In-situ muconic acid extraction reveals sugar consumption bottleneck in a xylose-utilizing Saccharomyces cerevisiae strain.

BACKGROUND: The current shift from a fossil-resource based economy to a more sustainable, bio-based economy requires development of alternative production routes based on utilization of biomass for the many chemicals that are currently produced from petroleum. Muconic acid is an attractive platform chemical for the bio-based economy because it can be converted in chemicals with wide industrial applicability, such as adipic and terephthalic acid, and because its two double bonds offer great versatility for chemical modification. RESULTS: We have constructed a yeast cell factory converting glucose and xylose into muconic acid without formation of ethanol. We consecutively eliminated feedback inhibition in the shikimate pathway, inserted the heterologous pathway for muconic acid biosynthesis from 3-dehydroshikimate (DHS) by co-expression of DHS dehydratase from P. anserina, protocatechuic acid (PCA) decarboxylase (PCAD) from K. pneumoniae and oxygen-consuming catechol 1,2-dioxygenase (CDO) from C. albicans, eliminated ethanol production by deletion of the three PDC genes and minimized PCA production by enhancing PCAD overexpression and production of its co-factor. The yeast pitching rate was increased to lower high biomass formation caused by the compulsory aerobic conditions. Maximal titers of 4 g/L, 4.5 g/L and 3.8 g/L muconic acid were reached with glucose, xylose, and a mixture, respectively. The use of an elevated initial sugar level, resulting in muconic acid titers above 2.5 g/L, caused stuck fermentations with incomplete utilization of the sugar. Application of polypropylene glycol 4000 (PPG) as solvent for in situ product removal during the fermentation shows that this is not due to toxicity by the muconic acid produced. CONCLUSIONS: This work has developed an industrial yeast strain able to produce muconic acid from glucose and also with great efficiency from xylose, without any ethanol production, minimal production of PCA and reaching the highest titers in batch fermentation reported up to now. Utilization of higher sugar levels remained conspicuously incomplete. Since this was not due to product inhibition by muconic acid or to loss of viability, an unknown, possibly metabolic bottleneck apparently arises during muconic acid fermentation with high sugar levels and blocks further sugar utilization.

Pediatric Airway Endoscopy: Recommendations of the Society for Pediatric Pneumology.

For many decades, pediatric bronchoscopy has been an integral part of the diagnosis and treatment of acute and chronic pulmonary diseases in children. Rapid technical advances have continuously influenced the performance of the procedure. Over the years, the application of pediatric bronchoscopy has considerably expanded to a broad range of indications. In this comprehensive and up-to-date guideline, the Special Interest Group of the Society for Pediatric Pneumology reviewed the most recent literature on pediatric bronchoscopy and reached a consensus on a safe technical performance of the procedure.

Dimensional compatibility of rigid ventilating bronchoscopes with pediatric airway anatomy using different recommendations for age-related sizing-A bench study.

BACKGROUND: Appropriate size selection of pediatric rigid bronchoscopes is fundamental to avoidance of airway trauma and to a high success rate with the first intubation attempt. The aim of the present study was to compare the outer diameters of pediatric rigid bronchoscopes with the anatomical data on the pediatric airway. METHODS: Outer diameters (OD) of pediatric rigid ventilating bronchoscopes as given by the manufacturer were compared with published computed tomography internal diameters (ID) of the cricoid outlet, the trachea as well as the left and right proximal mainstem bronchus, using six published recommendations for age-related size selection. The ratios between a specific OD and ID were calculated and given as percentage values (%) for lower and upper age ranges. RESULTS: Nominal sized 2.5, 3, 3.5, 3.7, 4, 4.5, 5, and 6 rigid bronchoscopes were included. In five recommendations, the rigid bronchoscopes' OD was larger than the internal cricoid diameter in the upper age range, whereas in one recommendation the ratio (OD rigid bronchoscope to ID of cricoid) ranged between 77% and 90% for the upper age and between 95% and 109% for the lower age range. In only one of the six recommendations was the rigid bronchoscope OD larger than the tracheal ID. Ratios for the left and right proximal mainstem bronchus ranged from 70% to 146% for the upper and from 78% to 156% for the lower age range, with the highest ratios for the left proximal mainstem bronchus. CONCLUSION: Based on this in vitro study, most of the recommendations analyzed result in a balanced fit of rigid bronchoscopes within the cricoid. Since the left mainstem bronchus is considerably smaller than the cricoid, any insertion on this level will require careful endoscopic guidance to avoid damage to the left mainstem bronchus.

