CRISPR-Cas9 enrichment, a new strategy in microbial metagenomics to investigate complex genomic regions: The case of an environmental integron.

Environmental integrons are ubiquitous in natural microbial communities, but they are mostly uncharacterized and their role remains elusive. Thus far, research has been hindered by methodological limitations. Here, we successfully used an innovative approach combining CRISPR-Cas9 enrichment with long-read nanopore sequencing to target, in a complex microbial community, a putative adaptive environmental integron, InOPS, and to unravel its complete structure and genetic context. A contig of 20 kb was recovered containing the complete integron from the microbial metagenome of oil-contaminated coastal sediments. InOPS exhibited typical integron features. The integrase, closely related to integrases of marine Desulfobacterota, possessed all the elements of a functional integron integrase. The gene cassettes harboured mostly unknown functions hampering inferences about their ecological importance. Moreover, the putative InOPS host, likely a hydrocarbonoclastic marine bacteria, raises questions as to the adaptive potential of InOPS in response to oil contamination. Finally, several mobile genetic elements were intertwined with InOPS highlighting likely genomic plasticity, and providing a source of genetic novelty. This case study showed the power of CRISPR-Cas9 enrichment to elucidate the structure and context of specific DNA regions for which only a short sequence is known. This method is a new tool for environmental microbiologists working with complex microbial communities to target low abundant, large or repetitive genetic structures that are difficult to obtain by classical metagenomics. More precisely, here, it offers new perspectives to comprehensively assess the eco-evolutionary significance of environmental integrons.

Les intégrons environnementaux sont omniprésents dans les communautés microbiennes naturelles, mais la plupart ne sont pas caractérisés et leur rôle reste obscur. A ce jour, les limitations méthodologiques ont restreint leur étude. Ici, nous avons utilisé avec succès une approche innovante, combinant l'enrichissement par CRISPR-Cas9 et le séquençage nanopore longs-fragments, pour cibler, dans une communauté microbienne complexe, un intégron environnemental potentiellement adaptatif, InOPS, et pour révéler sa structure complète et son contexte génétique. Un contig de 20 kb contenant l'intégron complet a été obtenu à partir du métagénome microbien de sédiments côtiers contaminés par du pétrole. InOPS présente les caractéristiques typiques d'un intégron. Son intégrase, proche des intégrases des Desulfobacterota marines, possède tous les éléments d'une intégrase d'intégron fonctionnelle. Les cassettes de gène ont des fonctions pour la plupart inconnues, ce qui empêche d'inférer leur importance écologique. De plus, l'hôte présumé d'InOPS, probablement une bactérie marine hydrocarbonoclaste, interroge sur le potentiel adaptatif d'InOPS en réponse à la contamination par le pétrole. En outre, la présence de plusieurs éléments génétiques mobiles dans le contig met en évidence une probable plasticité génomique qui pourrait être source de remaniements génétiques. Cette étude de cas a montré la puissance de l'enrichissement par CRISPR-Cas9 pour élucider la structure et le contexte de régions d'ADN spécifiques pour lesquelles seule une courte séquence est connue. Cette méthode fournit un nouvel outil aux microbiologistes environnementaux travaillant avec des communautés microbiennes complexes pour cibler des structures génétiques peu abondantes, larges ou répétées, qui sont difficiles à obtenir par métagénomique classique. Plus précisément, elle offre ici de nouvelles perspectives pour évaluer de manière exhaustive l'importance éco-évolutive des intégrons environnementaux.

Environmental integrons: the dark side of the integron world.

Integrons are bacterial genetic elements notorious for their role in spreading antibiotic resistance in clinical settings. In the natural environment, integrons present a wide and hidden diversity, raising questions as to their broader role in bacterial adaptation. From the One Health perspective, they must be considered a threatening pool of resistance determinants.

Epidemiology of Cardiac Events During Prehospital Care in Mountain Rescues Conducted in Aragón.

