The type II-A CRISPR-Cas system of streptococcus mutans : characterisation of bacteriophage-insensitive mutan(t)s

Les bactéries sont continuellement exposées à un danger, la prédation par des bactériophages. Pour se défendre, elles ont développé une grande variété de mécanismes. Parmi ceux-ci, on retrouve CRISPR-Cas (« clustered regularly interspaced palindromic repeats »), un système adaptatif que possèdent environ 45% des bactéries. Une caractéristique unique du système CRISPR-Cas est qu’il constitue en quelque sorte la mémoire de l’hôte. Par exemple, le système peut emmagasiner des petits fragments d’un génome viral, appelés espaceurs, et les introduire dans son CRISPR. Cette mémoire lui permet de se défendre contre une réinfection par le même virus ou un virus hautement apparenté. Par contre, malgré que l’acquisition de nouveaux espaceurs semble fréquente dans la nature, ce phénomène n’est que très rarement observé en conditions de laboratoire. Néanmoins, quelques bactéries font exception à la règle et l’une d’entre elles est Streptococcus mutans. Dans le cadre de cette étude, l’interaction entre la souche S. mutans P42S et le bactériophage virulent M102AD a été analysée en détail. De plus, certaines applications potentielles du système CRISPR-Cas ont également été approfondies. Le premier objectif de cette thèse était de caractériser le système CRISPR-Cas de S. mutans P42S au niveau moléculaire et de déterminer son rôle dans les interactions phage-bactérie. Le deuxième objectif était d’établir le potentiel de la protéine Cas9 de S. mutans P42S (SmutCas9) comme nouvel outil d’édition génomique. S. mutans P42S possède un système CRISPR-Cas de type II-A. Bien que ce type de système soit probablement le plus étudié, celui de S. mutans P42S présente plusieurs caractéristiques uniques lui permettant de se démarquer. En effet, ce dernier reconnaît un PAM différent de ce qui était auparavant connu pour cette espèce bactérienne, l’acquisition simultanée de multiples espaceurs semble fréquente, ce qui est probablement dû au phénomène de « priming ». Malgré le rôle de CRISPR-Cas dans la défense antivirale, S. mutans P42S dispose d’autres mécanismes de défense contre les phages. Des cellules mutantes sont résistantes aux phages en empêchant l’adsorption de particules virales à la cellule ont notamment été observées. D'autres mécanismes sont assurément impliqués dans la défense antivirale de S. mutans. Finalement, SmutCas9 s’est montrée efficace dans l’édition de génomes viraux et elle apparaît comme une candidate à explorer pour cette application.Bacteria are exposed to the constant threat of viral predation. To defend themselves, bacteria have developed a wide variety of different mechanisms. One of these mechanisms is CRISPR-Cas (clustered regularly interspaced palindromic repeats), an adaptive immune mechanism found in approximately 45% of bacteria. A unique feature of CRISPR-Cas systems compared to other antiviral defence mechanisms is that it has a memory. The system is capable of remembering previous viral encounters and protects the bacterial host from re-infection by the same or highly-related viruses. This memory is due to the acquisition of virus-derived genome fragments called spacers. Despite common acquisition of novel spacers in nature, and thereby the emergence of new immunity, acquisition of new spacers under laboratory conditions has been rarely observed. One of the few exceptions is Streptococcus mutans. In this study, the interactions between S. mutans strain P42S and its virulent bacteriophage M102AD are investigated in detail. In addition, possible applications of the CRISPR-Cas system are analysed. The first objective of this thesis was to characterise the CRISPR-Cas system of S. mutans P42S on the molecular level and to determine its role in antiviral defence. The second objective was to determine the potential of the Cas9 protein of S. mutans P42S (SmutCas9) in genome editing. S. mutans P42S possesses a type II-A CRISPR-Cas system. Although this is arguably the best studied system, the one found in the strain S. mutans P42S has several features that makes it stand out. It recognises a PAM different from what was known for this species, multiple spacer acquisitions are frequent, and this appears to be partially due to priming. Although CRISPR-Cas plays a role in antiviral defence, there are additional antiviral defence mechanisms that protect S. mutans against phages. Adsorption resistance is one of them, although additional unidentified antiviral defence mechanisms are likely involved. Finally, SmutCas9 has been shown functional in editing of viral genomes and appears to be a candidate for human genome editing

Exercise and Coronary Atherosclerosis: Observations, Explanations, Relevance, and Clinical Management.

