The Antibiotic Dosage of Fastest Resistance Evolution: Gene Amplifications Underpinning the Inverted-U.

To determine the dosage at which antibiotic resistance evolution is most rapid, we treated Escherichia coli in vitro, deploying the antibiotic erythromycin at dosages ranging from zero to high. Adaptation was fastest just below erythromycin's minimal inhibitory concentration (MIC) and genotype-phenotype correlations determined from whole genome sequencing revealed the molecular basis: simultaneous selection for copy number variation in three resistance mechanisms which exhibited an "inverted-U" pattern of dose-dependence, as did several insertion sequences and an integron. Many genes did not conform to this pattern, however, reflecting changes in selection as dose increased: putative media adaptation polymorphisms at zero antibiotic dosage gave way to drug target (ribosomal RNA operon) amplification at mid dosages whereas prophage-mediated drug efflux amplifications dominated at the highest dosages. All treatments exhibited E. coli increases in the copy number of efflux operons acrAB and emrE at rates that correlated with increases in population density. For strains where the inverted-U was no longer observed following the genetic manipulation of acrAB, it could be recovered by prolonging the antibiotic treatment at subMIC dosages.

The role of CPAP treatment in elderly patients with moderate obstructive sleep apnoea: a multicentre randomised controlled trial.

The efficacy of continuous positive airway pressure (CPAP) treatment in elderly patients with nonsevere obstructive sleep apnoea (OSA) is controversial. The objective of this study was to assess the effect of CPAP treatment in elderly patients with moderate OSA in terms of clinical, quality-of-life and neurocognitive measures.This was an open-label, randomised, multicentre clinical trial in 145 elderly patients (≥70 years old) with confirmed moderate OSA (apnoea-hypopnoea index 15-29.9 events·h-1) randomised to receive CPAP (n=73) or no CPAP (n=72) for 3 months. The primary end-point was the Epworth Sleepiness Scale (ESS) score, and the secondary end-points included quality of life (Quebec Sleep Questionnaire (QSQ) domain scores), sleep-related symptoms, presence of anxiety/depression, office-based blood pressure measurements and some neurocognitive tests. The analysis was performed according to the intention-to-treat principle.Mean±sd age was 74.9±4.6 years. The CPAP group achieved a greater improvement in the ESS score (adjusted difference of 2.6 (95% CI 3.6-1.6) points; effect size 1) in some sleep-related symptoms and in some dimensions of the QSQ questionnaire (nocturnal symptoms: -0.7 (95% CI -0.3--1.0) points; p<0.0001 and emotions: -0.4 (95% CI -0.1--0.7) points; p=0.023). However, CPAP did not demonstrate any effect on either neurocognitive tests (including anxiety and depression) or blood pressure levels. There was a positive correlation between the effect of CPAP and the improvement in ESS scores and quality of life domains.CPAP treatment resulted in a significant improvement in diurnal hypersomnia and some sleep-related symptoms and quality of life domains in elderly patients with moderate OSA.

Obstructive sleep apnoea in the elderly: role of continuous positive airway pressure treatment.

Almost all the information about the effect of continuous positive airway pressure (CPAP) in patients with obstructive sleep apnoea (OSA) comes from clinical trials involving only middle-aged patients. The objective of this study was to assess the effect of CPAP treatment in elderly patients with severe OSA on clinical, quality-of-life and neurocognitive spheres. We performed an open-label, randomised, multicentre clinical trial in a consecutive clinical cohort of 224 elderly (≥70 years old) patients with confirmed severe OSA (apnoea-hypopnea index ≥30) randomised to receive CPAP (n=115) or no CPAP (n=109) for 3 months. A sleep study was performed by either full polysomnography or respiratory polygraphy. CPAP titration was performed by an autoCPAP device. The primary endpoint was quality of life (Quebec Sleep Questionnaire) and secondary endpoints included sleep-related symptoms, presence of anxiety/depression, office-based blood pressure and some neurocognitive tests. The mean±sd age was 75.5±3.9 years. The CPAP group achieved a greater improvement in all quality-of-life domains (p<0.001; effect size: 0.41-0.98), sleep-related symptoms (p<0.001; effect size 0.31-0.91) as well as anxiety (p=0.016; effect size 0.51) and depression (p<0.001; effect size: 0.28) indexes and some neurocognitive tests (digit symbol test (p=0.047; effect size: 0.20) and Trail Making Test A (p=0.029; effect size: 0.44)) in an intention-to-treat analysis. In conclusion, CPAP treatment resulted in an improvement in quality of life, sleep-related symptoms, anxiety and depression indexes and some neurocognitive aspects in elderly people with severe OSA.