INTRODUCTION: Cardiac events are one of the leading causes of death in the Spanish population. Given the increase in the nontraumatic medical conditions found in mountain rescues, the objective of this study was to report on the heart conditions of patients rescued in the mountains of Aragón in the Spanish Pyrenees. METHODS: We conducted a retrospective observational study based on data collected from patients' medical histories for rescues undertaken in from 2010 to 2016 (at altitudes between 500 m [1640 ft] and 3404 m [11,168 ft]). RESULTS: Of the 2079 individuals rescued from 2010 to 2016, 34 (2%) were diagnosed with heart conditions, accounting for 21% of all nontraumatic medical conditions. The data showed a statistically significant increase in the age of the rescued patients with heart conditions (55±15 y) and those with acute coronary syndrome/sudden death (60±8 y). Eighty-five percent of the rescued patients with heart conditions were men, 62% were rescued above 2000 m (6500 ft), 42% had acute coronary syndrome (of whom 56% had inferior infarction), and 35% died suddenly. CONCLUSIONS: Aragón mountain rescues show an increase in patient age in recent years. The most common medical case among rescued individuals with heart conditions was a hiker over the age of 50 y with cardiovascular risk factors, inferior infarction, and occurrence at an altitude above 2000 m. Based on our observations, appropriate training should be undertaken, especially by older hikers, who may also benefit from cardiac screening, and rescue vehicles/personnel and mountain huts should be equipped with semiautomatic external defibrillators.

[Nontraumatic medical emergencies in mountain rescues]. / Utilidad de la ecografi ́a en urgencias: el protocolo RUSH.

OBJECTIVES: To describe the clinical and epidemiologic characteristics of patients with nontraumatic medical problems rescued by a Spanish mountain emergency response service (061 Aragon). MATERIAL AND METHODS: Retrospective observational analysis of records of mountain rescues completed between July 2010 and December 2016. RESULTS: A total of 164 patients with nontraumatic medical emergencies were rescued; 82.3% were males. Most patients were between the ages of 50 and 59 years. Environmentally related problems, most often hypothermia, accounted for 36.6% of the emergencies. Cardiac problems led to 20.7% and digestive problems to 12.8%. Eighty-two percent of the patients were hiking or engaged in general mountain activities (other than rock climbing, canyoning, hunting, or skiing). CONCLUSION: Recent years have seen a rise in the number of patients requiring rescue from mountains for nontraumatic medical emergencies, particularly heart problems. The typical patient to expect would be a man between the ages of 50 and 59 years who is hiking in the summer.

OBJETIVO: Describir las características clínico-epidemiológicas de los pacientes rescatados por la unidad de rescate de montaña del 061 de Aragón que presentaron patologías médicas no traumáticas. METODO: Estudio observacional retrospectivo de los rescates de montaña realizados entre julio de 2010 y diciembre de 2016. RESULTADOS: Se analizaron 164 pacientes con patología médica de origen no traumático durante el rescate (82,3% varones). El intervalo de edad más frecuente fue de 50 a 59 años. El 36,6% presentaban patologías relacionadas con el medio, siendo la hipotermia la más frecuente. El 20,7% fue patología cardiaca y el 12,8% fueron problemas digestivos. La actividad practicada por los pacientes fue el senderismo/montañismo en un 82% de las ocasiones. CONCLUSIONES: Se observa un aumento del número de pacientes rescatados con patología médica no traumática en los últimos años, y destaca el aumento de problemas cardiacos. El paciente tipo esperable sería un varón de entre 50 y 59 años practicando senderismo en época estival.

Multisector intervention to accelerate reductions in child stunting: an observational study from 9 sub-Saharan African countries.

BACKGROUND: In sub-Saharan Africa, ~ 40% of children <5 y old are stunted, with levels that have remained largely unchanged over the past 2 decades. Although the complex determinants of undernutrition are well recognized, few studies have evaluated strategies that combine nutrition-specific, health-based approaches with food system- and livelihood-based interventions. OBJECTIVE: We examined changes in childhood stunting and its determinants after 3 y of exposure to an integrated, multisector intervention and compared these changes with national trends. DESIGN: A prospective observational trial was conducted across rural sites in 9 sub-Saharan African countries with baseline levels of childhood stunting >20%. A stratified random sample of households and resident children <2 y old from villages exposed to the program were enrolled in the study. Main outcome measures included principal determinants of undernutrition and childhood stunting, which was defined as a height-for-age z score less than -2. National trends in stunting were generated from demographic and health surveys. RESULTS: Three years after the start of the program in 2005-2006, consistent improvements were observed in household food security and diet diversity, whereas coverage with child care and disease-control interventions improved for most outcomes. The prevalence of stunting in children <2 y old at year 3 of the program (2008-2009) was 43% lower (adjusted OR: 0.57; 95% CI: 0.38, 0.83) than at baseline. The average national stunting prevalence for the countries included in the study had remained largely unchanged over the past 2 decades. CONCLUSION: These findings provide encouraging evidence that a package of multisector interventions has the potential to produce reductions in childhood stunting.

