Respiratory diseases are a major cause of human morbidity and mortality and are a leading cause of economic loss to livestock producers. The respiratory tract is constantly in contact with dust, bacteria, fungi, viruses and other pathogenic agents that are found in the air. Normally, the body has the ability to clear these foreign particles. However, physiological and environmental stresses can impair airway defense mechanisms resulting in establishment of pulmonary infections. The microbes and their products engage various receptors in the lung to activate epithelium, endothelium, macrophages, neutrophils and other cells. The activation of inflammatory cascade in the lung results in recruitment of neutrophils, damage to air-blood barrier and development of edema. Although there have been significant advances in our understanding of mechanisms of lung inflammation, there have been a lack of any significant advances in the development of new therapeutics to manage lung disease, which may suggest that our understanding of the inflammatory mechanisms is still incomplete. Pentraxin 3 (PTX3) is an innate immune protein which has been implicated in a diverse range of inflammatory processes, such as recruitment of cells and production of cytokines. PTX3 is an acute phase protein, with low or undetectable levels in the circulation of healthy humans and animals, and rapid, dramatic increase in inflammatory diseases. The expression and function of this protein has not been characterized in the lungs of domestic animal species. Because of potential implications of PTX3 in lung inflammation, I studied the expression of PTX3 in normal and inflamed lungs of calves, pigs, horses, foals and humans. Lungs from all of these species showed expression of PTX3 in airway epithelium, alveolar septa, vascular endothelium and inflammatory cells. Western blot performed on homogenates from normal and inflamed lungs from calves and pigs show an increased expression of PTX3 in inflamed lungs (P<0.05). Because protein function is influenced by its location in the cell, I clarified the subcellular expression of PTX3 with immuno-electron microscopy on normal and inflamed calf and horse lungs. PTX3 was localized on pulmonary intravascular macrophages, monocytes, neutrophils and, unexpectedly, platelets. PTX3 was also present in the nuclei of neutrophils, monocytes and pulmonary intravascular macrophages. Neutrophils are critical regulators of acute lung inflammation. Having observed PTX3 in neutrophils, I investigated the effect of E. coli lipopolysaccharide-induced activation on PTX3 in neutrophils in vitro. Neutrophils challenged with E. coli LPS were examined at 30, 60, 90 and 120 minutes after the treatment. Normal peripheral blood neutrophils showed PTX3 expression. Neutrophils activated with LPS appeared ruffled and showed loss of PTX3 expression at 30 minutes followed by recovery of the expression. Western blots performed on normal and activated neutrophil homogenates did not show any differences (P=0.05). Collectively, the data show PTX3 in normal and inflamed lungs across multiple species. PTX3 was also detected in normal and activated neutrophils. While the function of intriguing localization of PTX3 in the nuclei as well as in platelets is not known, the similarity of expression across the species suggest a role for PTX3 in lung inflammation.

Bovine respiratory disease (BRD) is the principal cause of morbidity and mortality among feedlot cattle. Mannheimia haemolytica is consistently implicated in this condition, but treatment options are diminishing with the rise of antimicrobial resistance and intensifying consumer pressure to reduce reliance on conventional therapies. Thus, sustainable alternatives like vaccination are required. In this study, the phenotypic and genotypic diversity of BRD pathogens were examined with the objective to identify vaccine targets using reverse vaccinology, an innovative approach to identify antigens via genomic sequence. Preliminary surveillance confirmed M. haemolytica serotype 2 isolates were predominant in healthy animals (75.5%) while serotypes 1 (70.7%) and 6 (19.5%) were common in diseased animals. Pathogens of BRD, including M. haemolytica, Pasteurella multocida and Histophilus somni were also isolated from North American BRD mortalities, and compared using pulsed-field gel electrophoresis and antimicrobial susceptibility. Concurrently, polymerase chain reaction detection of bacterial and viral agents confirmed that M. haemolytica with bovine viral diarrhea virus were the most prevalent. Whereas isolates from live cattle were found to have a relatively low level of resistance, several pathogens from the mortalities were found to contain integrative conjugative elements (ICE) conferring resistance to seven antimicrobial classes. These ICEs were transferred via conjugation to other bacterial species, emphasizing the need for alternative antimicrobial therapies. Collectively, data from these investigations informed the selection of 11 diverse M. haemolytica strains for whole genome sequencing and comparative analyses. Several bacteriophage associated genes and CRISPR-Cas regulated gene expression systems were identified and are likely contributing to virulence in M. haemolytica. Coding sequences across all genomes were screened using pan-genome analysis, identifying 291 candidates with cell-surface associated signatures. Using a cell-free translation system and enzyme-linked immunosorbent assay the candidates were screened against serum from cattle challenged with serovar 1, 2 or 6 of M. haemolytica, and ranked according to immunogenicity. The top five vaccine candidates included Ssa1, ComE, a solute binding protein, an outer membrane protein, and the periplasmic component of an ABC transporter. With further characterization, these unique antigenic candidates could be developed into a vaccine to effectively reduce the dependence on antimicrobial therapies.