Mathematical Modeling of Photo- and Thermomorphogenesis in Plants.

Increased day lengths and warm conditions inversely affect plant growth by directly modulating nuclear phyB, ELF3, and COP1 levels. Quantitative measures of the hypocotyl length have been key to gaining a deeper understanding of this complex regulatory network, while similar quantitative data are the foundation for many studies in plant biology. Here, we explore the application of mathematical modeling, specifically ordinary differential equations (ODEs), to understand plant responses to these environmental cues. We provide a comprehensive guide to constructing, simulating, and fitting these models to data, using the law of mass action to study the evolution of molecular species. The fundamental principles of these models are introduced, highlighting their utility in deciphering complex plant physiological interactions and testing hypotheses. This brief introduction will not allow experimentalists without a mathematical background to run their own simulations overnight, but it will help them grasp modeling principles and communicate with more theory-inclined colleagues.

An Ion-Pair Induced Intermediate Complex Captured in Class D Carbapenemase Reveals Chloride Ion as a Janus Effector Modulating Activity.

Antibiotic-resistant Enterobacterales that produce oxacillinase (OXA)-48-like Class D ß-lactamases are often linked to increased clinical mortality. Though the catalytic mechanism of OXA-48 is known, the molecular origin of its biphasic kinetics has been elusive. We here identify selective chloride binding rather than decarbamylation of the carbamylated lysine as the source of biphasic kinetics, utilizing isothermal titration calorimetry (ITC) to monitor the complete reaction course with the OXA-48 variant having a chemically stable N-acetyl lysine. Further structural investigation enables us to capture an unprecedented inactive acyl intermediate wedged in place by a halide ion paired with a conserved active site arginine. Supported by mutagenesis and mathematical simulation, we identify chloride as a "Janus effector" that operates by allosteric activation of the burst phase and by inhibition of the steady state in kinetic assays of ß-lactams. We show that chloride-induced biphasic kinetics directly affects antibiotic efficacy and facilitates the differentiation of clinical isolates encoding Class D from Class A and B carbapenemases. As chloride is present in laboratory and clinical procedures, our discovery greatly expands the roles of chloride in modulating enzyme catalysis and highlights its potential impact on the pharmacokinetics and efficacy of antibiotics during in vivo treatment.

Seeking patterns of antibiotic resistance in ATLAS, an open, raw MIC database with patient metadata.

Antibiotic resistance represents a growing medical concern where raw, clinical datasets are under-exploited as a means to track the scale of the problem. We therefore sought patterns of antibiotic resistance in the Antimicrobial Testing Leadership and Surveillance (ATLAS) database. ATLAS holds 6.5M minimal inhibitory concentrations (MICs) for 3,919 pathogen-antibiotic pairs isolated from 633k patients in 70 countries between 2004 and 2017. We show most pairs form coherent, although not stationary, timeseries whose frequencies of resistance are higher than other databases, although we identified no systematic bias towards including more resistant strains in ATLAS. We sought data anomalies whereby MICs could shift for methodological and not clinical or microbiological reasons and found artefacts in over 100 pathogen-antibiotic pairs. Using an information-optimal clustering methodology to classify pathogens into low and high antibiotic susceptibilities, we used ATLAS to predict changes in resistance. Dynamics of the latter exhibit complex patterns with MIC increases, and some decreases, whereby subpopulations' MICs can diverge. We also identify pathogens at risk of developing clinical resistance in the near future.

COP1 dynamics integrate conflicting seasonal light and thermal cues in the control of Arabidopsis elongation.

