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The UK Synthetic Biology Research Centre, SYNBIOCHEM, hosted by the Manchester Institute of Biotechnology at the University of Manchester is delivering innovative technology platforms to facilitate the predictable engineering of microbial bio-factories for fine and speciality chemicals production. We provide an overview of our foundry activities that are being applied to grand challenge projects to deliver innovation in bio-based chemicals production for industrial biotechnology.

The fate of 2,4-dinitrochlorobenzene, a potent contact sensitizing chemical, and 2,4-dichloronitrobenzene, a non-sensitizer, was compared following their application to the skin of BALB/c mice. Although both chemicals were able to bind to protein in vitro and were capable of being absorbed across mouse skin in vivo, only 2,4-dinitrochlorobenzene was able to bind to cells in the skin and to induce the movement of these cells from the epidermis into the dermis and ultimately into the draining lymph nodes. The sensitization potential of a chemical may therefore be dependent on its ability to associate with and stimulate the efflux of cutaneous antigen-presenting cells. [ABSTRACT FROM AUTHOR]

The article describes the initial findings of a comparative study of with sex chromosome abnormalities and a group of matched controls of normal chromosome complement. Electroencephalogram (EEG) recordings were performed on 22 patients with sex chromosome abnormalities. The study examined the control group of patients' normal karyotype from the same hospital, matched for age, intelligence quotient (IQ) and duration of stay. It found that the subjects with sex chromosome abnormalities had a higher prevalence of EEG disturbance. The prevalence of EEG change was as common in the XYY as in the XXY syndrome.

Objective: To examine mothers' internet usage, in conjunction with social, health care, and virtual peer support navigations, when congenital anomalies were diagnosed in utero., Design: Qualitative descriptive, consisting of semistructured interviews., Setting: Interview data were collected over Zoom; mothers participated from locations of their choosing., Participants: Mothers of neonates discharged postoperatively from NICUs for uterine-diagnosed congenital anomalies. The sample was purposefully recruited from private Facebook groups for parents of children with congenital anomalies., Intervention/measurements: Analysis was done with deductive coding using concepts from the third iteration of the systems engineering initiative for patient safety theory. The a priori codes were health care, social, journey-benefit, journey-risk, task, and technology., Results: Twenty-two mothers signed up for an interview; 12 completed an interview, and 10 did not. The majority (n = 8, 66%) were White, had a bachelor's or graduate degree (n = 7, 58%) and were between 24 and 33 years of age (n = 8, 66%). Nine themes emerged: (a) Providers cautioned searching for diagnosis information but encouraged private Facebook groups for peer support, (b) Mothers' inquiries for their own care are lacking, (c) Search for information while recognizing parent-partner's coping differences, (d) Pace information from friends and family with patience and appreciation, (e) Manage inquiries from friends and family with group sharing, (f) Private Facebook groups provide a means of receiving and giving peer support, (g) Exposure to difficult stories on Facebook is a risk of stress, (h) Select a NICU, learn about their children's diagnoses, participate in virtual peer support, and (i) Device features frame search strategies., Conclusion: Mothers reflected on the internet as a burden and a source of support in their health care journeys. The ubiquity of internet access calls for mothers to include in their health care journeys the complexities of managing time spent on the internet., (Published by Elsevier Inc.)