A large number of studies have shown that both temperature and air pollution (eg, particulate matter and ozone) are associated with health outcomes. So far, it has received limited attention whether air pollution and temperature interact to affect health outcomes. A few studies have examined interactive effects between temperature and air pollution, but produced conflicting results. This thesis aimed to examine whether air pollution (including ozone and particulate matter) and temperature interacted to affect health outcomes in Brisbane, Australia and 95 large US communities. In order to examine the consistency across different cities and different countries, we used two datasets to examine interactive effects of temperature and air pollution. One dataset was collected in Brisbane City, Australia, during 1996-2000. The dataset included air pollution (PM10, ozone and nitrogen dioxide), weather conditions (minimum temperature, maximum temperature, relative humidity and rainfall) and different health outcomes. Another dataset was collected from the 95 large US communities, which included air pollution (ozone was used in the thesis), weather conditions (maximum temperature and dew point temperature) and mortality (all non-external cause mortality and cardiorespiratory mortality). Firstly, we used three parallel time-series models to examine whether maximum temperature modified PM10 effects on cardiovascular hospital admissions (CHA), respiratory hospital admissions (RHA), cardiovascular emergency visits (CEV), respiratory emergency visits (REV), cardiovascular mortality (CM) and non-external cause mortality (NECM), at lags of 0-2 days in Brisbane. We used a Poisson generalized additive model (GAM) to fit a bivariate model to explore joint response surfaces of both maximum temperature and particulate matter less than 10 μm in diameter (PM10) on individual health outcomes at each lag. Results show that temperature and PM10 interacted to affect different health outcomes at various lags. Then, we separately fitted non-stratification and stratification GAM models to quantify the interactive effects. In the non-stratification model, we examined the interactive effects by including a pointwise product for both temperature and the pollutant. In the stratification model, we categorized temperature into two levels using different cut-offs and then included an interactive term for both pollutant and temperature. Results show that maximum temperature significantly and positively modified the associations of PM10 with RHA, CEV, REV, CM and NECM at various lags, but not for CHA. Then, we used the above Poisson regression models to examine whether PM10 modified the associations of minimum temperature with CHA, RHA, CEV, REV, CM and NECM at lags of 0-2 days. In this part, we categorized PM10 into two levels using the mean as cut-off to fit the stratification model. The results show that PM10 significantly modified the effects of temperature on CHA, RHA, CM and NECM at various lags. The enhanced adverse temperature effects were found at higher levels of PM10, but there was no clear evidence for synergistic effects on CEV and REV at various lags. Three parallel models produced similar results, which strengthened the validity of these findings. Thirdly, we examined whether there were the interactive effects between maximum temperature and ozone on NECM in individual communities between April and October, 1987-2000, using the data of 60 eastern US communities from the National Morbidity, Mortality, and Air Pollution Study (NMMAPS). We divided these communities into two regions (northeast and southeast) according to the NMMAPS study. We first used the bivariate model to examine the joint effects between temperature and ozone on NECM in each community, and then fit a stratification model in each community by categorizing temperature into three levels. After that, we used Bayesian meta-analysis to estimate overall effects across