Aerobic growth of Rhodococcus aetherivorans BCP1 using selected naphthenic acids as the sole carbon and energy sources

Naphthenic acids (NAs) are an important group of toxic organic compounds naturally occurring in hydrocarbon deposits. This work shows that Rhodococcus aetherivorans BCP1 cells not only utilize a mixture of eight different NAs (8XNAs) for growth but they are also capable of marked degradation of two model NAs, cyclohexanecarboxylic acid (CHCA) and cyclopentanecarboxylic acid (CPCA) when supplied at concentrations from 50 to 500 mgL-1. The growth curves of BCP1 on 8XNAs, CHCA, and CPCA showed an initial lag phase not present in growth on glucose, which presumably was related to the toxic effects of NAs on the cell membrane permeability. BCP1 cell adaptation responses that allowed survival on NAs included changes in cell morphology, production of intracellular bodies and changes in fatty acid composition. Transmission electron microscopy (TEM) analysis of BCP1 cells grown on CHCA or CPCA showed a slight reduction in the cell size, the production of EPS-like material and intracellular electron-transparent and electron-dense inclusion bodies. The electron-transparent inclusions increased in the amount and size in NA-grown BCP1 cells under nitrogen limiting conditions and contained storage lipids as suggested by cell staining with the lipophilic Nile Blue A dye. Lipidomic analyses revealed significant changes with increases of methyl-branched (MBFA) and polyunsaturated fatty acids (PUFA) examining the fatty acid composition of NAs-growing BCP1 cells. PUFA biosynthesis is not usual in bacteria and, together with MBFA, can influence structural and functional processes with resulting effects on cell vitality. Finally, through the use of RT (Reverse Transcription)-qPCR, a gene cluster (chcpca) was found to be transcriptionally induced during the growth on CHCA and CPCA. Based on the expression and bioinformatics results, the predicted products of the chcpca gene cluster are proposed to be involved in aerobic NA degradation in R. aetherivorans BCP1. This study provides first insights into the genetic and metabolic mechanisms allowing a Rhodococcus strain to aerobically degrade NAs

Influence of Sociodemographic, Premorbid, and Injury-Related Factors on Post-Concussion Symptoms after Traumatic Brain Injury

Background: Post-concussion symptoms (PCS) are often reported as consequences of mild and moderate traumatic brain injury (TBI), but these symptoms are not well documented in severe TBI. There is a lack of agreement as to which factors and covariates affect the occurrence, frequency, and intensity of PCS among TBI severity groups. The present study therefore aims to examine the association between sociodemographic, premorbid, and injury-related factors and PCS. Methods: A total of 1391 individuals (65% male) from the CENTER-TBI study were included in the analyses. The occurrence, frequency (number of PCS), and intensity (severity) of PCS were assessed using the Rivermead Post-concussion Symptoms Questionnaire (RPQ) at six months after TBI. To examine the association between selected factors (age, sex, living situation, employment status, educational background, injury and TBI severity, and premorbid problems) and PCS, a zero-inflated negative binomial model (ZINB) for occurrence and frequency of PCS and a standard negative binomial regression (NB) for intensity were applied. Results: Of the total sample, 72% of individuals after TBI reported suffering from some form of PCS, with fatigue being the most frequent among all TBI severity groups, followed by forgetfulness, and poor concentration. Different factors contributed to the probability of occurrence, frequency, and intensity of PCS. While the occurrence of PCS seemed to be independent of the age and sex of the individuals, both the frequency and intensity of PCS are associated with them. Both injury and TBI severity influence the occurrence and frequency of PCS, but are associated less with its intensity (except "acute" symptoms such as nausea, vomiting, and headaches). Analyses focusing on the mTBI subgroup only yielded results comparable to those of the total sample. Discussion: In line with previous studies, the results support a multifactorial etiology of PCS and show the importance of differentiating between their occurrence, frequency, and intensity to better provide appropriate treatment for individual subgroups with different symptoms (e.g., multiple PCS or more intense PCS). Although PCS often occur in mild to moderate TBI, individuals after severe TBI also suffer from PCS or post-concussion-like symptoms that require appropriate treatment. The chosen statistical approaches (i.e., ZINB and NB models) permit an ameliorated differentiation between outcomes (occurrence, frequency, and intensity of PCS) and should be used more widely in TBI research

Prehospital Management of Traumatic Brain Injury across Europe: A CENTER-TBI Study