Introduction: Interstitial lung disease encompasses a group of rare lung conditions causing inflammation and scarring of lung tissue. The typical method of monitoring disease activity is through pulmonary function tests performed in a hospital setting. However, accessing care can be difficult for rural patients due to numerous barriers. This study assesses the feasibility and acceptability of home spirometry telemonitoring using MIR-Spirometers and the patientMpower home-monitoring platform for rural patients with interstitial lung disease., Methods: Unblinded, uncontrolled, prospective, multiple-methods study of the feasibility and utility of remote monitoring of 20 rural subjects with interstitial lung disease. Study assessments include adherence to twice weekly spirometry for 3 months in addition to mMRC dyspnea and EQ-5D-5L health-related quality of life questionnaires with each spirometry maneuver. Upon completion, subjects were encouraged to complete an 11-question satisfaction survey and participate in semi-structured qualitative interviews to further explore expectations and perceptions of rural patients to telehealth and remote patient monitoring., Results: 19 subjects completed the 3-month study period. Adherence to twice weekly spirometry was mean 53% ± 38%, with participants on average performing 2.26 ± 1.69 maneuvers per week. The median (Range) number of maneuvers per week was 2.0 (0.0, 7.0). The majority of participants responded favorably to the patient satisfaction survey questions. Themes regarding barriers to access included: lack of local specialty care, distance to center with expertise, and time, distance, and high cost associated with travel. Remote monitoring was well perceived amongst subjects as a way to improve access and overcome barriers., Conclusions: Remote spirometry monitoring through web-based telehealth is acceptable and feasible for rural patients. Perceived benefits include overcoming access barriers like time, distance, and travel costs. However, cost, reimbursement, and internet access must be addressed before implementing it widely. Future studies are needed to ensure long-term feasibility and to compare outcomes with usual care., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Characterising the small intestine absorptive membrane is essential to enable prediction of the systemic exposure of oral formulations. In particular, the ontogeny of key intestinal Drug Metabolising Enzymes and Transporter (DMET) proteins involved in drug disposition needs to be elucidated to allow for accurate prediction of the PK profile of drugs in the paediatric cohort. Using pinch biopsies from the paediatric duodenum (n = 36; aged 11 months to 15 years), the abundance of 21 DMET proteins and two enterocyte markers were quantified via LC-MS/MS. An established LCMS nanoflow method was translated to enable analysis on a microflow LC system, and a new stable-isotope-labelled QconCAT standard developed to enable quantification of these proteins. Villin-1 was used to standardise abundancy values. The observed abundancies and ontogeny profiles, agreed with adult LC-MS/MS-based data, and historic paediatric data obtained via western blotting. A linear trend with age was observed for duodenal CYP3A4 and CES2 only. As this work quantified peptides on a pinch biopsy coupled with a microflow method, future studies using a wider population range are very feasible. Furthermore, this DMET ontogeny data can be used to inform paediatric PBPK modelling and to enhance the understanding of oral drug absorption and gut bioavailability in paediatric populations., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Patients suspected of ventilator-associated lower respiratory tract infections (VA-LRTIs) commonly receive broad-spectrum antimicrobial therapy unnecessarily. We tested whether exhaled breath analysis can discriminate between patients suspected of VA-LRTI with confirmed infection, from patients with negative cultures. Breath from 108 patients suspected of VA-LRTI was analysed by gas chromatography-mass spectrometry. The breath test had a sensitivity of 98% at a specificity of 49%, confirmed with a second analytical method. The breath test had a negative predictive value of 96% and excluded pneumonia in half of the patients with negative cultures. Trial registration number: UKCRN ID number 19086, registered May 2015., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY. Published by BMJ.)

Inflammation is strongly implicated in both injury and repair processes occurring after stroke. In this exploratory study we assessed the feasibility of repeated sampling of exhaled volatile organic compounds and performed an untargeted metabolomic analysis of plasma collected at multiple time periods after stroke. Metabolic profiles were compared with the time course of the inflammatory markers C-reactive protein (CRP) and interleukin-6 (IL-6). Serial breath sampling was well-tolerated by all patients and the measurement appears feasible in this group. We found that exhaled decanal tracks CRP and IL-6 levels post-stroke and correlates with several metabolic pathways associated with a post-stroke inflammatory response. This suggests that measurement of breath and blood metabolites could facilitate development of novel therapeutic and diagnostic strategies. Results are discussed in relation to the utility of breath analysis in stroke care, such as in monitoring recovery and complications including stroke associated infection., (© 2021. The Author(s).)

