Microbial synthesis of wax esters.

Microbial synthesis of wax esters (WE) from low-cost renewable and sustainable feedstocks is a promising path to achieve cost-effectiveness in biomanufacturing. WE are industrially high-value molecules, which are widely used for applications in chemical, pharmaceutical, and food industries. Since the natural WE resources are limited, the WE production mostly rely on chemical synthesis from rather expensive starting materials, and therefore solution are sought from development of efficient microbial cell factories. Here we report to engineer the yeast Yarrowia lipolytica and bacterium Escherichia coli to produce WE at the highest level up to date. First, the key genes encoding fatty acyl-CoA reductases and wax ester synthase from different sources were investigated, and the expression system for two different Y. lipolytica hosts were compared and optimized for enhanced WE production and the strain stability. To improve the metabolic pathway efficiency, different carbon sources including glucose, free fatty acid, soybean oil, and waste cooking oil (WCO) were compared, and the corresponding pathway engineering strategies were optimized. It was found that using a lipid substrate such as WCO to replace glucose led to a 60-fold increase in WE production. The engineered yeast was able to produce 7.6 g/L WE with a yield of 0.31 (g/g) from WCO within 120 h and the produced WE contributed to 57% of the yeast DCW. After that, E. coli BL21(DE3), with a faster growth rate than the yeast, was engineered to significantly improve the WE production rate. Optimization of the expression system and the substrate feeding strategies led to production of 3.7-4.0 g/L WE within 40 h in a 1-L bioreactor. The predominant intracellular WE produced by both Y. lipolytica and E. coli in the presence of hydrophobic substrates as sole carbon sources were C36, C34 and C32, in an order of decreasing abundance and with a large proportion being unsaturated. This work paved the way for the biomanufacturing of WE at a large scale.

Using oils and fats to replace sugars as feedstocks for biomanufacturing: Challenges and opportunities for the yeast Yarrowia lipolytica.

More than 200 million tons of plant oils and animal fats are produced annually worldwide from oil, crops, and the rendered animal fat industry. Triacylglycerol, an abundant energy-dense compound, is the major form of lipid in oils and fats. While oils or fats are very important raw materials and functional ingredients for food or related products, a significant portion is currently diverted to or recovered as waste. To significantly increase the value of waste oils or fats and expand their applications with a minimal environmental footprint, microbial biomanufacturing is presented as an effective strategy for adding value. Though both bacteria and yeast can be engineered to use oils or fats as the biomanufacturing feedstocks, the yeast Yarrowia lipolytica is presented as one of the most attractive platforms. Y. lipolytica is oleaginous, generally regarded as safe, demonstrated as a promising industrial producer, and has unique capabilities for efficient catabolism and bioconversion of lipid substrates. This review summarizes the major challenges and opportunities for Y. lipolytica as a new biomanufacturing platform for the production of value-added products from oils and fats. This review also discusses relevant cellular and metabolic engineering strategies such as fatty acid transport, fatty acid catabolism and bioconversion, redox balances and energy yield, cell morphology and stress response, and bioreaction engineering. Finally, this review highlights specific product classes including long-chain diacids, wax esters, terpenes, and carotenoids with unique synthesis opportunities from oils and fats in Y. lipolytica.

Enzyme selection, optimization, and production toward biodegradation of post-consumer poly(ethylene terephthalate) at scale.

Poly(ethylene terephthalate) (PET) is one of the world's most widely used polyester plastics. Due to its chemical stability, PET is extremely difficult to hydrolyze in a natural environment. Recent discoveries in new polyester hydrolases and breakthroughs in enzyme engineering strategies have inspired enormous research on biorecycling of PET. This study summarizes our research efforts toward large-scale, efficient, and economical biodegradation of post-consumer waste PET, including PET hydrolase selection and optimization, high-yield enzyme production, and high-capacity enzymatic degradation of post-consumer waste PET. First, genes encoding PETase and MHETase from Ideonella sakaiensis and the ICCG variant of leaf-branch compost cutinase (LCCICCG ) were codon-optimized and expressed in Escherichia coli BL21(DE3) for high-yield production. To further lower the enzyme production cost, a pelB leader sequence was fused to LCCICCG so that the enzyme can be secreted into the medium to facilitate recovery. To help bind the enzyme on the hydrophobic surface of PET, a substrate-binding module in a polyhydroxyalkanoate depolymerase from Alcaligenes faecalis (PBM) was fused to the C-terminus of LCCICCG . The resulting four different LCCICCG variants (LCC, PelB-LCC, LCC-PBM, and PelB-LCC-PBM), together with PETase and MHETase, were compared for PET degradation efficiency. A fed-batch fermentation process was developed to produce the target enzymes up to 1.2 g L-1 . Finally, the best enzyme, PelB-LCC, was selected and used for the efficient degradation of 200 g L-1 recycled PET in a well-controlled, stirred-tank reactor. The results will help develop an economical and scalable biorecycling process toward a circular PET economy.

