Background and Purpose- The GLX (glycocalyx) is a protein/polysaccharide meshwork at the cellular surface. Consisting largely of glycosaminoglycans and proteoglycans, the GLX can shed in response to ...stress. In this study, we assay 11 components of the GLX in plasma from patients with ischemic stroke from a longitudinal cohort. Methods- Plasma samples from healthy individuals (N=8), and patients with ischemic stroke day ≥3, day 7, and day 90 (N=9-14) were immunoassayed for diverse components of the GLX. Results- Median stroke severity was mild (National Institutes of Health Stroke Scale 2.0 (range, 0-6) at day ≤3). Three (keratan-chondroitin-heparan-sulfate) of 4 glycosaminoglycans and CD44 (proteoglycan) were increased at day 7 and returned to baseline at day 90. Proteoglycan syndecan (Syn)-3 increased and Syn-2 levels decreased, significantly. Conclusions- Individual GLX components are often assayed as stand-alone biomarkers for endothelial health. This study suggests a full assessment of GLX components is more indicative of the endothelial health of an individual and represents a complex GLX signature that may be valuable as a composite biomarker of disease.
Bacterial airway infections, predominantly caused by
P. aeruginosa
, are a major cause of mortality and morbidity of CF patients. While short insertions and deletions as well as point mutations ...occurring during infection are well studied, there is a lack of understanding of how gene loss and acquisition play roles in bacterial adaptation to the human airways. Here, we investigated
P. aeruginosa
within-host evolution with regard to gene loss and acquisition. We show that during long-term infection
P. aeruginosa
genomes tend to lose genes, in particular, genes related to virulence. This adaptive strategy allows reduction of the genome size and evasion of the host’s immune response. This knowledge is crucial to understand the basic mutational steps that, on the timescale of years, diversify lineages and adds to the identification of bacterial genetic determinants that have implications for CF disease.
ABSTRACT
Genome analyses have documented that there are differences in gene repertoire between evolutionary distant lineages of the same bacterial species; however, less is known about microevolutionary dynamics of gene loss and acquisition within bacterial lineages as they evolve over years. Here, we analyzed the genomes of 45
Pseudomonas aeruginosa
lineages evolving in the lungs of cystic fibrosis (CF) patients to identify genes that are lost or acquired during the first years of infection. On average, lineage genome content changed by 88 genes (range, 0 to 473). Genes were more often lost than acquired, and prophage genes were more variable than bacterial genes. We identified convergent loss or acquisition of the same genes across lineages, suggesting selection for loss and acquisition of certain genes in the host environment. We found that a notable proportion of such genes are associated with virulence; a trait previously shown to be important for adaptation. Furthermore, we also compared the genomes across lineages to show that the within-lineage variable genes (i.e., genes that had been lost or acquired during the infection) often belonged to genomic content not shared across all lineages. In sum, our analysis adds to the knowledge on the pace and drivers of gene loss and acquisition in bacteria evolving over years in a human host environment and provides a basis to further understand how gene loss and acquisition play roles in lineage differentiation and host adaptation.
IMPORTANCE
Bacterial airway infections, predominantly caused by
P. aeruginosa
, are a major cause of mortality and morbidity of CF patients. While short insertions and deletions as well as point mutations occurring during infection are well studied, there is a lack of understanding of how gene loss and acquisition play roles in bacterial adaptation to the human airways. Here, we investigated
P. aeruginosa
within-host evolution with regard to gene loss and acquisition. We show that during long-term infection
P. aeruginosa
genomes tend to lose genes, in particular, genes related to virulence. This adaptive strategy allows reduction of the genome size and evasion of the host’s immune response. This knowledge is crucial to understand the basic mutational steps that, on the timescale of years, diversify lineages and adds to the identification of bacterial genetic determinants that have implications for CF disease.
► This study examines how organizations handle the internal dimension of crisis management and crisis communication before, during and after a crisis. ► During a crisis, employees typically produce ...more informal communication, lack information, and feel frustrated and insecure. ► The larger the organization the more crisis-prepared. ► The employees are communicatively active in a crisis situation. ► Public organizations have their own approach to internal crisis management and crisis communication.