Classical versus controlled rapid sequence induction and intubation in children with bleeding tonsils (a retrospective audit).

PURPOSE: To determine whether bag-mask ventilation between induction of anaesthesia and tracheal intubation in children with post-tonsillectomy bleeding reduces the incidence of hypoxaemia and difficult direct laryngoscopy without increasing perioperative respiratory complications. METHODS: Medical records, anaesthesia protocols and vital sign data were analysed from February 2005 to March 2017 for patients undergoing anaesthesia for surgical revision of bleeding tonsils. Type of rapid sequence induction and intubation (RSII; classical, ie, apnoeic, vs controlled, ie, with gentle bag-mask ventilation) was noted. Primary outcomes were incidence of moderate and severe hypoxaemia, grade of direct laryngoscopic views as well as occurrence of noted tracheal intubation difficulties. Haemodynamic alterations during RSII and perioperative adverse events such as noted gastric regurgitation, pulmonary aspiration and perioperative pulmonary morbidity were also recorded. RESULTS: A classical RSII was performed for 22 surgical revisions in 22 children and a controlled RSII was used for 88 surgical revisions in 81 children. Patients undergoing controlled RSII had less incidence of severe hypoxaemia (1 vs 3; P = .025), better direct laryngoscopic views (P = .048) and less hypertension (5 vs 9; P < .001) than those patients managed by classical RSII. No tracheal intubation difficulties occurred. There was no significant perioperative pulmonary morbidity reported in either group. CONCLUSIONS: Controlled RSII had advantages over classical RSII in children with post-tonsillectomy bleeding and may become a strategic option in these patients to avoid hypoxaemia, difficult laryngoscopy and hypertension during induction of anaesthesia and tracheal intubation. Bag-mask ventilation in patients with bleeding tonsils did not lead to pulmonary morbidity.

Assessment of the multiplex PCR-based assay Unyvero pneumonia application for detection of bacterial pathogens and antibiotic resistance genes in children and neonates.

BACKGROUND: Pneumonia is a major healthcare problem. Rapid pathogen identification is critical, but often delayed due to the duration of culturing. Early, broad antibacterial therapy might lead to false-negative culture findings and eventually to the development of antibiotic resistances. We aimed to assess the accuracy of the new application Unyvero P50 based on multiplex PCR to detect bacterial pathogens in respiratory specimens from children and neonates. METHODS: In this prospective study, bronchoalveolar lavage fluids, tracheal aspirates, or pleural fluids from neonates and children were analyzed by both traditional culture methods and Unyvero multiplex PCR. RESULTS: We analyzed specimens from 79 patients with a median age of 1.8 (range 0.01-20.1). Overall, Unyvero yielded a sensitivity of 73.1% and a specificity of 97.9% compared to culture methods. Best results were observed for non-fermenting bacteria, for which sensitivity of Unyvero was 90% and specificity 97.3%, while rates were lower for Gram-positive bacteria (46.2 and 93.9%, respectively). For resistance genes, we observed a concordance with antibiogram of 75% for those specimens in which there was a cultural correlate. CONCLUSIONS: Unyvero is a fast and easy-to-use tool that might provide additional information for clinical decision making, especially in neonates and in the setting of nosocomial pneumonia. Sensitivity of the PCR for Gram-positive bacteria and important resistance genes must be improved before this application can be widely recommended.

ERS statement: interventional bronchoscopy in children.

Paediatric airway endoscopy is accepted as a diagnostic and therapeutic procedure, with an expanding number of indications and applications in children. The aim of this European Respiratory Society task force was to produce a statement on interventional bronchoscopy in children, describing the evidence available at present and current clinical practice, and identifying areas deserving further investigation. The multidisciplinary task force panel performed a systematic review of the literature, focusing on whole lung lavage, transbronchial and endobronchial biopsy, transbronchial needle aspiration with endobronchial ultrasound, foreign body extraction, balloon dilation and occlusion, laser-assisted procedures, usage of airway stents, microdebriders, cryotherapy, endoscopic intubation, application of drugs and other liquids, and caregiver perspectives. There is a scarcity of published evidence in this field, and in many cases the task force had to resort to the collective clinical experience of the committee to develop this statement. The highlighted gaps in knowledge underline the need for further research and serve as a call to paediatric bronchoscopists to work together in multicentre collaborations, for the benefit of children with airway disorders.

