Bacterial consortia in acid mine groundwater seepage: final report

What is expected to be there (bacteria) is there and much more. This study reiterates the versatility and powerfulness of the 16s rDNA sequencing technology in the identification of microbial community in a natural environmental setting without prior cultivation. The challenge at hand is to assess activity and abundance of the culprit and beneficial bacteria in the groundwater seepage path and develop a knowledge-based strategy for the reduction of acidity to combat acid mine drainage problem. The importance of a multidisciplinary approach in this effort cannot be overemphasized

Camphor pathway redux: functional recombinant expression of 2,5- and 3,6-diketocamphane monooxygenases of Pseudomonas putida ATCC 17453 with their cognate flavin reductase catalyzing Baeyer-Villiger reactions

Whereas the biochemical properties of the monooxygenase components that catalyze the oxidation of 2,5-diketocamphane and 3,6-diketocamphane (2,5-DKCMO and 3,6-DKCMO, respectively) in the initial catabolic steps of (+) and (−) isomeric forms of camphor metabolism in Pseudomonas putida ATCC 17453 are relatively well characterized, the actual identity of the flavin reductase (Fred) component that provides the reduced flavin to the oxygenases is hitherto ill-defined. In this study, a 37-kDa Fred was purified from camphor-induced culture of P. putida ATCC 17453 and this facilitated cloning and characterization of the requisite protein. The active Fred is a homodimer with a subunit molecular mass of 18-kDa that uses NADH as electron donor (Km = 32 μM) and it catalyzes the reduction of FMN (Km = 3.6 μM; kcat = 283 s-1) in preference to FAD (Km = 19 μM; kcat = 128 s-1). Sequence determination of ∼40-kb of the camphor (CAM) degradation plasmid revealed the locations of two isofunctional 2,5-DKCMO genes (camE25-1 for 2,5-DKCMO-1, and camE25-2 for 2,5-DKCMO-2) as well as that of 3,6-DKCMO-encoding gene (camE36). In addition, by pulsed-field gel electrophoresis, the CAM plasmid was established to be linear and ∼533-kb in length. To enable functional assessment of the two-component monooxygenase system in Baeyer-Villiger oxidations, recombinant plasmids expressing Fred in tandem with the respective 2,5-DKCMO and 3,6-DKCMO encoding genes in Escherichia coli were constructed. Comparative substrate profiling of the isofunctional 2,5-DCKMOs did not yield obvious differences in Baeyer-Villiger biooxidations but they are distinct from 3,6-DKCMO in the stereoselective oxygenations with various mono- and bicyclic ketone substrates

Multidimensional endotyping in patients with severe asthma reveals inflammatory heterogeneity in matrix metalloproteinases and chitinase 3–like protein 1

BackgroundDisease heterogeneity in patients with severe asthma and its relationship to inflammatory mechanisms remain poorly understood.ObjectiveWe aimed to identify and replicate clinicopathologic endotypes based on analysis of blood and sputum parameters in asthmatic patients.MethodsOne hundred ninety-four asthmatic patients and 21 control subjects recruited from 2 separate centers underwent detailed clinical assessment, sputum induction, and phlebotomy. One hundred three clinical, physiologic, and inflammatory parameters were analyzed by using topological data analysis and Bayesian network analysis.ResultsSevere asthma was associated with anxiety and depression, obesity, sinonasal symptoms, decreased quality of life, and inflammatory changes, including increased sputum chitinase 3–like protein 1 (YKL-40) and matrix metalloproteinase (MMP) 1, 3, 8, and 12 levels. Topological data analysis identified 6 clinicopathobiologic clusters replicated in both geographic cohorts: young, mild paucigranulocytic; older, sinonasal disease; obese, high MMP levels; steroid resistant TH2 mediated, eosinophilic; mixed granulocytic with severe obstruction; and neutrophilic, low periostin levels, severe obstruction. Sputum IL-5 levels were increased in patients with severe particularly eosinophilic forms, whereas IL-13 was suppressed and IL-17 levels did not differ between clusters. Bayesian network analysis separated clinical features from intricately connected inflammatory pathways. YKL-40 levels strongly correlated with neutrophilic asthma and levels of myeloperoxidase, IL-8, IL-6, and IL-6 soluble receptor. MMP1, MMP3, MMP8, and MMP12 levels were associated with severe asthma and were correlated positively with sputum IL-5 levels but negatively with IL-13 levels.ConclusionIn 2 distinct cohorts we have identified and replicated 6 clinicopathobiologic clusters based on blood and induced sputum measures. Our data underline a disconnect between clinical features and underlying inflammation, suggest IL-5 production is relatively steroid insensitive, and highlight the expression of YKL-40 in patients with neutrophilic inflammation and the expression of MMPs in patients with severe asthma