Consensus in diagnostic definitions for bone or joint infections in children by a Delphi method with European French-speaking experts.

AIM: Various diagnostic criteria have been proposed for bone or joint infection. This study used a Delphi process to determine the consensual definitions for arthritis, osteomyelitis and bone or joint infections in general in children. METHODS: A group of European French-speaking experts participated in an email Delphi process. Definitions were identified during a systematic search of the PubMed database. Five definitions of arthritis, eight for osteomyelitis and five for bone or joint infections in general were included in a three-round process. We sought two sorts of definitions: definitions for 'definitive' diagnoses for epidemiological studies and definitions for 'probable' diagnoses for clinical or therapeutic studies, considering enlarged criteria. RESULTS: Ten experts were involved in the Delphi process. A consensus was reached for a definitive diagnosis of arthritis, osteomyelitis and bone or joint infections in general. A consensus was also reached for a probable diagnosis of bone or joint infections in general. CONCLUSION: This Delphi process made consensus definitions and criteria available for bone or joint infections that could improve the comparability of both epidemiological and clinical studies. This is a first step to standardise diagnostic criteria and distinguish definitive and probable bone or joint infections in children.

Recent Advances in Biological Recycling of Polyethylene Terephthalate (PET) Plastic Wastes.

Polyethylene terephthalate (PET) is one of the most commonly used polyester plastics worldwide but is extremely difficult to be hydrolyzed in a natural environment. PET plastic is an inexpensive, lightweight, and durable material, which can readily be molded into an assortment of products that are used in a broad range of applications. Most PET is used for single-use packaging materials, such as disposable consumer items and packaging. Although PET plastics are a valuable resource in many aspects, the proliferation of plastic products in the last several decades have resulted in a negative environmental footprint. The long-term risk of released PET waste in the environment poses a serious threat to ecosystems, food safety, and even human health in modern society. Recycling is one of the most important actions currently available to reduce these impacts. Current clean-up strategies have attempted to alleviate the adverse impacts of PET pollution but are unable to compete with the increasing quantities of PET waste exposed to the environment. In this review paper, current PET recycling methods to improve life cycle and waste management are discussed, which can be further implemented to reduce plastics pollution and its impacts on health and environment. Compared with conventional mechanical and chemical recycling processes, the biotechnological recycling of PET involves enzymatic degradation of the waste PET and the followed bioconversion of degraded PET monomers into value-added chemicals. This approach creates a circular PET economy by recycling waste PET or upcycling it into more valuable products with minimal environmental footprint.

Increased detection rate of Kawasaki disease using new diagnostic algorithm, including early use of echocardiography.

OBJECTIVE: To determine the incidence of Kawasaki disease (KD) in Northern France by using new American Heart Association (AHA) criteria. STUDY DESIGN: A 1-year prospective multicenter cohort study was performed in all pediatric departments. Patients <18 years old, who were admitted for prolonged but initially unexplained fever or suspected KD were included. All patients received the standard treatment considered appropriate by their physicians. A descriptive analysis and comparison of patients with complete and incomplete forms of KD were performed. The incidence of confirmed cases of KD (complete and incomplete forms) was calculated. RESULTS: Seventy-seven children were included (39 in whom KD was diagnosed). Of the patients with KD, 26 (67%) met the classic AHA case definition, and 7 (18%) had incomplete KD. Cardiac ultrasound scanning was helpful in the diagnosis of 6 of 7 patients with incomplete KD (86%). The final incidence of confirmed KD was 9 of 100,000 children <5 years of age. In 6 children (15%) the diagnosis of KD was uncertain, but they were successfully treated for it. Coronary disease was identified in 48% of patients with confirmed KD. CONCLUSION: The incidence of KD is higher than previously reported, in part because earlier reports did not include incomplete forms. New AHA criteria (laboratory tests and early echocardiography) were helpful for the diagnosis of incomplete forms of KD.