regions and temperature levels from the stratification model. The bivariate model shows that temperature obviously modified ozone effects in most of the northeast communities, but the trend was not obviously in the southeast region. Bayesian meta-analysis shows that in the northeast region, a 10-ppb increment in ozone was associated with 2.2% (95% posterior interval [PI]: 1.2%, 3.1 %), 3.1% (95% PI: 2.2%, 3.8 %) and 6.2 % (95% PI: 4.8%, 7.6 %) increase in mortality for low, moderate and high temperature levels, respectively, while in the southeast region, a 10-ppb increment in ozone was associated with 1.1% (95% PI: -1.1%, 3.2 %), 1.5% (95% PI: 0.2%, 2.8%) and 1.3% (95% PI: -0.3%, 3.0 %) increase in mortality. In addition, we examined whether temperature modified ozone effects on cardiovascular mortality in 95 large US communities between May and October, 1987-2000 using the same models as the above. We divided the communities into 7 regions according to the NMMAPS study (Northeast, Industrial Midwest, Upper Midwest, Northwest, Southeast, Southwest and Southern California). The bivariate model shows that temperature modified ozone effects in most of the communities in the northern regions (Northeast, Industrial Midwest, Upper Midwest, Northwest), but such modification was not obvious in the southern regions (Southeast, Southwest and Southern California). Bayesian meta-analysis shows that temperature significantly modified ozone effects in the Northeast, Industrial Midwest and Northwest regions, but not significant in Upper Midwest, Southeast, Southwest and Southern California. Nationally, temperature marginally positively modified ozone effects on cardiovascular mortality. A 10-ppb increment in ozone was associated with 0.4% (95% posterior interval [PI]: -0.2, 0.9 %), 0.3% (95% PI: -0.3%, 1.0%) and 1.6% (95% PI: 4.8%, 7.6%) increase in mortality for low, moderate and high temperature levels, respectively. The difference of overall effects between high and low temperature levels was 1.3% (95% PI: - 0.4%, 2.9%) in the 95 communities. Finally, we examined whether ozone modified the association between maximum temperature and cardiovascular mortality in 60 large eastern US communities during the warmer days, 1987-2000. The communities were divided into the northeast and southeast regions. We restricted the analyses to the warmer days when temperature was equal to or higher than the median in each community throughout the study period. We fitted a bivariate model to explore the joint effects between temperature and ozone on cardiovascular mortality in individual communities and results show that in general, ozone positively modified the association between temperature and mortality in the northeast region, but such modification was not obvious in the southeast region. Because temperature effects on mortality might partly intermediate by ozone, we divided the dataset into four equal subsets using quartiles as cut-offs. Then, we fitted a parametric model to examine the associations between temperature and mortality across different levels of ozone using the subsets. Results show that the higher the ozone concentrations, the stronger the temperature-mortality associations in the northeast region. However, such a trend was not obvious in the southeast region. Overall, this study found strong evidence that temperature and air pollution interacted to affect health outcomes. PM10 and temperature interacted to affect different health outcomes at various lags in Brisbane, Australia. Temperature and ozone also interacted to affect NECM and CM in US communities and such modification varied considerably across different regions. The symmetric modification between temperature and air pollution was observed in the study. This implies that it is considerably important to evaluate the interactive effect while estimating temperature or air pollution effects and further investigate reasons behind the regional variability.

