcare visit is feasible, vaccine education can diminishes health misconceptions, and vaccination mandates are ways to reach some students.

Infection with influenza virus generally produces an immune response in immunocompetent individuals that will protect from further infection of the same strain of virus. The experiments carried out in this study were designed to identify correlates of protection induced by primary experimental influenza virus infection and to compare to/and assess Particle Mediated Immunotherapeutic Delivery (PMID) as a method of DNA immunisation. Initial experiments characterised the protective immune response observed following intranasal influenza virus infection with regard to the humoral and cellular responses generated. It became clear that a combination of systemic and mucosal antibody responses and strong virus-specific CD8+ T cell response in the lungs were important in the control of viral replication. The data on the immune responses generated following influenza virus infection was used as a "gold standard" to compare the responses observed following PMID immunisation of influenza virus nucleoprotein (NP) DNA. When PMID was assessed for its ability to generate an immune response, a mucosal immune response was observed in the D-NALT as well as a more general systemic response. Whilst a single immunisation induced specific cellular responses, it proved inefficient at generating a humoral response. Protection studies showed that priming by PMID of NP DNA, resulted in a level of protection that reduced viral replication in the lungs by 2 logs. Investigation into the phenotype of virus-specific T cells generated by either infection or immunisation showed that they have a common CD44hiCD11ahlCD62L10 phenotype in various systemic and mucosal tissues. The cells isolated from the D-NALT displayed an intriguing intermediate level of CD44 expression. In vivo migration studies showed a tendency for activated splenic virus-specific CD8+ T cells to home to the D-NALT perhaps implying a role for the D-NALT as a site where effector/memory T cells actively home as part of their role in immune surveillance.

Plasmid-based gene therapy and vaccination require the production of purified plasmid DNA. There is a current understanding that supercoiled plasmid DNA is the best form for administration and FDA guidelines state that a specification for the minimum level of supercoiled plasmid DNA, in the final product, should be made. Currently, plasmids of 10 kb or smaller are being used in gene therapy trials, but if plasmid-based vaccination is to become a reality then much larger plasmids, with the ability to carry multi-variant genes, will be required, this raises questions of feasibility of production. This study examined two main issues. Firstly a fed-batch strategy to maximise the level of supercoiling of a 6.9 kb plasmid resulting from fermentation of a recombinant Escherichia coli strain was developed. Secondly issues relating to the production of larger plasmids, for gene therapy and vaccination, were examined. A fed-batch fermentation strategy for the production of a 6.9 kb plasmid, pSV, in E. coli DH5, on a semi-defined medium, was developed. Starvation of amino acids was shown to induce plasmid amplification and in batch fermentation a maximum biomass of 3.5 g/L dry cell weight (DCW) and maximum plasmid yields of 40 mg/L and 11.3 mg/g DCW were achieved. However, amplified plasmid levels were not maintained. After 31 h 90% of plasmid was in the supercoiled form but only remained so for 1 h. Maximum plasmid yield and maximum plasmid supercoiling did not occur simultaneously. Hence a strategy to delay amplification was investigated. A dual feeding strategy consisting of a linear amino acids feed and an exponential D-glucose feed was employed. In fed-batch culture a mean maximum biomass of 4.9 g/L dry cell weight and mean maximum plasmid yields of 44 mg/L and 9.1 mg/g DCW were achieved. 90% of plasmid was in the supercoiled form after 25 h and remained at this level until harvest at 35 h. An important consideration in the production of large plasmids is the ability to supply sufficient oxygen to the fermentation. E. coli DH5 harbouring either the plasmid pBGS18, a 4.4 kb pUC-based plasmid, or pQR150, a 20 kb derivative of pBGS18, was grown in D-glucose-limited chemostat culture to investigate the effects of plasmid size and recombinant protein production on oxygen demand. Under conditions where no recombinant protein was expressed the cellular oxygen demand of the two strains was not significantly different. When recombinant protein was expressed cells harbouring pBGS18 demonstrated a statistically insignificant increase in mean specific oxygen uptake rate while those harbouring pQR150 demonstrated a statistically significant increase. It was concluded that plasmid size does not significantly affect oxygen demand. The increase in oxygen demand reported with an increase in plasmid size by other researchers is due to production of recombinant protein. The production of a 116 kb plasmid, p5176, in E. coli DH10 on complex and semi-defined media was investigated. Fermentations on complex medium were poorly reproducible while those on semi-defined medium were highly reproducible. Maximum plasmid yields were comparable between the two media, while higher biomass yields and lower oxygen requirements were observed with semi-defined medium. Lysis and primary recovery were investigated and indicated that, using current methods, the manufacture of a large plasmid product may be difficult. A fed-batch fermentation strategy which allows an increased yield of supercoiled plasmid DNA has been developed. It has been established that oxygen supply is not an issue in the production of large plasmids and a base for the production of large plasmids has been established for future work.