Association of risk of suicide attempts with methylphenidate treatment

IMPORTANCE Patients with attention-deficit/hyperactivity disorder (ADHD) are at an increased risk of attempting suicide. Stimulants, such as methylphenidate hydrochloride, are the most common treatment for ADHD, but the association between their therapeutic use and suicide is unclear. OBJECTIVE To investigate the association between methylphenidate and the risk of suicide attempts. DESIGN, SETTING, AND PARTICIPANTS A population-based, electronic medical records database from the Hong Kong Clinical Data Analysis & Reporting System was used to identify 25 629 individuals aged 6 to 25 years who were treated with methylphenidate between January 1, 2001, and December 31, 2015. Those who had attempted suicide were included in the analysis. A self-controlled case series design was used to control for time-invariant characteristics of the patients. MAIN OUTCOMES AND MEASURES Relative incidence of suicide attempt during periods when patients were exposed to methylphenidate compared with nonexposed periods. RESULTS Among 25 629 patients with methylphenidate prescriptions, 154 had their first recorded suicide attempt within the study period; of these individuals, 111 (72.1%) were male; mean (SD) age at baseline was 7.15 (2.19) years. The overall incidence of suicide attempts duringmethylphenidate treatment was 9.27 per 10 000 patient-years. An increased risk of suicide attempts was detected during the 90-day period before methylphenidate was initiated, with an incidence rate ratio (IRR) of 6.55 (95%CI, 3.37-12.72). The IRR remained elevated during the first 90 days of treatment (IRR, 3.91; 95%CI, 1.62-9.42) before returning to baseline levels during ongoing treatment (IRR, 1.35; 95%CI, 0.77-2.38). When the risk during the first 90 days of treatment was compared with the 90 days preceding first treatment, the incidence of suicide attempts was not elevated (IRR, 0.78; 95%CI, 0.26-2.35). CONCLUSIONS AND RELEVANCE The incidence of suicide attempts was higher in the period immediately before the start ofmethylphenidate treatment. The risk remained elevated immediately after the start ofmethylphenidate treatment and returned to baseline levels during continuation of methylphenidate treatment. The observed higher risk of suicide attempts before treatment may reflect emerging psychiatric symptoms that trigger medical consultations that result in a decision to begin ADHD treatment. Therefore, this study’s results do not support a causal association between methylphenidate treatment and suicide attempts

Epithelial IL-6 trans-signaling defines a new asthma phenotype with increased airway inflammation

Background: Although several studies link high levels of IL-6 and soluble IL-6 receptor (sIL-6R) to asthma severity and decreased lung function, the role of IL-6 trans-signaling (IL-6TS) in asthmatic patients is unclear. Objective: We sought to explore the association between epithelial IL-6TS pathway activation and molecular and clinical phenotypes in asthmatic patients. Methods: An IL-6TS gene signature obtained from air-liquid interface cultures of human bronchial epithelial cells stimulated with IL-6 and sIL-6R was used to stratify lung epithelial transcriptomic data (Unbiased Biomarkers in Prediction of Respiratory Disease Outcomes [U-BIOPRED] cohorts) by means of hierarchical clustering. IL-6TS-specific protein markers were used to stratify sputum biomarker data (Wessex cohort). Molecular phenotyping was based on transcriptional profiling of epithelial brushings, pathway analysis, and immunohistochemical analysis of bronchial biopsy specimens. Results: Activation of IL-6TS in air-liquid interface cultures reduced epithelial integrity and induced a specific gene signature enriched in genes associated with airway remodeling. The IL-6TS signature identified a subset of patients with IL-6TS-high asthma with increased epithelial expression of IL-6TS-inducible genes in the absence of systemic inflammation. The IL-6TS-high subset had an overrepresentation of frequent exacerbators, blood eosinophilia, and submucosal infiltration of T cells and macrophages. In bronchial brushings Toll-like receptor pathway genes were upregulated, whereas expression of cell junction genes was reduced. Sputum sIL-6R and IL-6 levels correlated with sputum markers of remodeling and innate immune activation, in particular YKL-40, matrix metalloproteinase 3, macrophage inflammatory protein 1 beta, IL-8, and IL-1 beta. Conclusions: Local lung epithelial IL-6TS activation in the absence of type 2 airway inflammation defines a novel subset of asthmatic patients and might drive airway inflammation and epithelial dysfunction in these patients.Peer reviewe