Vacinas inativadas de vírus inteiro (WIV) são amplamente utilizadas para o controle de influenza A em suínos (SIV) nos Estados Unidos e Europa e conferem proteção contra cepas homólogas à vacina reduzindo a doença clínica. Porém, a proteção da WIV pode ser limitada contra um desafio heterólogo. O aumento da doença respiratória associado à vacina (VAERD) foi descrito em suínos vacinados com uma WIV contendo uma cepa δ1-cluster-H1N2 e desafiado com uma cepa heteróloga contendo a mesma hemaglutinina, H1N1pdm09. Nesse contexto, os suínos apresentaram pneumonia severa com aumento das lesões pulmonares associado ao uso da WIV. No primeiro manuscrito foi avaliado se a formulação da WIV, o tempo entre o reforço vacinal e o desafio, e a idade de vacinação tinham impacto em VAERD. Suínos foram vacinados com duas vacinas bivalentes, WIV-δ1H1N2/H3N2 (MN08-TX98) ou WIV-δ1H1N2/H1N1pdm09 (MN08-CA09), administradas em duas doses com intervalo de três semanas, e desfiados com H1N1pdm09, três semanas após o reforço. Além disso, dois grupos foram vacinados com uma WIV-δ1H1N2 (MN08) e desafiados três ou seis semanas após o reforço. Em um estudo subsequente, dois grupos vacinados com uma WIV-δ1H1N2 com duas doses da vacina em diferentes idades: a primeira dose com 4 ou 9 semanas de idade e o reforço com 7 ou 12 semanas de idade. E um grupo vacinado com uma vacina viva atenuada de influenza (LAIV). Os animais foram desafiados com H1N1pdm09 às 15 semanas de idade. Com exceção dos grupos MN08-CA09, LAIV e controles, todos os grupos WIV apresentaram lesões pulmonares consistentes com VAERD. Nenhum grupo induziu anticorpos neutralizantes para o H1N1pdm09, exceto MN08-CA09. No entanto, grupos vacinados com a WIV-MN08 em diferentes idades induziram anticorpos IgG não-neutralizantes com reatividade cruzada com o H1N1pdm09 no soro e no pulmão, comparado com LAIV e controles. Esses dados sugerem que a idade da primeira dose da vacina, a formulação com a vacina bivalente MN08-TX98 e o desafio seis semanas após o reforço vacinal, não preveniram o desenvolvimento de VAERD. Em contraste, a vacinação com a LAIV demonstrou proteção até oito semanas após reforço. No segundo manuscrito foi avaliado o impacto dos adjuvantes na formulação da WIV no modelo de VAERD. Cinco grupos foram vacinados com WIV-δ1H1N2 formuladas com diferentes adjuvantes comerciais: óleo em água 1 (OW1), OW2, nano-emulsão (NE), gel polímero (GP) e partículas imuno-estimulantes aquosa (IMP). O protocolo vacinal/desafio foi utilizado com três semanas de intervalo entre as doses. Os grupos vacinados com WIV-OW apresentaram lesões macroscópicas nos pulmões consistentes com VAERD comparado com WIV-NE, WIV-GP e controles. Nenhum dos grupos induziu anticorpos neutralizantes contra o vírus do desafio (H1N1pdm09). Similarmente, os grupos WIV-OW, WIV-NE e WIV-GP induziram anticorpos não neutralizantes IgG de reatividade cruzada contra o H1N1pdm09 no soro e pulmão. Apesar dos anticorpos não neutralizantes com reatividade cruzada nos grupos WIV-NE e WIV-GP, não apresentaram lesões pulmonares tão severas como os grupos OW. Além disso, os adjuvantes tiveram um papel significativo na imunogenicidade da WIV e no desenvolvimento de VAERD. / Whole inactivated virus (WIV) vaccines reduce clinical disease against homologous influenza A virus (IAV) infection and are widely used in swine; however, WIV has been associated with vaccine-associated enhanced respiratory disease (VAERD) when challenged with heterologous IAV of the same hemagglutinin subtype. Here, we evaluated the impact of age at vaccination and timing between vaccination and challenge on development of VAERD using a model with a human-like 1δ cluster swine MN08 H1N2 as the vaccine component and pandemic H1N1 (H1N1pdm09) virus as the challenge strain. Pigs were vaccinated with bivalent WIV vaccine containing H1N2-MN08/H3N2-TX98 or H1N2-MN08/H1N1pdm09-CA09. All groups were challenged with H1N1pdm09 at 3 weeks post-boost (wpb). In addition, a monovalent WIV MN08 H1N2 group was challenged at 6 wpb to determine if time post-vaccination played a role on VAERD. In a follow up study, we compared the time of first WIV vaccination (4 or 9 weeks of age) and the boost three weeks later (7 or 12 weeks of age) to determine if age at first vaccination or if 8 