Endoscopic airway findings in children with or without prior endotracheal intubation.

BACKGROUND: Airway alterations found after endotracheal intubation are usually associated with mechanical trauma from the tube. However, no studies are available concerning alterations in airways that have never been intubated before. It was the aim of the study to compare endoscopic findings in the larynx and trachea of children who had undergone prior endotracheal intubation with findings in children who had not been intubated before. METHODS: In 1021 children aged from 0 to 6 years, rigid endoscopies were performed before planned elective endotracheal intubation. The anonymized endoscopy videos were reviewed and graded by five international airway experts. Data was compared between the two groups using the chi-square test (P ≤ 0.05). RESULTS: Endoscopic records of 971 children (473 with prior intubation; 498 without prior airway intubation) were included in the final calculations. Most patients (93.7%) with prior intubation had been intubated with a cuffed tube. The number of intubations ranged from 1 to 27. The median interval between intubation and endoscopy was 0.53 years (0.003-5.57 years). Abnormal findings were observed in 31.7% and 26.8% of patients with and without prior intubation, respectively (P = 0.063). Glottic granulomas were significantly more common after intubation (3.6% vs 1.4%; P = 0.028). The incidence of other abnormal findings was similar in both groups. CONCLUSION: Endoscopic airway alterations can be observed in about one-quarter of children presenting for routine surgery without prior intubation. Except for glottic granulomas, the abnormalities are found with similar frequency in patients with and without prior intubation. No relevant airway damage from short-term endotracheal intubation was found.

[Current concepts of weaning children from invasive ventilation]. / Respiratorentwöhnung--Definition und klinischer Kontext aus Sicht der Pädiatrie.

Timely weaning from invasive ventilation is of major importance to limit time of invasive ventilation and improve outcomes. However, in pediatrics only limited knowledge on the optimal weaning approach is available. In this review evidence from recent trials on weaning in pediatrics is summarized. Standardized daily evaluation of weaning readiness, daily interruption of sedation, use of pediatric sedation protocols, application of noninvasive ventilation and prophylactic treatment with steroids of patients with high risk for post-extubation upper airway obstruction have been shown to decrease duration of invasive ventilation or to decrease the risk of extubation failure. However, due to the heterogeneous patient population in pediatric intensive care units, an individual approach might be necessary for certain subgroups of pediatric patients.

Long-term follow-up and treatment of congenital alveolar proteinosis.

BACKGROUND: Clinical presentation, diagnosis, management and outcome of molecularly defined congenital pulmonary alveolar proteinosis (PAP) due to mutations in the GM-CSF receptor are not well known. CASE PRESENTATION: A 2 1/2 years old girl was diagnosed as having alveolar proteinosis. Whole lung lavages were performed with a new catheter balloon technique, feasible in small sized airways. Because of some interstitial inflammation in the lung biopsy and to further improve the condition, empirical therapy with systemic steroids and azathioprin, and inhaled and subcutaneous GMCSF, were used. Based on clinical measures, total protein and lipid recovered by whole lung lavages, all these treatments were without benefit. Conversely, severe respiratory viral infections and an invasive aspergillosis with aspergilloma formation occurred. Recently the novel homozygous stop mutation p.Ser25X of the GMCSF receptor alpha chain was identified in the patient. This mutation leads to a lack of functional GMCSF receptor and a reduced response to GMCSF stimulation of CD11b expression of mononuclear cells of the patient. Subsequently a very intense treatment with monthly lavages was initiated, resulting for the first time in complete resolution of partial respiratory insufficiency and a significant improvement of the overall somato-psychosocial condition of the child. CONCLUSIONS: The long term management from early childhood into young adolescence of severe alveolar proteinosis due to GMCSF receptor deficiency requires a dedicated specialized team to perform technically demanding whole lung lavages and cope with complications.

Heterozygous SOX9 mutations allowing for residual DNA-binding and transcriptional activation lead to the acampomelic variant of campomelic dysplasia.