Cellular and metabolic engineering of oleaginous yeast Yarrowia lipolytica for bioconversion of hydrophobic substrates into high-value products.

The non-conventional oleaginous yeast Yarrowia lipolytica is able to utilize both hydrophilic and hydrophobic carbon sources as substrates and convert them into value-added bioproducts such as organic acids, extracellular proteins, wax esters, long-chain diacids, fatty acid ethyl esters, carotenoids and omega-3 fatty acids. Metabolic pathway analysis and previous research results show that hydrophobic substrates are potentially more preferred by Y. lipolytica than hydrophilic substrates to make high-value products at higher productivity, titer, rate, and yield. Hence, Y. lipolytica is becoming an efficient and promising biomanufacturing platform due to its capabilities in biosynthesis of extracellular lipases and directly converting the extracellular triacylglycerol oils and fats into high-value products. It is believed that the cell size and morphology of the Y. lipolytica is related to the cell growth, nutrient uptake, and product formation. Dimorphic Y. lipolytica demonstrates the yeast-to-hypha transition in response to the extracellular environments and genetic background. Yeast-to-hyphal transition regulating genes, such as YlBEM1, YlMHY1 and YlZNC1 and so forth, have been identified to involve as major transcriptional factors that control morphology transition in Y. lipolytica. The connection of the cell polarization including cell cycle and the dimorphic transition with the cell size and morphology in Y. lipolytica adapting to new growth are reviewed and discussed. This review also summarizes the general and advanced genetic tools that are used to build a Y. lipolytica biomanufacturing platform.

Critical and reasoned review of the psychosomatic question in French-speaking psychoanalysts.

European psychosomatics, as a branch of psychoanalytic psychopathology, has existed for almost two centuries, but it is a constantly moving body of thought. From epistemology to aetiology, its questions are still evolving and concern the debate between the psyche/soma dualism and the monistic alternative, but also the psychic meaning that is to be attributed to somatic symptoms. In France, the discipline has not had its last word, for it is constantly seeking to position itself as a field of research and clinical practice within psychoanalysis. However, it still needs to affirm itself and open itself more towards other branches of knowledge, such as medicine, neuroscience, or the social sciences. In this context, are the processes of somatisation still to be thought of as boundary objects for psychoanalysis? On what epistemological, theoretical, methodological, and practical conditions is the psychosomatic question still or at last possible? In particular, is it possible to reintroduce the question of the "meaning" of the somatic phenomenon in order to breathe new life into it, without returning, however, to the symbolic interpretation of symptoms, while preserving the subjectivising approach of psychoanalysis and its logic of interpolation? Can we speak in the human sciences of a truly psychosomatic revolution in the image of psychoanalytic advances?

[Impact of the consensus conference on the ambulatory treatment of bronchiolitis in infants]. / Impact de la conférence de consensus sur le traitement ambulatoire des bronchiolites du nourrisson.

OBJECTIVE: To assess the impact of guidelines of the consensus conference in September 2000 on the ambulatory management of acute bronchiolitis in infants. METHODS: Prospective multicenter study, in four hospitals in the North department in France, during three epidemic periods: the winter preceding the consensus conference and the two following winters. All the infants between 30 days and 2 years of age, admitted to an emergency care unit for a first or second episode of bronchiolitis were included. The infants' characteristics and ambulatory treatments prescribed were collected. RESULTS: Six hundred thirty-eight infants were included: 169 before the conference and 469 after. Seventy-seven percent had consulted previously and a prescription had been drawn-up for 77%. There was no difference in the frequency of drug prescriptions or physiotherapy between the three periods. After the conferences, inhaled beta(2) agonists were still prescribed in 21% of cases, inhaled corticosteroids in 15%, oral corticosteroids in 34%, antibiotics in 53% and mucolytics in 54%. Physiotherapy was prescribed for 58% of the infants. CONCLUSION: Two years after the consensus conference, the ambulatory treatment of bronchiolitis has no changed, corticosteroids, beta(2) agonists, antibiotics and mucolytics are still prescribed in excess. Further studies, within a few years, are required to reassess the application of the guidelines, not only in outpatient but also in inpatient management.