The aim of this paper is to present and discuss some of the main findings from a large survey of internal crisis management and crisis communication conducted in the spring of 2011 among public and private organizations in Denmark (the ICMCC survey). The survey was conducted among the 367 largest private companies (selected from DK 1000, established by Børsen business magasin) and among 98 public organizations (municipalities). The overall goal was to get a preliminary idea of how these companies or organizations perceive, plan, coordinate and implement internal crisis management and crisis communication activities before, during and after a crisis. The survey questionnaire comprised 36 questions and was sent to respondents who typically are responsible for the crisis-preparedness of their organizations. The results from the survey show that the vast majority of organizations have a crisis or contingency plan, and most of these plans contain an internal dimension relating to the management and communication with the internal stakeholders during a crisis. Thus, the study shows a rather professional and formalized behavior towards crisis management in general, but also when it comes to managing a crisis in relation to the internal organizational stakeholders in specific. In addition, the results clearly indicate a strong relation between organizational size and crisis management; the larger the organization the more likely to have a crisis plan. This particularly pertained to the private organizations. The ICMCC survey forms part of a major collaborative research project, financed by the Danish Council for Independent Research/Social Sciences (2011–2014), entitled Internal Crisis Management and Crisis Communication in Danish Organizations. The purpose of this three-year long project is to shed light upon the role of internal crisis management and crisis communication before, during and after an organizational crisis and/or a societal crisis leading to downsizing or major changes within an organization or an organizational field.
species are increasingly being detected in patients with cystic fibrosis (CF), and this emerging pathogen is associated with antibiotic resistance and more-severe disease outcomes. Nonetheless, ...little is known about the extent of transmission and antibiotic resistance development in
infections. We sequenced the genomes of 101
clinical isolates (identified as
based on matrix-assister laser desorption ionization-time of flight MALDI-TOF or API N20 typing) collected from 51 patients with CF-the largest longitudinal data set to date. We performed phylogenetic analysis on the genomes and combined this with epidemiological and antibiotic resistance data to identify patient-to-patient transmission and the development of antibiotic resistance. We confirmed that the MALDI-TOF or API N20 method was not sufficient for
species-level typing and that the population of
isolates was composed of five different species, among which
accounted for 52% of infections. Most patients were infected by unique
clone types; nonetheless, suspected patient-to-patient transmission cases identified by shared clone types were observed in 35% (
= 18) of patients. In 15 of 16 cases, the suspected transmissions were further supported by genome- or clinic visit-based epidemiological analysis. Finally, we found that resistance developed over time. We show that whole-genome sequencing (WGS) is essential for
species typing and identification of patient-to-patient transmission, which was revealed for
,
, and, for the first time,
Furthermore, we show that the development of antibiotic resistance is associated with chronic
infections. Our findings emphasize that transmission and antibiotic resistance should be considered in future treatment strategies.
Traditional studies on the evolution of antibiotic resistance development use approaches that can range from laboratory-based experimental studies, to epidemiological surveillance, to sequencing of ...clinical isolates. However, evolutionary trajectories also depend on the environment in which selection takes place, compelling the need to more deeply investigate the impact of environmental complexities and their dynamics over time. Herein, we explored the within-patient adaptive long-term evolution of a Pseudomonas aeruginosa hypermutator lineage in the airways of a cystic fibrosis (CF) patient by performing a chronological tracking of mutations that occurred in different subpopulations; our results demonstrated parallel evolution events in the chromosomally encoded class C β-lactamase (
). These multiple mutations within
shaped diverse coexisting alleles, whose frequency dynamics responded to the changing antibiotic selective pressures for more than 26 years of chronic infection. Importantly, the combination of the cumulative mutations in
provided structural and functional protein changes that resulted in a continuous enhancement of its catalytic efficiency and high level of cephalosporin resistance. This evolution was linked to the persistent treatment with ceftazidime, which we demonstrated selected for variants with robust catalytic activity against this expanded-spectrum cephalosporin. A "gain of function" of collateral resistance toward ceftolozane, a more recently introduced cephalosporin that was not prescribed to this patient, was also observed, and the biochemical basis of this cross-resistance phenomenon was elucidated. This work unveils the evolutionary trajectories paved by bacteria toward a multidrug-resistant phenotype, driven by decades of antibiotic treatment in the natural CF environmental setting.
Antibiotics are becoming increasingly ineffective to treat bacterial infections. It has been consequently predicted that infectious diseases will become the biggest challenge to human health in the near future. Pseudomonas aeruginosa is considered a paradigm in antimicrobial resistance as it exploits intrinsic and acquired resistance mechanisms to resist virtually all antibiotics known. AmpC β-lactamase is the main mechanism driving resistance in this notorious pathogen to β-lactams, one of the most widely used classes of antibiotics for cystic fibrosis infections. Here, we focus on the β-lactamase gene as a model resistance determinant and unveil the trajectory P. aeruginosa undertakes on the path toward a multidrug-resistant phenotype during the course of two and a half decades of chronic infection in the airways of a cystic fibrosis patient. Integrating genetic and biochemical studies in the natural environment where evolution occurs, we provide a unique perspective on this challenging landscape, addressing fundamental molecular mechanisms of resistance.