Epithelial IL-6 trans-signaling defines a new asthma phenotype with increased airway inflammation

Background: Although several studies link high levels of IL-6 and soluble IL-6 receptor (sIL-6R) with asthma severity and decreased lung function, the role of IL-6 trans-signaling (IL-6TS) in asthma is unclear. Objective: To explore the association between epithelial IL-6TS pathway activation and molecular and clinical phenotypes in asthma. Methods: An IL-6TS gene signature, obtained from air-liquid interface (ALI) cultures of human bronchial epithelial cells stimulated with IL-6 and sIL-6R, was used to stratify lung epithelium transcriptomic data (U-BIOPRED cohorts) by hierarchical clustering. IL-6TS-specific protein markers were used to stratify sputum biomarker data (Wessex cohort). Molecular phenotyping was based on transcriptional profiling of epithelial brushings, pathway analysis and immunohistochemical analysis of bronchial biopsies. Results: Activation of IL-6TS in ALI cultures reduced epithelial integrity and induced a specific gene signature enriched in genes associated with airway remodeling. The IL-6TS signature identified a subset of IL-6TS. High asthma patients with increased epithelial expression of IL-6TS inducible genes in absence of systemic inflammation. The IL-6TS High subset had an overrepresentation of frequent exacerbators, blood eosinophilia, and submucosal infiltration of T cells and macrophages. In bronchial brushings, TLR pathway genes were up-regulated while the expression of tight junction genes was reduced. Sputum sIL-6R and IL-6 levels correlated with sputum markers of remodeling and innate immune activation, in particular YKL-40, MMP3, MIP-1β, IL-8 and IL-1β. Conclusions: Local lung epithelial IL-6TS activation in absence of type 2 airway inflammation defines a novel subset of asthmatics and may drive airway inflammation and epithelial dysfunction in these patients

Monitoring of Baeyer-Villiger biotransformation kinetics and fingerprinting using reactIR 4000 spectroscopy

NRC publication: Ye

Direct fermentation of triticale starch to lactic acid by Rhizopus oryzae

The production of lactic acid from triticale starch was demonstratedfor the first time. Direct fermentation by Rhizopus oryzae in batch modeled to a conversion yield of 0.74 g lactic acid/g starch. Kinetics analysisof the fermentation suggested a multiphase process which consistedof quick hydrolysis of starch, rapid glucose accumulation followedby fast production of lactic acid. In batch fermentation mode, correcttiming and proper dosage of a neutralizing agent (calcium carbonate)were found to be critical factors that affect the organic acid yield.Addition of CaCO3 at the time point when glucose accumulationhad reached its peak (24 h) resulted in the highest lactic acid yield.The best ratio (by weight) of triticale starch to CaCO3 for lactic acidproduction was 1:1. It was important to maintain a pH of about 5during the whole fermentation process. Fermentation carried out inan optimized, commercially available small-scale parallel bioreactor(DASGIP) yielded up to 0.87 g lactic acid/g triticale starch whenR. oryzae spores were directly added to the fermentation medium. Ourfermentation results indicated that triticale starch from this nonfoodcrop is a promising renewable feedstock for production of lactic acidPeer reviewed: YesNRC publication: Ye

Two-layer continuous-process design for the biodesulfurization of diesel oils under bacterial growth conditions

An aqueous\ufffdorganic two-layer partitioning and continuous process was designed to make efficient use of growing cells for the biodesulfurization of diesel oils. This biphasic system was shown to have a significant advantage over batch and fed-batch processes in the maintenance of the biodesulfurization activity for an extended period of time. Specifically, it was demonstrated that in a 2-L bioreactor 1500 ppm total sulfur was removed from 250 ml diesel oil within a 5-day period.NRC publication: Ye