Educating Asthmatic Children in European Ambulatory Pediatrics: Facts and Insights.

The aim of this study was to assess the role of European ambulatory pediatricians in caring for asthmatic children, especially in terms of their therapeutic education. We developed a survey that was observational, declarative, retrospective and anonymous in nature. 436 ambulatory pediatricians in Belgium, France, Germany, Italy, Luxembourg and Slovenia were asked to participate in the survey providing information on three children over 6 years old suffering from persistent asthma, who had been followed for at least 6 months. We considered the pediatricians' profile, and their role in the therapeutic education of children. 277 pediatricians (64%) responded: 81% were primary care pediatricians; 46% participated in networks; 4% had specific training in Therapeutic Patient Education; 69% followed more than 5 asthmatic children per month, and over long periods (7 ± 4 years). The profiles of 684 children were assessed. Answers diverged concerning the provision of a Personalized Action Plan (60-88%), training the child to measure and interpret his Peak Expiratory Flow (31-99%), and the prescription of pulmonary function tests during the follow-up programme of consultations (62-97%). Answers converged on pediatricians' perception of their role in teaching children about their condition and its treatment (99%), about inhalation techniques (96%), and in improving the children's ability to take preventive measures when faced with risk situations (97%). This study highlights the role of European pediatricians in caring for asthmatic children, and their lack of training in Therapeutic Patient Education. Programmes and tools are required in order to train ambulatory pediatricians in Therapeutic Patient Education, and such resources should be integrated into primary health care, and harmonized at the European level.

The value of body weight measurement to assess dehydration in children.

Dehydration secondary to gastroenteritis is one of the most common reasons for office visits and hospital admissions. The indicator most commonly used to estimate dehydration status is acute weight loss. Post-illness weight gain is considered as the gold-standard to determine the true level of dehydration and is widely used to estimate weight loss in research. To determine the value of post-illness weight gain as a gold standard for acute dehydration, we conducted a prospective cohort study in which 293 children, aged 1 month to 2 years, with acute diarrhea were followed for 7 days during a 3-year period. The main outcome measures were an accurate pre-illness weight (if available within 8 days before the diarrhea), post-illness weight, and theoretical weight (predicted from the child's individual growth chart). Post-illness weight was measured for 231 (79%) and both theoretical and post-illness weights were obtained for 111 (39%). Only 62 (21%) had an accurate pre-illness weight. The correlation between post-illness and theoretical weight was excellent (0.978), but bootstrapped linear regression analysis showed that post-illness weight underestimated theoretical weight by 0.48 kg (95% CI: 0.06-0.79, p<0.02). The mean difference in the fluid deficit calculated was 4.0% of body weight (95% CI: 3.2-4.7, p<0.0001). Theoretical weight overestimated accurate pre-illness weight by 0.21 kg (95% CI: 0.08-0.34, p = 0.002). Post-illness weight underestimated pre-illness weight by 0.19 kg (95% CI: 0.03-0.36, p = 0.02). The prevalence of 5% dehydration according to post-illness weight (21%) was significantly lower than the prevalence estimated by either theoretical weight (60%) or clinical assessment (66%, p<0.0001).These data suggest that post-illness weight is of little value as a gold standard to determine the true level of dehydration. The performance of dehydration signs or scales determined by using post-illness weight as a gold standard has to be reconsidered.

Risk of bacterial meningitis in young children with a first seizure in the context of fever: a systematic review and meta-analysis.

BACKGROUND: Of major concern in any febrile child presenting with a seizure is the possibility of bacterial meningitis (BM). We did a systematic review to estimate the risk of BM among various subgroups of young children with a first seizure in the context of fever, and to assess the utility of routine lumbar puncture (LP) in children with an apparent first FS. METHODS/PRINCIPAL FINDINGS: MEDLINE, INIST, and the COCHRANE Library databases were searched from inception to December 2011 for published studies, supplemented by manual searches of bibliographies of potentially relevant articles and review articles. Studies reporting the prevalence of BM in young children presenting to emergency care with a first: i) "seizure and fever", ii) apparent simple FS, and iii) apparent complex FS were included. Fourteen studies met the inclusion criteria. In children with a first "seizure and fever", the pooled prevalence of BM was 2.6% (95% CI 0.9-5.1); the diagnosis of BM might be suspected from clinical examination in 95% of children >6 months. In children with an apparent simple FS, the average prevalence of BM was 0.2% (range 0 to 1%). The pooled prevalence of BM among children with an apparent complex FS was 0.6% (95% CI 0.2-1.4). The utility of routine LP for diagnosis of CNS infections requiring immediate treatment in children with an apparent first FS was low: the number of patients needed to test to identify one case of such infections was 1109 in children with an apparent first simple FS, and 180 in those with an apparent first complex FS. CONCLUSION: The values provided from this study provide a basis for an evidence-based approach to the management of different subgroups of children presenting to emergency care with a first seizure in the context of fever.