Esophageal eosinophilic infiltration responds to proton pump inhibition in most adults.

BACKGROUND & AIMS: Despite consensus recommendations, eosinophilic esophagitis (EoE) is commonly diagnosed upon esophageal eosinophilic infiltration (EEI; based on ≥ 15 eosinophils per high power field; eo/HPF). We evaluated the prevalence of EEI before and after proton pump inhibitor (PPI) therapy and assessed the accuracy of EEI and pH monitoring analyses. METHODS: Biopsies were taken from the upper-middle esophagus of 712 adults with upper gastrointestinal symptoms who were referred for endoscopy due to upper gastrointestinal symptoms. Patients with EEI were treated with rabeprazole (20 mg, twice daily) for 2 months. EoE was defined by persistent symptoms and >15 eo/HPF following PPI therapy. RESULTS: Thirty-five patients (4.9%) had EEI, of whom 55% had a history of allergies, and 70% had food impaction or dysphagia as their primary complaint. Twenty-six EEI patients (75%) achieved clinicopathological remission with PPI therapy; of these, 17 had GERD-like profile (EEI <35 eo/HPF and objective evidence of reflux, based on endoscopy or pH monitoring), and 9 had EoE-like profile (EEI 35-165 eo/HPF, typical EoE symptoms and endoscopic findings). The PPI response was 50% in the EoE-like profile patients. The PPI-response was 50% in EoE-like profile patients. Likewise, PPI-responsive EEI occurred with normal (33%) and pathologic (80%) pH monitoring. Higher histologic cut-off values improved specificity and positive predictive for EoE (35%-35% for >20 eo/HPF; 46%-39% for >24 eo/HPF; 65%-50% for 35 eo/HPF). CONCLUSIONS: In adults with EEI, 75% of unselected patients and 50% with an EoE phenotype respond to PPI therapy; pH monitoring is poorly predictive of response. Patients with PPI-responsive EEI >35 eo/HPF are phenotypically undistinguishable from EoE patients. EoE might be overestimated without clinical and pathologic follow-up of patient response to PPI.

[Cardiovascular disease epidemiology and risk factors in primary care]. / Epidemiología de las enfermedades cardiovasculares y factores de riesgo en atención primaria.

INTRODUCTION AND OBJECTIVES: To study the prevalence of and risk factors for cardiovascular disease in primary care. PATIENTS AND METHOD: A cross-sectional study was carried out at an urban health center in Barcelona, Spain. In total, 2248 patients > or =15 years old were selected randomly from medical records. The study investigated cardiovascular diseases such as ischemic heart disease, cerebrovascular disease and peripheral arterial disease, and cardiovascular risk factors such as age, sex, smoking, high blood pressure, hypercholesterolemia, hypertriglyceridemia, and diabetes mellitus. RESULTS: The patients' mean age was 49.1 (18.9) years and 53.5% were male. Cardiovascular risk factor prevalences were: smoking, 35.2%; high blood pressure, 33.7%; hypercholesterolemia, 21.9%; hypertriglyceridemia,12.7%; and diabetes mellitus, 15.8%. Overall, 57.9% of patients had at least 1 cardiovascular risk factor. Significantly more males presented with each risk factor (P<.05), apart from high blood pressure. The prevalence of all risk factors, except smoking, increased with age until 74 years and then stabilized, except high blood pressure, which continued to increase. Around 10% had cardiovascular disease, with myocardial ischemia in 5.5%, cerebrovascular disease in 3.7%, and peripheral arterial disease in 2.4%. All except cerebrovascular disease were significantly more common in males (P<.05). The prevalence of cardiovascular disease was low in individuals <55 years old, particularly women, and increased with age for all forms of disease. Some 68.3% were > or =65 years old. CONCLUSIONS: The high prevalence of cardiovascular risk factors was confirmed. Cardiovascular disease was more common in males and the elderly.