Campomelic dysplasia is a malformation syndrome with multiple symptoms including characteristic shortness and bowing of the long bones (campomelia). CD, often lethal due to airway malformations, is caused by heterozygous mutations in SOX9, an SRY-related gene regulating testis and chondrocyte development including expression of many cartilage genes such as type II collagen. Male to female sex reversal occurs in the majority of affected individuals with an XY karyotype. A mild form without campomelia exists, in which sex-reversal may be also absent. We report here two novel SOX9 missense mutations in a male (c.495C>G; p.His165Gln) and a female (c.337A>G; p.Met113Val) within the DNA-binding domain leading to non-lethal acampomelic CD. Functional analyses of mutant proteins demonstrate residual DNA-binding and transactivation of SOX9-regulated genes. Combining our data and reports from the literature we postulate a genotype-phenotype correlation: SOX9 mutations allowing for residual function lead to a mild form of CD in which campomelia and sex reversal may be absent.

Expression, regulation and clinical significance of soluble and membrane CD14 receptors in pediatric inflammatory lung diseases.

BACKGROUND: Inflammatory lung diseases are a major morbidity factor in children. Therefore, novel strategies for early detection of inflammatory lung diseases are of high interest. Bacterial lipopolysaccharide (LPS) is recognized via Toll-like receptors and CD14. CD14 exists as a soluble (sCD14) and membrane-associated (mCD14) protein, present on the surface of leukocytes. Previous studies suggest sCD14 as potential marker for inflammatory diseases, but their potential role in pediatric lung diseases remained elusive. Therefore, we examined the expression, regulation and significance of sCD14 and mCD14 in pediatric lung diseases. METHODS: sCD14 levels were quantified in serum and bronchoalveolar lavage fluid (BALF) of children with infective (pneumonia, cystic fibrosis, CF) and non-infective (asthma) inflammatory lung diseases and healthy control subjects by ELISA. Membrane CD14 expression levels on monocytes in peripheral blood and on alveolar macrophages in BALF were quantified by flow cytometry. In vitro studies were performed to investigate which factors regulate sCD14 release and mCD14 expression. RESULTS: sCD14 serum levels were specifically increased in serum of children with pneumonia compared to CF, asthma and control subjects. In vitro, CpG induced the release of sCD14 levels in a protease-independent manner, whereas LPS-mediated mCD14 shedding was prevented by serine protease inhibition. CONCLUSIONS: This study demonstrates for the first time the expression, regulation and clinical significance of soluble and membrane CD14 receptors in pediatric inflammatory lung diseases and suggests sCD14 as potential marker for pneumonia in children.

Whole-lung lavage in infants and children with pulmonary alveolar proteinosis.

BACKGROUND: Pulmonary alveolar proteinosis (PAP) is a rare alveolar filling syndrome where the mainstay of treatment is therapeutic whole-lung lavage (WLL). WLL techniques used in adults have to be modified for children because of their small-diameter airways. AIM: To describe a technique for WLL adapted for small children. METHODS: We describe a WLL technique that combines safe single-lung ventilation with the use of an age-appropriate endotracheal tube and selective occlusion of the other main bronchus with a balloon catheter through which the lavage is performed. Effectiveness measured by change in oxygen requirements and adverse effects was noted. RESULTS: We performed 64 WLL procedures in four children (age 13 months to 7 years; body weight 4.7-14 kg). Two children had idiopathic and two had secondary PAP. At referral, all children had dyspnoea at rest and required continuous oxygen supplementation. Two patients showed significantly decreased oxygen demands and radiological improvement after WLL. Two patients showed no significant response. The only adverse effect observed was transient hypoxemia. Complications comprised fluid leak at the balloon (4), balloon rupture (1), and pneumothorax (1). CONCLUSIONS: This technique for WLL combining single-lung ventilation with an endotracheal tube and lung exclusion for lavage with a balloon catheter can be safely and effectively performed in small children with PAP.

A sustainable wood biorefinery for low-carbon footprint chemicals production.

The profitability and sustainability of future biorefineries are dependent on efficient feedstock use. Therefore, it is essential to valorize lignin when using wood. We have developed an integrated biorefinery that converts 78 weight % (wt %) of birch into xylochemicals. Reductive catalytic fractionation of the wood produces a carbohydrate pulp amenable to bioethanol production and a lignin oil. After extraction of the lignin oil, the crude, unseparated mixture of phenolic monomers is catalytically funneled into 20 wt % of phenol and 9 wt % of propylene (on the basis of lignin weight) by gas-phase hydroprocessing and dealkylation; the residual phenolic oligomers (30 wt %) are used in printing ink as replacements for controversial para-nonylphenol. A techno-economic analysis predicts an economically competitive production process, and a life-cycle assessment estimates a lower carbon dioxide footprint relative to that of fossil-based production.