Background Prehospital care for traumatic brain injury (TBI) is important to prevent secondary brain injury. We aim to compare prehospital care systems within Europe and investigate the association of system characteristics with the stability of patients at hospital arrival. Methods We studied TBI patients who were transported to CENTER-TBI centers, a pan-European, prospective TBI cohort study, by emergency medical services between 2014 and 2017. The association of demographic factors, injury severity, situational factors, and interventions associated with on-scene time was assessed using linear regression. We used mixed effects models to investigate the case mix adjusted variation between countries in prehospital times and interventions. The case mix adjusted impact of on-scene time and interventions on hypoxia (oxygen saturation Results Among 3878 patients, the greatest driver of longer on-scene time was intubation (+8.3 min, 95% CI: 5.6-11.1). Secondary referral was associated with shorter on-scene time (-5.0 min 95% CI: -6.2- -3.8). Between countries, there was a large variation in response (range: 12-25 min), on-scene (range: 16-36 min) and travel time (range: 15-32 min) and in prehospital interventions. These variations were not explained by patient factors such as conscious level or severity of injury (expected OR between countries: 1.8 for intubation, 1.8 for IV fluids, 2.0 for helicopter). On-scene time was not associated with the regional EMS policy (p= 0.58). Hypotension and/or hypoxia were seen in 180 (6%) and 97 (3%) patients in the overall cohort and in 13% and 7% of patients with severe TBI (GCS Discussion Hypoxia and hypotension continue to occur in patients who suffer a TBI, and remain relatively common in severe TBI. Substantial variation in prehospital care exists for patients after TBI in Europe, which is only partially explained by patient factors.</div

Prediction of global functional outcome and post-concussive symptoms after mild Traumatic Brain Injury: external validation of prognostic models in the Collaborative European NeuroTrauma Effectiveness Research in Traumatic Brain Injury (CENTER-TBI) study

The majority of traumatic brain injuries (TBIs) are categorized as mild, according to a baseline Glasgow Coma Scale (GCS) score of 13-15. Prognostic models that were developed to predict functional outcome and persistent post-concussive symptoms (PPCS) after mild TBI have rarely been externally validated. We aimed to externally validate models predicting 3-12-month Glasgow Outcome Scale Extended (GOSE) or PPCS in adults with mild TBI. We analyzed data from the Collaborative European NeuroTrauma Effectiveness Research in Traumatic Brain Injury (CENTER-TBI) project, which included 2862 adults with mild TBI, with 6-month GOSE available for 2374 and Rivermead Post-Concussion Symptoms Questionnaire (RPQ) results available for 1605 participants. Model performance was evaluated based on calibration (graphically and characterized by slope and intercept) and discrimination (C-index). We validated five published models for 6-month GOSE and three for 6-month PPCS scores. The models used different cutoffs for outcome and some included symptoms measured 2 weeks post-injury. Discriminative ability varied substantially (C-index between 0.58 and 0.79). The models developed in the Corticosteroid Randomisation After Significant Head Injury (CRASH) trial for prediction of GOSE <5 discriminated best (C-index 0.78 and 0.79), but were poorly calibrated. The best performing models for PPCS included 2-week symptoms (C-index 0.75 and 0.76). In conclusion, none of the prognostic models for early prediction of GOSE and PPCS has both good calibration and discrimination in persons with mild TBI. In future studies, prognostic models should be tailored to the population with mild TBI, predicting relevant end-points based on readily available predictors.Analysis and support of clinical decision makingDevelopment and application of statistical models for medical scientific researc

Fundamental deficits of auditory perception in Wernicke’s aphasia

Objective: This work investigates the nature of the comprehension impairment in Wernicke’s aphasia, by examining the relationship between deficits in auditory processing of fundamental, non-verbal acoustic stimuli and auditory comprehension. Wernicke’s aphasia, a condition resulting in severely disrupted auditory comprehension, primarily occurs following a cerebrovascular accident (CVA) to the left temporo-parietal cortex. Whilst damage to posterior superior temporal areas is associated with auditory linguistic comprehension impairments, functional imaging indicates that these areas may not be specific to speech processing but part of a network for generic auditory analysis. Methods: We examined analysis of basic acoustic stimuli in Wernicke’s aphasia participants (n = 10) using auditory stimuli reflective of theories of cortical auditory processing and of speech cues. Auditory spectral, temporal and spectro-temporal analysis was assessed using pure tone frequency discrimination, frequency modulation (FM) detection and the detection of dynamic modulation (DM) in “moving ripple” stimuli. All tasks used criterion-free, adaptive measures of threshold to ensure reliable results at the individual level. Results: Participants with Wernicke’s aphasia showed normal frequency discrimination but significant impairments in FM and DM detection, relative to age- and hearing-matched controls at the group level (n = 10). At the individual level, there was considerable variation in performance, and thresholds for both frequency and dynamic modulation detection correlated significantly with auditory comprehension abilities in the Wernicke’s aphasia participants. Conclusion: These results demonstrate the co-occurrence of a deficit in fundamental auditory processing of temporal and spectrotemporal nonverbal stimuli in Wernicke’s aphasia, which may have a causal contribution to the auditory language comprehension impairment Results are discussed in the context of traditional neuropsychology and current models of cortical auditory processing