It is unclear whether recurrent sputum culture with Pseudomonas aeruginosa from patients with chronic obstructive pulmonary disease (COPD) is caused by intermittent airway carriage by different ...P. aeruginosa lineages or persistent carriage by the same lineage, and whether lineages genetically adapt during carriage.
Whole-genome sequencing was performed for P. aeruginosa isolates sampled longitudinally from sputum cultures in patients with COPD who were enrolled in an ongoing randomized controlled trial (clinicaltrials.gov: NCT03262142).
A total of 153 P. aeruginosa isolates were sequenced for 23 patients during 365 days of follow-up. Recurrent presence of P. aeruginosa was seen in 19 patients (83%) and was caused by persistence of the same clonal lineage in all but one patient. We identified 38 genes mutated in parallel in two or more lineages, suggesting positive selection for adaptive mutations. Mutational enrichment analysis revealed genes important in antibiotic resistance and chronic infections to be more frequently mutated.
Recurrent P. aeruginosa was common and carried for a prolonged time after initial detection in the airways of patients with COPD. Recurrence was caused by persistence of the same clonal lineage and was associated with genetic adaptation. Trial data on possible clinical benefits of attempting antibiotic eradication of P. aeruginosa in COPD are warranted.
The small RNA ErsA of
was previously suggested to be involved in biofilm formation via negative post-transcriptional regulation of the
gene that encodes the virulence-associated enzyme AlgC, which ...provides sugar precursors for the synthesis of several polysaccharides. In this study, we show that a knock-out
mutant strain forms a flat and uniform biofilm, not characterized by mushroom-multicellular structures typical of a mature biofilm. Conversely, the knock-out mutant strain showed enhanced swarming and twitching motilities. To assess the influence of ErsA on the
transcriptome, we performed RNA-seq experiments comparing the knock-out mutant with the wild-type. More than 160 genes were found differentially expressed in the knock-out mutant. Parts of these genes, important for biofilm formation and motility regulation, are known to belong also to the AmrZ transcriptional regulator regulon. Here, we show that ErsA binds
and positively regulates
mRNA at post-transcriptional level
suggesting an interesting contribution of the ErsA-
mRNA interaction in biofilm development at several regulatory levels.
Type II toxin-antitoxin (TA) systems are most commonly composed of two genes encoding a stable toxin, which harms the cell, and an unstable antitoxin that can inactivate it. TA systems were initially ...characterized as selfish elements, but have recently gained attention for regulating general stress responses responsible for pathogen virulence, formation of drug-tolerant persister cells and biofilms-all implicated in causing recalcitrant chronic infections. We use a bioinformatics approach to explore the distribution and evolution of type II TA loci of the opportunistic pathogen,
, across longitudinally sampled isolates from cystic fibrosis lungs. We identify their location in the genome, mutations, and gain/loss during infection to elucidate their function(s) in stabilizing selfish elements and pathogenesis. We found (1) 26 distinct TA systems, where all isolates harbor four in their core genome and a variable number of the remaining 22 on genomic islands; (2) limited mutations in core genome TA loci, suggesting they are not under negative selection; (3) no evidence for horizontal transmission of elements with TA systems between clone types within patients, despite their ability to mobilize; (4) no gain and limited loss of TA-bearing genomic islands, and of those elements partially lost, the remnant regions carry the TA systems supporting their role in genomic stabilization; (5) no significant correlation between frequency of TA systems and strain ability to establish as chronic infection, but those with a particular TA, are more successful in establishing a chronic infection.
Antibiotic resistance has become a serious threat to human health (WHO
. Geneva: World Health Organization; 2017), and the ability to predict antibiotic resistance from genome sequencing has become a ...focal point for the medical community. With this genocentric prediction in mind, we were intrigued about two particular findings for a collection of clinical
isolates (Marvig
.
2015;47:57-64; Frimodt-Møller
.
2018;8:12512; Bartell
.
2019;10:629): (i) 15 out of 52 genes found to be frequently targeted by adaptive mutations during the initial infection stage of cystic fibrosis airways ('candidate pathoadaptive genes') (Marvig
.
2015;47:57-64) were associated with antibiotic resistance (López-Causapé
.
2018;9:685; López-Causapé
.
2018;62:e02583-17); (ii) there was a parallel lack of resistance development and linkage to the genetic changes in these antibiotic-resistance-associated genes (Frimodt-Møller
.
2018;8:12512; Bartell
.
2019;10:629). In this review, we highlight alternative selective forces that potentially enhance the infection success of
and focus on the linkage to the 15 pathoadaptive antibiotic-resistance-associated genes, thereby showing the problems we may face when using only genomic information to predict and inform about relevant antibiotic treatment.
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