Introduction: Obstructive sleep apnea (OSA) is an independent cerebrovascular risk factor and highly prevalent in patients with ischemic stroke and transient ischemic attack (TIA). Timely diagnosis and treatment of OSA is important as clinical data suggest that treatment of OSA in the setting of acute ischemic stroke improves functional outcomes. We aimed to assess polysomnography (PSG) utilization in US. Veterans with acute stroke or TIA over a 2-year period., Methods: Veterans with acute ischemic stroke or TIA presenting to a Veterans Administration Medical Center (VAMC) between October 1, 2015, and June 30, 2017, were included. Demographic, clinical data, and PSG within 12 months of hospital discharge were obtained from the VA Corporate Data Warehouse to determine the rate of PSG testing among those with acute ischemic stroke or TIA. Fisher's exact test and two-sample t tests were used to compare demographic and clinical characteristics for those receiving and not receiving PSG. Mixed effect logistic regression was used to model the association of clinical and demographic characteristics with PSG receipt., Results: In fiscal years (FYs) 2016 and 2017, 9,200 Veterans were admitted to a VAMC with ischemic stroke (6,011) or TIA (3,089). Veterans were elderly (70.5 ± 11.1 years), predominantly male (95.7%), and largely Caucasian (68.0% Caucasian, 26.3% African-American). Just 6.0% of Veterans underwent PSG within 1 year of acute ischemic stroke or TIA in FY 2016, compared to 6.2% in FY 2017 (p = 0.72). Compared to Veterans ≥80 years, those <60 had adjusted OR of 6.73 (4.10-11.05), those 60-69 had OR 4.29 (2.73-6.74), and those 70-79 had OR 2.63 (1.66-4.18) of having PSG. Veterans with diabetes or heart failure had significantly higher odds, whereas those with dementia had significantly lower odds of receiving PSG., Conclusion: PSG utilization among US Veterans is low and stable over time, despite recent guidelines recommending PSG among those having stroke or TIA. Older Veterans and those with dementia were unlikely to get PSG, representing especially vulnerable populations., (© 2019 S. Karger AG, Basel.)

Methods for breath sampling and analysis require robust quality assessment to minimise the risk of false discoveries. Planning large-scale multi-site breath metabolite profiling studies also requires careful consideration of systematic and random variation as a result of sampling and analysis techniques. In this study we use breath sample data from the recent U-BIOPRED cohort to evaluate and discuss some important methodological considerations such as batch variation and correction, variation between sites, storage and transportation, as well as inter-instrument analytical differences. Based on this we provide a summary of recommended best practices for new large scale multi-site studies.

Reduction of double bonds of α,β-unsaturated carboxylic acids and esters by ene-reductases remains challenging and it typically requires activation by a second electron-withdrawing moiety, such as a halide or second carboxylate group. We showed that profen precursors, 2-arylpropenoic acids and their esters, were efficiently reduced by Old Yellow Enzymes (OYEs). The XenA and GYE enzymes showed activity towards acids, while a wider range of enzymes were active towards the equivalent methyl esters. Comparative co-crystal structural analysis of profen-bound OYEs highlighted key interactions important in determining substrate binding in a catalytically active conformation. The general utility of ene reductases for the synthesis of (R)-profens was established and this work will now drive future mutagenesis studies to screen for the production of pharmaceutically-active (S)-profens.