As the summer approaches, plants experience enhanced light inputs and warm temperatures, two environmental cues with an opposite morphogenic impact. Key components of this response are PHYTOCHROME B (phyB), EARLY FLOWERING 3 (ELF3), and CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1). Here, we used single and double mutant/overexpression lines to fit a mathematical model incorporating known interactions of these regulators. The fitted model recapitulates thermal growth of all lines used and correctly predicts thermal behavior of others not used in the fit. While thermal COP1 function is accepted to be independent of diurnal timing, our model shows that it acts at temperature signaling only during daytime. Defective response of cop1-4 mutants is epistatic to phyB-9 and elf3-8, indicating that COP1 activity is essential to transduce phyB and ELF3 thermosensory function. Our thermal model provides a unique toolbox to identify best allelic combinations enhancing climate change resilience of crops adapted to different latitudes.

Characterization of COPD Admissions During the First COVID-19 Outbreak.

Purpose: Exacerbations of COPD (ECOPD) are a frequent cause of hospitalization that seemed to ameliorate during the COVID outbreak. We aimed to evaluate the clinical characteristics of COPD-related hospital admissions and mortality in relation to the presence of COVID-19. Patients and Methods: We conducted a case-control study of patients admitted in four teaching hospitals throughout Spain between March 15 and April 30, 2020. Hospital admissions of respiratory cause with and without PCR-proven SARS-CoV-2 infection in patients with COPD were evaluated. Baseline and episode-related clinical characteristics were analyzed. Logistic regression analysis was performed to evaluate the risk for mortality. Results: During the study period, 2101 patients were admitted for respiratory worsening, 1200 (57.1%) with COVID-19. A total of 228 (10.8%) were admitted due to COPD worsening, of whom 52 (22.8%) tested positive for COVID-19. COPD patients with COVID-19, when compared to those without COVID-19, were more frequently males with better lung function (FEV1 postbronchodilator 71% vs 46% respectively, p<0.001) and had higher mortality (44.9% vs 13.6% respectively, p<0.001) despite similar age, comorbidities, total days of hospitalization and admission to intensive care unit. COVID-19 and eosinopenia were the strongest risk factors for mortality in the multivariate analysis in the overall COPD population. Inhaled corticosteroid use was not associated to mortality. Conclusion: Hospitalizations for ECOPD without COVID-19 were more frequent than COPD with COVID-19 during the first outbreak, but the latter were associated with higher mortality and low eosinophil counts that warrant further analysis.

From genotypes to organisms: State-of-the-art and perspectives of a cornerstone in evolutionary dynamics.

Understanding how genotypes map onto phenotypes, fitness, and eventually organisms is arguably the next major missing piece in a fully predictive theory of evolution. We refer to this generally as the problem of the genotype-phenotype map. Though we are still far from achieving a complete picture of these relationships, our current understanding of simpler questions, such as the structure induced in the space of genotypes by sequences mapped to molecular structures, has revealed important facts that deeply affect the dynamical description of evolutionary processes. Empirical evidence supporting the fundamental relevance of features such as phenotypic bias is mounting as well, while the synthesis of conceptual and experimental progress leads to questioning current assumptions on the nature of evolutionary dynamics-cancer progression models or synthetic biology approaches being notable examples. This work delves with a critical and constructive attitude into our current knowledge of how genotypes map onto molecular phenotypes and organismal functions, and discusses theoretical and empirical avenues to broaden and improve this comprehension. As a final goal, this community should aim at deriving an updated picture of evolutionary processes soundly relying on the structural properties of genotype spaces, as revealed by modern techniques of molecular and functional analysis.

Populations of genetic circuits are unable to find the fittest solution in a multilevel genotype-phenotype map.

The evolution of gene regulatory networks (GRNs) is of great relevance for both evolutionary and synthetic biology. Understanding the relationship between GRN structure and its function can allow us to understand the selective pressures that have shaped a given circuit. This is especially relevant when considering spatio-temporal expression patterns, where GRN models have been shown to be extremely robust and evolvable. However, previous models that studied GRN evolution did not include the evolution of protein and genetic elements that underlie GRN architecture. Here we use toyLIFE, a multilevel genotype-phenotype map, to show that not all GRNs are equally likely in genotype space and that evolution is biased to find the most common GRNs. toyLIFE rules create Boolean GRNs that, embedded in a one-dimensional tissue, develop a variety of spatio-temporal gene expression patterns. Populations of toyLIFE organisms choose the most common GRN out of a set of equally fit alternatives and, most importantly, fail to find a target pattern when it is very rare in genotype space. Indeed, we show that the probability of finding the fittest phenotype increases dramatically with its abundance in genotype space. This phenotypic bias represents a mechanism that can prevent the fixation in the population of the fittest phenotype, one that is inherent to the structure of genotype space and the genotype-phenotype map.