Enhancing Production of Bio-Isoprene Using Hybrid MVA Pathway and Isoprene Synthase in E. coli

The depleting petroleum reserve, increasingly severe energy crisis, and global climate change are reigniting enthusiasm for seeking sustainable technologies to replace petroleum as a source of fuel and chemicals. In this paper, the efficiency of the MVA pathway on isoprene production has been improved as follows: firstly, in order to increase MVA production, the source of the “upper pathway” which contains HMG-CoA synthase, acetyl-CoA acetyltransferase and HMG-CoA reductase to covert acetyl-CoA into MVA has been changed from Saccharomyces cerevisiae to Enterococcus faecalis; secondly, to further enhance the production of MVA and isoprene, a alanine 110 of the mvaS gene has been mutated to a glycine. The final genetic strain YJM25 containing the optimized MVA pathway and isoprene synthase from Populus alba can accumulate isoprene up to 6.3 g/L after 40 h of fed-batch cultivation

Genome-Scale Reconstruction and Analysis of the Pseudomonas putida KT2440 Metabolic Network Facilitates Applications in Biotechnology

A cornerstone of biotechnology is the use of microorganisms for the efficient production of chemicals and the elimination of harmful waste. Pseudomonas putida is an archetype of such microbes due to its metabolic versatility, stress resistance, amenability to genetic modifications, and vast potential for environmental and industrial applications. To address both the elucidation of the metabolic wiring in P. putida and its uses in biocatalysis, in particular for the production of non-growth-related biochemicals, we developed and present here a genome-scale constraint-based model of the metabolism of P. putida KT2440. Network reconstruction and flux balance analysis (FBA) enabled definition of the structure of the metabolic network, identification of knowledge gaps, and pin-pointing of essential metabolic functions, facilitating thereby the refinement of gene annotations. FBA and flux variability analysis were used to analyze the properties, potential, and limits of the model. These analyses allowed identification, under various conditions, of key features of metabolism such as growth yield, resource distribution, network robustness, and gene essentiality. The model was validated with data from continuous cell cultures, high-throughput phenotyping data, 13C-measurement of internal flux distributions, and specifically generated knock-out mutants. Auxotrophy was correctly predicted in 75% of the cases. These systematic analyses revealed that the metabolic network structure is the main factor determining the accuracy of predictions, whereas biomass composition has negligible influence. Finally, we drew on the model to devise metabolic engineering strategies to improve production of polyhydroxyalkanoates, a class of biotechnologically useful compounds whose synthesis is not coupled to cell survival. The solidly validated model yields valuable insights into genotype–phenotype relationships and provides a sound framework to explore this versatile bacterium and to capitalize on its vast biotechnological potential

Brain death and postmortem organ donation: Report of a questionnaire from the CENTER-TBI study

Background: We aimed to investigate the extent of the agreement on practices around brain death and postmortem organ donation. Methods: Investigators from 67 Collaborative European NeuroTrauma Effectiveness Research in Traumatic Brain Injury (CENTER-TBI) study centers completed several questionnaires (response rate: 99%). Results: Regarding practices around brain death, we found agreement on the clinical evaluation (prerequisites and neurological assessment) for brain death determination (BDD) in 100% of the centers. However, ancillary tests were required for BDD in 64% of the centers. BDD for nondonor patients was deemed mandatory in 18% of the centers before withdrawing life-sustaining measures (LSM). Also, practices around postmortem organ donation varied. Organ donation after circulatory arrest was forbidden in 45% of the centers. When withdrawal of LSM was contemplated, in 67% of centers the patients with a ventricular drain in situ had this removed, either sometimes or all of the time. Conclusions: This study showed both agreement and some regional differences regarding practices around brain death and postmortem organ donation. We hope our results help quantify and understand potential differences, and provide impetus for current dialogs toward further harmonization of practices around brain death and postmortem organ donation