Bacterial skin infections in children hospitalized with varicella: a possible negative impact of non-steroidal anti-inflammatory drugs?

This 1-year multicentre prospective study in northern France sought to evaluate the incidence of secondary bacterial skin complications related to varicella, describe these superinfections, and analyse risk factors for their onset. The study included every child admitted to a district paediatric unit with a varicella infection. Patients with varicella infection, with and without secondary bacterial skin complication, were compared. The study included 159 children, 43 of whom had a secondary bacterial skin complication on admission, 21 of them had a severe secondary bacterial skin complication (respective incidence: 7.5 and 3.7/100,000 children younger than 16 years old). Persistence or recurrence of fever > or =38.5 degrees C for > or =3 days after the beginning of varicella infection (adjusted odds ratio (aOR)=8.1; 95% confidence interval (CI): 2.3-28.4) and the use of non-steroidal anti-inflammatory drugs (aOR=4.8; 95% CI: 1.6-14.4) were independent factors associated with severe secondary bacterial skin complication.

[Value of history and clinical and laboratory data for the diagnosis of dehydration due to acute diarrhea in children younger than 5 years]. / Valeur des données anamnestiques, cliniques et biologiques pour le diagnostic de déshydratation par diarrhée aiguë chez l'enfant de moins de 5 ans.

BACKGROUND: Acute diarrhea is frequent, costly because of the number of hospital admissions required, and sometimes serious, even fatal to children in France. The clinical diagnosis of dehydration is difficult, but essential to determine management. OBJECTIVE: To summarize the published data on the value of clinical history, clinical signs and laboratory results for diagnosing dehydration during acute diarrhea in young (1 month-5 years) non-malnourished children. METHODS: Four databases (Medline, INIST, Ovid, and Cochrane) were searched through November 2006, with the key words "dehydration" subcategories "diagnosis, or etiology, or history", "diarrhea" subcategory "diagnosis", and age limits "infant or preschool child". We selected the articles and reviews that included as an endpoint for dehydration "weight gain > 5% after recovery" (the gold standard). RESULTS: Thirteen studies were selected. No single clinical history item, clinical sign or laboratory value was sufficient to discriminate between children with and without dehydration. The reproducibility of clinical signs varied substantially between studies. Persistent skin folds and signs of vasoconstriction contributed the most information, with good specificity but sensitivity < 50%. The combination of at least 3 clinical signs was most discriminative for dehydration. No dehydration scale has been validated. None of the studies selected had a very high level of proof (level 1 and 2); neither signs nor scores have been validated internally or externally because of the low number of subjects. CONCLUSION: The diagnosis of dehydration due to acute diarrhea in young children depends on the number of signs present, since no individual element of clinical history, clinical picture or laboratory tests distinguished dehydration. Other studies are necessary.

Paediatric skin disorders encountered in an emergency hospital facility: a prospective study.

To determine the frequency of skin disorders encountered in a paediatric emergency care unit and to evaluate the benefits of advice from a dermatologist, we prospectively recorded data of children admitted with skin disorders to the emergency care unit during a 5-month period. Diagnostic agreement between paediatricians and dermatologists evaluating the patients separately was assessed. Three hundred and ninety-five children (median age 3 years; interquartile 1-6) were included. Skin disorders represented 4% of all paediatric emergency care unit visits. Visits were considered as appropriate in 19-30% of cases according to different criteria. Six diseases accounted for 57% of cases: viral exanthema, urticaria, atopic dermatitis, varicella, diaper dermatitis and herpetic gingivostomatitis. The dermatologist modified the diagnosis in 42% of cases and the treatment in 30%. Greater emphasis on teaching the skin disorders encountered in this setting and efforts to provide easy access to advice from dermatologist would improve the quality of care.