Parsimonious Scenario for the Emergence of Viroid-Like Replicons De Novo.

Viroids are small, non-coding, circular RNA molecules that infect plants. Different hypotheses for their evolutionary origin have been put forward, such as an early emergence in a precellular RNA World or several de novo independent evolutionary origins in plants. Here, we discuss the plausibility of de novo emergence of viroid-like replicons by giving theoretical support to the likelihood of different steps along a parsimonious evolutionary pathway. While Avsunviroidae-like structures are relatively easy to obtain through evolution of a population of random RNA sequences of fixed length, rod-like structures typical of Pospiviroidae are difficult to fix. Using different quantitative approaches, we evaluated the likelihood that RNA sequences fold into a rod-like structure and bear specific sequence motifs facilitating interactions with other molecules, e.g., RNA polymerases, RNases, and ligases. By means of numerical simulations, we show that circular RNA replicons analogous to Pospiviroidae emerge if evolution is seeded with minimal circular RNAs that grow through the gradual addition of nucleotides. Further, these rod-like replicons often maintain their structure if independent functional modules are acquired that impose selective constraints. The evolutionary scenario we propose here is consistent with the structural and biochemical properties of viroids described to date.

Data on samara morphology and wind dispersal in the invasive tree Ailanthus altissima.

The data presented in this paper is supporting the research article "Estimating wind dispersal potential in Ailanthus altissima: The need to consider the three-dimensional structure of samaras" [1]. We analyzed the estimation of samara's wind dispersal potential through a group of morphological variables that succeed in describing the three-dimensional nature of samaras. We present here a dataset containing 8 morphological variables of 200 samaras belonging to 5 different individuals of the invasive tree Ailanthus altissima (Mill.) Swingle. Additionally, we present the average descent velocity of each of the samaras, which was recorded by releasing 5 times each samara under controlled and reproducible conditions. The data set is structured in a single spreadsheet where we also included the samara and the individual identity code of the tree.

On the networked architecture of genotype spaces and its critical effects on molecular evolution.

Evolutionary dynamics is often viewed as a subtle process of change accumulation that causes a divergence among organisms and their genomes. However, this interpretation is an inheritance of a gradualistic view that has been challenged at the macroevolutionary, ecological and molecular level. Actually, when the complex architecture of genotype spaces is taken into account, the evolutionary dynamics of molecular populations becomes intrinsically non-uniform, sharing deep qualitative and quantitative similarities with slowly driven physical systems: nonlinear responses analogous to critical transitions, sudden state changes or hysteresis, among others. Furthermore, the phenotypic plasticity inherent to genotypes transforms classical fitness landscapes into multiscapes where adaptation in response to an environmental change may be very fast. The quantitative nature of adaptive molecular processes is deeply dependent on a network-of-networks multilayered structure of the map from genotype to function that we begin to unveil.

Adding levels of complexity enhances robustness and evolvability in a multilevel genotype-phenotype map.

Robustness and evolvability are the main properties that account for the stability and accessibility of phenotypes. They have been studied in a number of computational genotype-phenotype maps. In this paper, we study a metabolic genotype-phenotype map defined in toyLIFE, a multilevel computational model that represents a simplified cellular biology. toyLIFE includes several levels of phenotypic expression, from proteins to regulatory networks to metabolism. Our results show that toyLIFE shares many similarities with other seemingly unrelated computational genotype-phenotype maps. Thus, toyLIFE shows a high degeneracy in the mapping from genotypes to phenotypes, as well as a highly skewed distribution of phenotypic abundances. The neutral networks associated with abundant phenotypes are highly navigable, and common phenotypes are close to each other in genotype space. All of these properties are remarkable, as toyLIFE is built on a version of the HP protein-folding model that is neither robust nor evolvable: phenotypes cannot be mutually accessed through point mutations. In addition, both robustness and evolvability increase with the number of genes in a genotype. Therefore, our results suggest that adding levels of complexity to the mapping of genotypes to phenotypes and increasing genome size enhances both these properties.