weeks duration between vaccination and challenge impacted VAERD. A live-attenuated influenza virus (LAIV) vaccine administered at 4 and 7 weeks of age was also included. Pigs from study 2 were challenged with H1N1pdm09 at 15 weeks of age. All mismatched WIV groups had significantly higher macroscopic and microscopic lung lesions compared with LAIV, bivalent MN08-CA09 and non-vaccinated challenge controls. These data suggest that the age of first vaccination or length of time between booster dose and subsequent challenge do not alter the development VAERD in WIV vaccinated pigs. In the second paper, we evaluated the impact of adjuvant in WIV vaccine on VAERD-affected pigs. Pigs were vaccinated with WIV containing swine MN08/H1N2 that were formulated with five different commercially available adjuvants: OW1, OW2, nano-emulsion (NE), gel polymer (GP), and aqueous immunostimulating particulate (IMP). Pigs were vaccinated with 2 doses, 3 weeks apart, by the intramuscular (WIV-OW1, -OW2, -NE, and -GP) or intranasal (WIV-IMP) routes. Non-vaccinated and challenged pigs (NV/C) and non-vaccinated, non-challenged pigs (NV/NC) were included as controls. Following challenge with H1N1pdm09, WIV-OW1 and WIV-OW2 groups had significantly higher percentages of macroscopic lung lesions consistent with VAERD compared to the NV/C controls, and in contrast to WIV-NE, WIV-GP and WIV-IMP. Prior to challenge, WIV-OW, NE and GP groups had the hemagglutination inhibition antibody (HI) titers to the vaccine strain compared with controls. The WIV-IMP group had HI mean titers below the positive cut-off, with no response to any immune parameter measured, so it could not be implicated or excluded in the VAERD model. None of the groups had cross-reactive HI antibodies against the H1N1pdm09. WIV-OW groups had significantly higher levels of IgG antibody in serum against homologous and heterologous virus; however, the WIV-NE and WIV-GP groups also had serum IgG antibody against both viruses, so presence of total virus IgG alone did not explain the different VAERD outcomes. Adjuvant played a significant role in WIV immunogenicity and in VAERD; however the mechanism requires further investigation.

Master of Science / Department of Diagnostic Medicine/Pathobiology / Richard Hesse / Bovine Coronavirus (BCoV) is known to cause enteric and respiratory diseases, such as calf diarrhea, winter dysentery, calf respiratory disease, and bovine respiratory disease complex (BRD). All of these diseases are believed to be caused by the same genotype of BCoV. BCoV exhibits tissue tropism for both the gastrointestinal and respiratory tracts. This tropism is due to 9-O-acetylated sialic acid receptor on both epithelial cells in the respiratory and enteric tract. Currently, the only vaccine available for BCoV targets the enteric form of the disease. This study addresses the hypothesis that antibodies from the enteric form of the disease can cross neutralize the respiratory form of the virus. Data from surveillance studies suggest that BCoV is one of the major contributors to BRD, for which there is no currently approved vaccine for the respiratory form of the disease. Our approach to answering this question is to sequence and analyze the complete genome of 11 respiratory and enteric coronavirus isolates using next generation sequencing (NGS). Following the NGS, viruses were selected based on phylogenetic analysis and ability to grow and be maintained in cell culture. These viruses were then be used as serum neutralization indicator viruses in SN assays. 147 bovine serums submitted to KSVDL were used to determine if there are any serological differences between the immune response to respiratory versus enteric viruses based on the antibodies produced by the animal. The overall results show that there are few differences between the enteric and respiratory isolates at the genomic level and the serological response from the animal to these viruses. The differences between enteric and respiratory virus will need to be further addressed and analyzed to conclude if there is a noteworthy difference between the viruses with different tropisms. Other factors, such as host immune response and environment, are believed to be involved in the virus tropism to certain areas of the body.