[Risk of Suffering from Cardiovascular Diseases because of the Clustering of the Modifiable Cardiovascular Risk Factors in the Population Older than 15 years of a Health Care Center in Barcelona]. / riesgo de presentación de eventos cardiovasculares según la agrupación de los factores de riesgo modificables en la población mayor de 15 de años de un centro de salud de barcelona.

OBJECTIVE: Over recent decades, the death rate due to cardiovascular diseases has shown a downward trend in developed countries, as has also been the case in Spain. However, are still the leading cause of death. This study is aimed at studying the relationship between cardiovascular diseases and different modifiable cardiovascular risk factors related to their clustering. METHODS: Descriptive cross-sectional study conducted at an urban healthcare center, which included 2248 individuals ages 15 and above selected by simple random sampling of the medical record files. The risk of having some cardiovascular disease (ischemic cardiopathy, cerebrovascular disease or peripheral arteriopathy of lower limbs) was calculated in the form of an odds ration (OR) in relation to the clustering of the risk factors of smoking, high blood pressure,hypercholesterolemia, hypertriglyceridemia and diabetes mellitus,adjusted by age, sex and risk factors. RESULTS: The individuals studied had 224 cardiovascular diseases.For smoking, the OR as an isolated risk factor was 1.5 (95% CI:1.0-2.2) and 1.6 (95% CI: 0.9-2.5) with the clustering with the other four risk factors; with high blood pressure, respectively of 2.1 (95% CI: 1.5-2.9) and 1.7 (95% CI: 1.1-2.6); with hypercholesterolemia, respectively of 1.7 (95% CI: 1.2-2.4) and 1.6 (95% CI: 1.1-2.4); and with hypertriglyceridemia, respectively of 1.8(95% CI: 1.2-2.8) and 1.3 (95% CI: 0.8-2.1). The OR's showed a similar behavior on layering by each cardiovascular disease, although the highest OR's (2.4 - 3.1 range) corresponded to the clustering of diabetes mellitus and smoking. CONCLUSIONS: The risk of having cardiovascular diseasesremains high with the clustering of cardiovascular risk factors,although differences among them are found to exist.

OBJETIVO: En la últimas décadas la mortalidad por enfermedades cardiovasculares ha mostrado una tendencia decreciente en los países desarrollados, confirmada asimismo en España. No obstante siguen siendo la principal causa de mortalidad El objetivo de este trabajo es estudiar la asociación entre las enfermedades cardiovasculares y diversos factores de riesgo cardiovascular modificables en relación a su agrupación (clustering). METODOS: Estudio descriptivo transversal realizado en un centro de salud urbano, que incluyó a 2.248 personas de 15 o más años, seleccionadas por muestreo aleatorio simple del archivo de historias clínicas. Se calculó en forma de odds ratio (OR) el riesgo de tener alguna enfermedad cardiovascular (cardiopatía isquémica, enfermedad cerebrovascular o arteriopatia periférica de extremidades inferiores)en relación al clustering de los factores de riesgo tabaquismo,hipertensión arterial, hipercolesterolemia, hipertrigliceridemia y diabetes mellitus, ajustado por edad, sexo y factores de riesgo. RESULTADOS: Las personas estudiadas tenían 224 enfermedades cardiovasculares. En el tabaquismo la OR como factor de riesgo aislado fue de 1,5 (IC95%: 1,0-2,2) y de 1,6 (IC95%: 0,9-2,5) con el clustering con los otros 4 factores de riesgo; con la hipertensión arterial de 2,1 (IC95%: 1,5-2,9) y de 1,7 (IC95%: 1,1-2,6), respectivamente; con la hipercolesterolemia de 1,7 (IC95%: 1,2-2,4) y de 1,6 (IC95%: 1,1-2,4), respectivamente; con la diabetes de 2,5 (IC95%: 1,7-3,5) y 2,0 (IC95%: 1,3-3,0), respectivamente y con la hipertrigliceridemia de 1,8 (IC95%: 1,2-2,8) y 1,3 (IC95%: 0,8-2,1), respectivamente. Las OR se comportaron de manera similar al estratificar por cada enfermedad cardiovascular, aunque las OR más elevadas (entre 2,4 y 3,1) correspondieron al clustering de diabetes mellitus y tabaquismo. CONCLUSIONES: El riesgo de tener enfermedades cardiovasculares sigue siendo elevado con el clustering de factores de riesgo cardiovascular,aunque se observan diferencias entre ellos.