Validation of clinical control in COPD as a new tool for optimizing treatment.

BACKGROUND: It has recently been proposed that the concept of clinical control in COPD may be useful for deciding treatment in COPD, but the original control criteria (OCC) were considered too restrictive. OBJECTIVE: Define and subsequently validate "modified" control criteria (MCC) of COPD. METHOD: Prospective observational study in COPD patients with a 1-year follow-up. Control was defined as the presence of low clinical impact and clinical stability. To evaluate clinical impact, the following clinical parameters were assessed: the degree of dyspnea, use of rescue medication, physical activity, and sputum color. Stability was assessed by clinical changes and exacerbations in the last 3 months. The COPD assessment test score and their changes were also evaluated as alternative control criteria. To define the MCC, adjustment for disease severity using BODEx index (MCC-B) or FEV1 (MCC-F) was evaluated, and the best cutoff point was established. Time to first combined event (emergency visit, hospitalization, or death) was analyzed to evaluate the predictive capacity of risk of the OCC, MCC-B, and MCC-F. RESULTS: We included 265 patients, 224 (83.9%) men, with a mean age (±SD) of 68±9 years and FEV1 of 58%±17%. The proportion of controlled patients was higher using clinical MCC-B or MCC-F (61.5% and 59.6%) than OCC (27.5%). Similar percentages were found using COPD assessment test scores. The time to the first combined event was significantly greater in controlled patients using MCC criteria (P<0.001, all cases). The predictive capacity of risk was similar in MCC-B (c-statistic [C]=0.639) and MCC-F (C=0.637) and higher than OCC (C=0.589). CONCLUSIONS: The new MCC identified a higher number of controlled COPD patients. These patients have a better quality of life and lower risk of poor outcomes. The concept of control and the new MCC could be a useful tool to optimize therapy.

Interobserver variability in rectum contouring in high-dose-rate brachytherapy for prostate cancer: A multi-institutional prospective analysis.

PURPOSE: The aim of this study was to evaluate the interobserver variability (IOV) of rectum contouring, and its dosimetric consequences, for high-dose-rate brachytherapy in patients with prostate cancer across multiple institutions. METHODS AND MATERIALS: Five radiation oncologists contoured rectums in 10 patients on transperineal ultrasound image sets after establishing a delineation consensus. The D0.1cc, D1cc, and D2cc rectum volume parameters were determined. The mean, standard deviation, and range of each dose-volume histogram parameter were evaluated for each patient. The IOV was determined using the coefficient of variation, and the dosimetric impacts on the total dose were analyzed by estimating the biologically equivalent dose (EQD2α/ß = 3). RESULTS: The interobserver coefficients of variation (±standard deviation) for the reported D0.1cc, D1cc, and D2cc were 5 ± 1.84%, 4 ± 1.26%, and 4 ± 1.33%, respectively. As for the impact on the total dose, the mean dose differences for D0.1cc, D1cc, and D2cc were 10 Gy, 7.3 Gy, and 6.6 Gy, respectively. CONCLUSIONS: The D2cc is robust as evident by the low IOV (<5%). However, some variability ranges almost overlap with the clinical threshold level, which may present dosimetric and clinical complications. General rectal contouring guidelines for prostate high-dose-rate brachytherapy are desirable to reduce discrepancies in delineation.

Adaptive multiscapes: an up-to-date metaphor to visualize molecular adaptation.