Doctor of Philosophy / Department of Diagnostic Medicine/Pathobiology / Robert L. Larson / Bradley J. White / There are multiple modalities available to evaluate health or stress status of animals. The objective of my research was to evaluate different modalities including behavior, rectal and nasal temperature, and blood samples to determine the relationship with these outcomes of interest in bovine respiratory disease (BRD) events, environmental conditions, transportation, and Mannheimia haemolytica challenge model. The objective for the final project was to determine whether diagnostic sensitivity or specificity resulted in greater economic value for the industry using simulation models for identification of BRD. There was a positive association with rectal temperature and probability of not finishing the production cycle normally, but this relationship was not linear. Rectal temperature of feedlot calves at first treatment for BRD had limited value as a prognostic indicator of whether those calves would finish the production cycle normally. A positive association between rectal temperature and ambient temperature and temperature-humidity index was determined. Environmental conditions must be considered when rectal temperature is used as a diagnostic tool. At 48 hours after initiation of transportation there were no differences in body weight, rectal temperature, and time spent at various locations in the pen detected between transported and non-transported control heifers. Transportation of heifers during periods of high ambient temperatures caused transient changes in physiologic and behavioral indices of heifers. Calves challenged with Mannheimia haemolytica had more changes in behavior, body weight, and blood biomarkers during high ambient temperatures compared to control calves. Results of this study may guide research in development of objective assessment tools for identification and management of cattle affected with BRD during extreme summer conditions. For both low and high apparent prevalence cohorts, increasing diagnostic specificity resulted in more rapid, positive change in net returns compared to change in increasing sensitivity. Improvement of diagnostic specificity, perhaps through a confirmatory test or pen-level diagnostics, can increase diagnostic value. Mortality risk was the primary driver for net returns. Results from this study are important for determining future research priorities to analyze diagnostic techniques for BRD and provide a novel way for modeling diagnostic tests.

Doctor of Philosophy / Diagnostic Medicine/Pathobiology / Michael D. Apley / Bovine respiratory disease (BRD) remains a major disease in beef production systems. The administration of antimicrobials for both the control and treatment of acute BRD is common. According to most published accounts, antimicrobial resistance among BRD pathogens is increasing; therefore, judicious antimicrobial usage is vital for continued efficacy. The introduction of a novel antimicrobial class has not occurred for well over a decade, therefore it is paramount to maximize efficacy of the antimicrobials currently available. The challenge is targeting the perfect scenario: maximizing clinical efficacy while minimizing antimicrobial resistance. The host-pathogen-drug interaction is very complex and despite current sophisticated technology, this interaction is still not well understood for many infectious diseases. This dissertation work sought to investigate the effects of the administration of a macrolide for both control and treatment of acute BRD on the prevalence of resistance among isolated Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni. Whole genome sequencing of M. haemolytica was utilized to investigate the presence/absence of macrolide resistance genes and their relationship to the observed minimum inhibitory concentration. Cattle were sampled (plasma and pulmonary epithelial lining fluid) after administration of gamithromycin for drug concentration analysis. A non-linear mixed effects approach was used to fit a compartmental model to the resulting sparse pharmacokinetic data so that a complete time concentration curve could be simulated. From these curves, the CMAX and AUC were measured and used to calculate standard PKPD indices using the MIC values of the isolated bacteria. Clear associations between the use of gamithromycin for control and treatment of BRD and a statistically significantly increased likelihood of macrolide resistance were not found, possibly due to sample size limitations. The calculation of pharmacokinetic-pharmacodynamic indices found that a longer drug exposure was more closely associated with a successful treatment outcome, but there was not a statistically significant correlation. However, there were few clinical failures in this study giving further credence to the complexity of the in vivo system. There are many factors beyond pharmacokinetics/pharmacodynamics and MICs that contribute to the success of a treatment regimen for cattle suffering from BRD.