Physical activity and exercise training are effective strategies for reducing the risk of cardiovascular events, but multiple studies have reported an increased prevalence of coronary atherosclerosis, usually measured as coronary artery calcification, among athletes who are middle-aged and older. Our review of the medical literature demonstrates that the prevalence of coronary artery calcification and atherosclerotic plaques, which are strong predictors for future cardiovascular morbidity and mortality, was higher in athletes compared with controls, and was higher in the most active athletes compared with less active athletes. However, analysis of plaque morphology revealed fewer mixed plaques and more often only calcified plaques among athletes, suggesting a more benign composition of atherosclerotic plaques. This review describes the effects of physical activity and exercise training on coronary atherosclerosis in athletes who are middle-aged and older and aims to contribute to the understanding of the potential adverse effects of the highest doses of exercise training on the coronary arteries. For this purpose, we will review the association between exercise and coronary atherosclerosis measured using computed tomography, discuss the potential underlying mechanisms for exercise-induced coronary atherosclerosis, determine the clinical relevance of coronary atherosclerosis in middle-aged athletes and describe strategies for the clinical management of athletes with coronary atherosclerosis to guide physicians in clinical decision making and treatment of athletes with elevated coronary artery calcification scores

Epidemiology of Heart Failure

Cardiovascular mortality rates have decline'd significantly in most industrialized countries over the past three decades.2 Nevertheless, cardiovascular disease remains one of the most important causes of morbidity and mortality in Western society, especially as the average age of the population increases.'" Heart failure is rapidly becoming one of the most prevalent cardiovascular disorders and the incidence of heart failure is expected to continue to increase for some time to come.5,6 Unfortunately, it appears that the declining fatality rate of acute coronary events/ resulting in a larger group of persons at increased risk of developing chronic cardiovascular disease, contributes to the rise of heart failure. The paradox of better care is expanded by the observation that treatment of hypertension may actually postpone rather than prevent the onset of heart failure. 8 The prognosis of heart failure is poor9 and the economic impact of heart failure on health services is considerable because of the long-tenn pharmacological treatment and frequent hospitalizations associated with the syndrome. This burden is set to increase further as the prognosis of patients with heart failure is improved by medical and surgical interventions10- 13 and the proportion of the elderly increases in Western society

More Than One Way to Solve the Healthcare Innovation Crisis With Digital Platforms. Various Forms of Platform Openness Impacting Primary Healthcare

Whereas open digital platforms drive innovation in industries, platforms in primary healthcare are mostly closed. Policy-makers have been looking for ways to open up primary healthcare platforms to stimulate collaboration and innovation and need to do so even more due to the ongoing COVID-19 crisis. Yet, there is not one way of opening up platforms in primary healthcare, just as it is unclear how different ways of openness can lead to more innovation. This paper analyzes the opportunities and challenges in realizing platform openness while examining alternative forms of openness. To answer this, we (1) conceptualize different forms of platform openness (sponsor-provider-platform-user openness), (2) examine how these forms of openness can resolve barriers to innovation, and (3) examine what challenges need to be overcome to realize that form of openness in practice, such as complexity in roles, regulations, and ICT infrastructure. The findings are relevant to structure further research on how platform openness leads to more innovations in healthcare

Cardiac imaging to detect coronary artery disease in athletes aged 35 years and older. A scoping review.

Sudden cardiac death (SCD) is a devastating event in athletes. Screening efforts that were first directed at athletes younger than 35 years, are now focusing on the rapidly growing group of older sportspersons. Athletes aged ≥35 years have a 10-fold increased risk of exercise-related cardiac arrest, mostly due to coronary artery disease (CAD). Although cardiac imaging is pivotal in identifying CAD, the role of imaging modalities in screening asymptomatic older sportspersons remains unclear. We performed a scoping review to identify the role of cardiac imaging to detect CAD in older sportspersons and to identify gaps in the existing literature. We searched Medline, Embase and the Cochrane library for studies reporting data on cardiac imaging of CAD in sportspersons ≥35 years. The systematic search yielded 1737 articles and 14 were included in this scoping review. Imaging modalities included 2 echocardiography, 1 unenhanced Computed Tomography (CT) for coronary artery calcium scoring (CACS), 3 CACS and contrast-enhanced CT angiography (CCTA), 2 CACS and Cardiac Magnetic Resonance (CMR), 1 CCTA with CMR and echocardiography, 2 CCTA, 2 CMR, and 1 myocardial perfusion imaging article. The low number of relevant articles and the selection bias introduced by studying specific groups, like veteran marathon runners, indicate the need for future research. Cardiac CT (CACS and CCTA) probably has the highest potential for pre-participation screening, with high diagnostic value to detect CAD and low radiation dose. However, currently there is insufficient evidence for incorporating routine cardiac imaging in the pre-participation screening of asymptomatic sportspersons over 35 years

Non-adherence to ivabradine and placebo and outcomes in chronic heart failure: an analysis from SHIFT