Pediatric pulmonology year in review 2017: Part 1.

Pediatric Pulmonology publishes original research, case reports and review articles on topics related to a wide range of children's respiratory disorders. In this article (Part 1 of a series), we summarize the past year's publications in our major topic areas, as well as selected literature in these areas from other journals. In Part 1, we review selected articles on diagnostic testing/endoscopy, respiratory complications of neuromuscular disorders, and rare lung diseases.

Childhood Stroke: Awareness, Interest, and Knowledge Among the Pediatric Community.

Objective: Acute childhood stroke is an emergency requiring a high level of awareness among first-line healthcare providers. This survey serves as an indicator of the awareness of, the interest in, and knowledge of childhood stroke of German pediatricians. Methods: Thousand six hundred and ninety-seven physicians of pediatric in- and outpatient facilities in Bavaria, Germany, were invited via email to an online-survey about childhood stroke. Results: The overall participation rate was 14%. Forty-six percent of participants considered a diagnosis of childhood stroke at least once during the past year, and 47% provide care for patients who have suffered childhood stroke. The acronym FAST (Face-Arm-Speech-Time-Test) was correctly cited in 27% of the questionnaires. Most commonly quoted symptoms of childhood stroke were hemiparesis (90%), speech disorder (58%), seizure (44%), headache (40%), and impaired consciousness (33%). Migraine (63%), seizure (39%), and infections of the brain (31%) were most frequently named as stroke mimics. Main diagnostic measures indicated were magnetic resonance imaging (MRI) (96%) and computer tomography (CT) (55%). Main therapeutic strategies were thrombolysis (80%), anticoagulation (41%), neuroprotective measures, and thrombectomies (15% each). Thirty-nine percent of participants had taken part in training sessions, 61% studied literature, 37% discussed with colleagues, and 25% performed internet research on childhood stroke. Ninety-three percent of participants approve skill enhancement, favoring training sessions (80%), publications (43%), and web based offers (35%). Consent for offering a flyer on the topic to caregivers in facilities was given in 49%. Conclusion: Childhood stroke constitutes a topic of clinical importance to pediatricians. Participants demonstrate a considerable level of comprehension concerning the subject, but room for improvement remains. A multi-modal approach encompassing an elaborate training program, regular educational publications in professional journals, and web based offers could reach a broad range of health care providers. Paired with a public adult and childhood stroke awareness campaign, these efforts could contribute to optimize the care for children suffering from stroke.

Foreign body removal in children: Recommendations versus real life-A survey of current clinical management in Germany.

BACKGROUND: Task forces of the European Respiratory Society (ERS, 2003) and the American Thoracic Society (ATS, 2015) both recommend rigid bronchoscopy for foreign body removal in children. However, there is an increasing number of publications recommending flexible bronchoscopy as primary method for foreign body removal in children. METHODS: We therefore initiated a nationwide survey, investigating the routinely used method of foreign body removal in children in different centers in Germany. RESULTS: In total, 259 centers took part in the survey. Twenty percent used flexible bronchoscopy as primary method of choice for foreign body removal. Of note, only 40% of the respondents reported that they had attended formal training courses in flexible bronchoscopy, 15% in rigid bronchoscopy, and 42% had never attended a formal training. Twenty-five percent indicated that they had already encountered an emergency situation during or after foreign body removal, requiring intensive care. CONCLUSION: Published data shows that rigid bronchoscopy involves fewer complications and is more successful in foreign body removal in children. It is consequently questionable, why 20% of the medical centers report a preference for the flexible method. One reason might be the poor availability of rigid bronchoscopy and the difficulty gaining sufficient expertise in rigid bronchoscopy in Germany. Pediatr Pulmonol. 2017;52:656-661. © 2016 Wiley Periodicals, Inc.

Pediatric Pulmonology year in review 2016: Part 1.

Pediatric Pulmonology continues to publish research and clinical topics related to the entire range of children's respiratory disorders. As we have done annually in recent years, we here summarize the past year's publications in our major topic areas, as well as selected literature in these areas from other core journals relevant to our discipline. This review (Part 1) covers selected articles on sleep, diagnostic testing/endoscopy, respiratory complications of neuromuscular disorders, and rare lung diseases.