The purpose of this thesis was to extend our understanding of the assessment and interpretation of aerobic exercise function of paediatric patients with cystic fibrosis (CF). The first investigation sought to establish (1) the validity of traditional criteria to verify maximal oxygen (V ̇O2max) during a maximal cardiopulmonary exercise test (CPET); and (2) the utility of supramaximal verification (Smax) to confirm V ̇O2max. Traditional criteria significantly underreported V ̇O2max, whilst Smax was shown to provide a valid measurement in this patient group. The reproducibility of this CPET protocol, over the short- (48 h) and medium- (4-6 weeks) term, was then established in study two. V ̇O2max was repeatedly determined with no learning effect over 48 h (typical error (TE): ∆150 mL; ∆9.3%) and 4-6 weeks (TE: ∆160 mL; ∆13.3%). Supplementary maximal and submaximal CPET parameters should be incorporated for a comprehensive evaluation of a patient, however they are characterised by greater variability over time. The influence of mild-to-moderate CF on aerobic exercise function and the matching of muscle O2 delivery-to-O2 utilisation during ramp incremental exercise to exhaustion were then examined in study three. Aerobic function was impaired in CF, indicated by very likely reduced fat-free mass normalised V ̇O2max (mean difference, ±90% CI: -7.9 mL∙kg-1∙min-1, ±6.1), very likely lower V ̇O2 gain (-1.44 mL∙min-1∙W-1, ±1.12) and a likely slower V ̇O2 mean response time (MRT) (11 s, ±13). Arterial oxygen saturation was lower in CF, supporting the notion that centrally mediated O2 delivery may be impaired during ramp incremental exercise. Although a faster rate of fractional O2 extraction would be expected in the face of reduced O2 delivery, this was not observed, suggesting additional impairment in O2 extraction and utilisation at the periphery in CF. The fourth study then demonstrated the clinical utility of CPET to assess the response to 12 weeks treatment with Ivacaftor, using a case-based design. Whilst one patient with relatively mild disease demonstrated no meaningful change in V ̇O2max, the second demonstrated a 30% improvement in V ̇O2max, due to increased O2 delivery and extraction. Furthermore, changes in aerobic function were detected earlier than spirometric indices of pulmonary function. This study demonstrated that CPET represents an important and comprehensive clinical assessment tool and its use as an outcome measure in the functional assessment of patients is encouraged. Study five investigated the V ̇O2 kinetics in this patient group. During moderate intensity cycling, the phase II V ̇O2 time constant (τ) (p = 0.84, effect size (ES) = 0.11) and overall MRT (p = 0.52, ES=0.33) were not slower in CF. However, both were slowed during very heavy intensity cycling (p = 0.02, ES = 1.28 and p = 0.01, ES = 1.40, respectively) in CF. Cardiac output and muscle deoxygenation dynamics were unaltered in CF, however, the arterial-venous O2 content difference (C(a-v ̅)O2) was reduced (p=0.03) during VH and ∆C(a-v ̅)O2 correlated with the phase II τ (r= -0.85; p=0.02) and MRT (r = -0.79; p=0.03) in CF. This study showed that impaired oxidative muscle metabolism in this group is exercise intensity-dependent and mechanistically linked to an intrinsic intramuscular impairment, which limits O2 extraction and utilisation. In conclusion, this thesis has provided guidelines for a valid and reproducible CPET protocol for children and adolescents with mild-to-moderate CF, demonstrated the utility of CPET as clinical outcome measure and furthered our understanding of the factors responsible for impaired aerobic exercise function in this patient group.