Aims In heart failure, non-adherence increases events; in turn, the effect of hospitalization on adherence is incompletely understood. We explored the relationship of non-adherence to outcomes, hospitalizations with non-adherence, and the influence of non-adherence on treatment effects of heart rate lowering with ivabradine. Methods and results In the randomized, controlled Systolic Heart failure treatment with the If-inhibitor ivabradine Trial (SHIFT), we studied the effect of non-adherence (n = 1287) compared with adherence (n = 5204) on cardiovascular outcomes. After adjustment, non-adherence was associated with the primary composite endpoint of cardiovascular death and heart failure hospitalization (hazard ratio 3.47, 95% confidence interval 2.91–4.13, P < 0.0001). No interaction with the treatment groups of placebo or ivabradine (P for interaction 0.54) occurred. Similar results for cardiovascular death and heart failure hospitalization, as well as for cardiovascular hospitalization, heart failure death, and total death were observed. The effect of ivabradine was maintained in patients being adherent or becoming non-adherent during the trial (P for interaction = 0.54). Patients with a previous hospitalization were more likely to become non-adherent thereafter. Conclusions Non-adherence identifies a group at particularly high cardiovascular event risk independent of treatment allocation. Non-adherent patients in the ivabradine group maintain a treatment benefit. Patients with previous hospitalizations are more likely to become non-adherent and represent a group of particularly high-risk patients in whom special attention to stimulate adherence may be valuable

Historical trends in survival of hospitalized heart failure patients: 2000 versus 1995

BACKGROUND: Population-based secular trends in survival of patients with congestive heart failure (CHF) are central to public health research on the burden of the syndrome. METHODS: Patients 35–79 years old with a CHF discharge code in 1995 or 2000 were identified in 22 Minneapolis-St. Paul hospitals. A sample of the records was abstracted (50% of 1995 records; 38% of 2000 records). A total of 2,257 patients in 1995 and 1,825 patients in 2000 were determined to have had a CHF-related hospitalization. Each patient was followed for one year to ascertain vital status. RESULTS: The risk profile of the 2000 patient cohort was somewhat worse than that of the 1995 cohort in both sex groups, but the distributions of age and left ventricular ejection fraction were similar. Within one year of admission in 2000, 28% of male patients and 27% of female patients have died, compared to 36% and 27% of their counterparts in 1995, respectively. In various Cox regression models the average year effect (2000 vs. 1995) was around 0.75 for men and 0.95 to 1.00 for women. The use of angiotensin converting-enzyme inhibitors and beta-blockers was associated with substantially lower hazard of death during the subsequent year. CONCLUSION: Survival of men who were hospitalized for CHF has improved during the second half of the 1990s. The trend in women was very weak, compatible with little to no change. Documented benefits of angiotensin converting-enzyme inhibitors and beta-blockers were evident in these observational data in both men and women

The Relationship Between Lifelong Exercise Volume and Coronary Atherosclerosis in Athletes.

Background -Higher levels of physical activity are associated with a lower risk of cardiovascular events. Nevertheless, there is debate on the dose-response relationship of exercise and CVD outcomes and whether high volumes of exercise may accelerate coronary atherosclerosis. We aimed to determine the relationship between lifelong exercise volumes and coronary atherosclerosis. Methods -Middle aged men engaged in competitive or recreational leisure sports underwent a non-contrast and contrast-enhanced computed tomography scan to assess coronary artery calcification (CAC) and plaque characteristics. Participants reported lifelong exercise history patterns. Exercise volumes were multiplied by Metabolic Equivalent of Task (MET) scores to calculate MET-min/week. Participants were categorized as 2000 MET-min/week. Results -284 men (55±7 years) were included. CAC was present in 150/284 (53%) participants with a median CAC score of 35.8 [9.3-145.8). Athletes with a lifelong exercise volume >2000 MET-min/week (n=75) had a significantly higher CAC score (9.4 [0-60.9] versus 0 [0-43.5], p=.02) and prevalence of CAC (68%,ORadjusted=3.2 (95%CI: 1.6-6.6)) and plaque (77%, ORadjusted=3.3 (95%CI: 1.6-7.1)) compared to 0, there was no difference in CAC score (p=.20), area (p=.21), density (p=.25) and regions of interest (p=.20) across exercise volume groups. Among participants with plaque, the most active group (>2000 MET-min/week) had a lower prevalence of mixed plaques (48% versus 69%, ORadjusted=0.35 (95%CI: 0.15-0.85) and more often had only calcified plaques (38% versus 16%, ORadjusted=3.57 (95%CI: 1.28-9.97)) compared to the least active group (2000 MET-min/week group had a higher prevalence of CAC and atherosclerotic plaques. The most active group did however have a more benign composition of plaques, with fewer mixed plaques and more often only calcified plaques. These observations may explain the increased longevity typical of endurance athletes despite the presence of more coronary atherosclerotic plaque in the most active participants