BACKGROUND: Wright's metaphor of the fitness landscape has shaped and conditioned our view of the adaptation of populations for almost a century. Since its inception, and including criticism raised by Wright himself, the concept has been surrounded by controversy. Among others, the debate stems from the intrinsic difficulty to capture important features of the space of genotypes, such as its high dimensionality or the existence of abundant ridges, in a visually appealing two-dimensional picture. Two additional currently widespread observations come to further constrain the applicability of the original metaphor: the very skewed distribution of phenotype sizes (which may actively prevent, due to entropic effects, the achievement of fitness maxima), and functional promiscuity (i.e. the existence of secondary functions which entail partial adaptation to environments never encountered before by the population). RESULTS: Here we revise some of the shortcomings of the fitness landscape metaphor and propose a new "scape" formed by interconnected layers, each layer containing the phenotypes viable in a given environment. Different phenotypes within a layer are accessible through mutations with selective value, while neutral mutations cause displacements of populations within a phenotype. A different environment is represented as a separated layer, where phenotypes may have new fitness values, other phenotypes may be viable, and the same genotype may yield a different phenotype, representing genotypic promiscuity. This scenario explicitly includes the many-to-many structure of the genotype-to-phenotype map. A number of empirical observations regarding the adaptation of populations in the light of adaptive multiscapes are reviewed. CONCLUSIONS: Several shortcomings of Wright's visualization of fitness landscapes can be overcome through adaptive multiscapes. Relevant aspects of population adaptation, such as neutral drift, functional promiscuity or environment-dependent fitness, as well as entropic trapping and the concomitant impossibility to reach fitness peaks are visualized at once. Adaptive multiscapes should aid in the qualitative understanding of the multiple pathways involved in evolutionary dynamics. REVIEWERS: This article was reviewed by Eugene Koonin and Ricard Solé.

toyLIFE: a computational framework to study the multi-level organisation of the genotype-phenotype map.

The genotype-phenotype map is an essential object to understand organismal complexity and adaptability. However, its experimental characterisation is a daunting task. Thus, simple models have been proposed and investigated. They have revealed that genotypes differ in their robustness to mutations; phenotypes are represented by a broadly varying number of genotypes, and simple point mutations suffice to navigate the space of genotypes while maintaining a phenotype. Nonetheless, most current models focus only on one level of the map (folded molecules, gene regulatory networks, or networks of metabolic reactions), so that many relevant questions cannot be addressed. Here we introduce toyLIFE, a multi-level model for the genotype-phenotype map based on simple genomes and interaction rules from which a complex behaviour at upper levels emerges -remarkably plastic gene regulatory networks and metabolism. toyLIFE is a tool that permits the investigation of how different levels are coupled, in particular how and where mutations affect phenotype or how the presence of certain metabolites determines the dynamics of toyLIFE gene regulatory networks. The model can easily incorporate evolution through more complex mutations, recombination, or gene duplication and deletion, thus opening an avenue to explore extended genotype-phenotype maps.

Internal consistency and validity of the Spanish version of the quality of life questionnaire specific for obstructive sleep apnea: sleep apnea quality of life index.

INTRODUCTION: The objective of the present study was to validate the Spanish version of the SAQLI, which is a health-related quality of life (HRQL) questionnaire specific for sleep apnea-hypopnea syndrome (SAHS), and to assess its sensitivity to change. MATERIAL AND METHODS: A multicenter study performed in a group of patients with SAHS (apnea-hypopnea index [AHI] ≥5) who had been referred to the centers' Sleep Units. All patients completed the following questionnaires: SF-36, FOSQ, SAQLI and Epworth scale. The psychometric properties (internal consistency, construct validity, concurrent validity, predictive value, repeatability and responsiveness to change) of the SAQLI were assessed (four domains: daily function, social interactions, emotional function and symptoms; an optional fifth domain is treatment-related symptoms). RESULTS: One hundred sixty-two patients were included for study (mean age: 58±12; Epworth: 10±4; BMI: 33±5.9kg m(-2); AHI: 37±15hour(-1)). The factorial analysis showed a construct of four factors with similar distribution to the original questionnaire domains. Internal consistency (Cronbach's alpha between 0.78 and 0.82 for the different domains), concurrent validity for SF-36, Epworth scale and FOSQ, and test-retest reliability were appropriate. The predictive validity of the questionnaire showed no significant correlations with the severity of SAHS. SAQLI showed good sensitivity to change in all the domains of the questionnaire (p<0,01). CONCLUSIONS: The Spanish version of the SAQLI is a valid HRQL measurement with appropriate psychometric properties for use in patients with SAHS and it is sensitive to change.