La capacité musculaire et la performance à l’effort des patients sont devenues des enjeux prioritaires dans la prise en soins des pathologies respiratoires aigües ou chroniques. Il est maintenant bien établi que dans les secteurs de réanimation ou de l’insuffisance respiratoire chronique obstructive, la fonction musculaire est un facteur indépendamment associé au pronostic vital des patients. Les investigations pour l’étude des réserves cardio-respiratoires et musculaires sont primordiales pour optimiser la prise en soins des patients. Dans le cadre de cette thèse, nous avons recherché à explorer de façon non invasive les capacités des patients à déployer leur réserve cardio-circulatoire et respiratoire au cours de différentes situations d’effort (du sevrage de la ventilation mécanique au réentraînement à l’effort). Dans la première partie nous avons observé sur 124 patients intubés en cours de sevrage de la ventilation mécanique leur force musculaire inspiratoire. Nous avons défini la faiblesse des muscles respiratoires par une pression inspiratoire maximale inférieure à 30 cmH2O. Nous avons observé que la faiblesse musculaire inspiratoire était un facteur indépendamment associé à la mortalité à un an de l’extubation. Ensuite nous avons montré sur 90 patients que la mesure des pressions respiratoires maximales effectuée via une sonde d’intubation étaient fiables. Pour finir nous avons différenciés l’atteinte musculaire inspiratoire et l’atteinte musculaire périphérique sur 99 patients et nous avons observés qu’une atteinte combinée était associée à une augmentation de la mortalité à court terme. Dans la seconde partie nous avons évaluer la réponse cardio-vasculaire et la microcirculation musculaire de 20 patients intubés et sédatés au cours de 4 techniques de réhabilitation précoce. Nous avons constaté que 3 techniques sur 4 augmentaient peu le métabolisme à l’effort. Seule une technique combinant mouvement et contraction musculaire induite par stimulation électrique (FES-Cycling) induisait une réponse physiologique suggérant une activité musculaire plus intense. Dans la dernière partie, nous avons étudié la FES-Cycling dans le cadre d’une séance de réentrainement à l’effort d’intensité modérée (50% de la VO2pic) sur 25 patients BPCO. Comparativement à un placebo, la FES-Cycling entrainait une augmentation de la VO2 au cours de l’effort sans augmentation de la dyspnée ressentie, suggérant une intensité de travail atteinte plus importante. Ainsi, nous avons utilisé plusieurs méthodes non invasives pour étudier et optimiser la condition musculaire des patients handicapés respiratoire au cours de l’effort. / Muscular and exercise capacities of patients have become priority issues in taking care in acute or chronic respiratory disease. It is now well established in Intensive Care Unit or Chronic Obstructive Pulmonary Disease that muscle function is an independently factor associated with the prognosis of patients. Investigations for the study of cardiorespiratory and muscle reserves are essential to optimize the taking care of patients. As part of this thesis, we sought to explore by non-invasive ways the ability of patients to deploy their cardiopulmonary reserve during different conditions (to weaning from mechanical ventilation to pulmonary rehabilitation). In the first part we observed the inspiratory muscle strength on 124 intubated patients during weaning from mechanical ventilation. We have defined the weakness of the respiratory muscles by a maximum inspiratory pressure less than 30 cmH2O. We observed that the inspiratory muscle weakness at extubation was an independently factor associated with one year-mortality. Then we have shown on 90 patients that the maximum respiratory pressure measurement via a tube were reliable. Finally we have differentiated the inspiratory muscle damage and peripheral muscle weakness on 99 patients and we have seen that an overlap weakness was associated with an increase in the short term mortality. In the second part we evaluated the cardiovascular response and muscle microcirculation of 20 sedated patients and under mechanical ventilation during 4 early rehabilitation techniques. We have found that 3 techniques on 4 did not alter metabolism to the effort. Only one technique combining movement and muscle contraction induced by functional electrical stimulation (FES-Cycling) induced a physiological response suggesting efficient muscle activity. In the last part, we studied on 25 COPD patients the FES-Cycling in a single session of pulmonary rehabilitation at a moderate intensity (50% of the VO2peak). Compared to placebo, the FES-Cycling training increased VO2 during exercise without increased the perceived dyspnoea, suggesting an intensity reached more important. Thereby, we used several non-invasive methods to study and optimize the muscle condition during exerise of respiratory disabled patients.

No description available.