Although cold acclimation is a key process in plants from temperate climates, the mechanisms sensing low temperature remain obscure. Here, we show that the accumulation of the organic acid fumaric acid, mediated by the cytosolic fumarase FUM2, is essential for cold acclimation of metabolism in the cold-tolerant model species Arabidopsis (Arabidopsis thaliana). A nontargeted metabolomic approach, using gas chromatography-mass spectrometry, identifies fumarate as a key component of the cold response in this species. Plants of T-DNA insertion mutants, lacking FUM2, show marked differences in their response to cold, with contrasting responses both in terms of metabolite concentrations and gene expression. The fum2 plants accumulated higher concentrations of phosphorylated sugar intermediates and of starch and malate. Transcripts for proteins involved in photosynthesis were markedly down-regulated in fum2.2 but not in wild-type Columbia-0. Plants of fum2 show a complete loss of the ability to acclimate photosynthesis to low temperature. We conclude that fumarate accumulation plays an essential role in low temperature sensing in Arabidopsis, either indirectly modulating metabolic or redox signals or possibly being itself directly involved in cold sensing., (© 2016 American Society of Plant Biologists. All rights reserved.)

myo-Inositol phosphates possessing the 1,2,3-trisphosphate motif share the remarkable ability to completely inhibit iron-catalysed hydroxyl radical formation. The simplest derivative, myo-inositol 1,2,3-trisphosphate [Ins(1,2,3)P(3)], has been proposed as an intracellular iron chelator involved in iron transport. The binding conformation of Ins(1,2,3)P(3) is considered to be important to complex Fe(3+) in a 'safe' manner. Here, a pyrene-based fluorescent probe, 4,6-bispyrenoyl-myo-inositol 1,2,3,5-tetrakisphosphate [4,6-bispyrenoyl Ins(1,2,3,5)P(4)], has been synthesised and used to monitor the conformation of the 1,2,3-trisphosphate motif using excimer fluorescence emission. Ring-flip of the cyclohexane chair to the penta-axial conformation occurs upon association with Fe(3+), evident from excimer fluorescence induced by pi-pi stacking of the pyrene reporter groups, accompanied by excimer formation by excitation at 351 nm. This effect is unique amongst biologically relevant metal cations, except for Ca(2+) cations exceeding a 1 : 1 molar ratio. In addition, the thermodynamic constants for the interaction of the fluorescent probe with Fe(3+) have been determined. The complexes formed between Fe(3+) and 4,6-bispyrenoyl Ins(1,2,3,5)P(4) display similar stability to those formed with Ins(1,2,3)P(3), indicating that the fluorescent probe acts as a good model for the 1,2,3-trisphosphate motif. This is further supported by the antioxidant properties of 4,6-bispyrenoyl Ins(1,2,3,5)P(4), which closely resemble those obtained for Ins(1,2,3)P(3). The data presented confirms that Fe(3+) binds tightly to the unstable penta-axial conformation of myo-inositol phosphates possessing the 1,2,3-trisphosphate motif.

Natural myo-inositol phosphate antioxidants containing the 1,2,3-trisphosphate motif bind Fe(3+) in the unstable penta-axial conformation.

This study evaluated a single intradermal injection model in the guinea pig with subsequent inhalation challenge and serological analysis as a method to predict the potential of chemicals to induce respiratory allergy. Four known respiratory allergens (trimellitic anhydride, diphenyl methane diisocyanate, phthalic anhydride and toluene diisocyanate (TDI)) were screened by two industrial research laboratories using this protocol. Dinitrochlorobenzene, a potent contact allergen, was included as a negative control material. In both laboratories, the respiratory allergens, but not the contact allergen, induced high titre antigen-specific antibodies in treated animals. The inhalation challenge results were similar in both laboratories but were less conclusive in that exposure to free TDI failed to induce pulmonary responses, probably because it fails to penetrate to the deep lung in sufficient concentration. Although the assay shows promise as a means of identifying chemical respiratory sensitisers, its use as a routine screen for the prediction of the ability of materials to induce respiratory allergy in man is probably questionable.

Epicutaneous application of 2,4-dinitrothiocyanobenzene (DNTB) is said to result in specific immunological hyporesponsiveness and fails to induce contact sensitization. However, we demonstrate that topical exposure to DNTB causes activation of the draining lymph node in mice and the induction of contact sensitization in both rodents and a single human volunteer. In mice and rats, pre-exposure to DNTB failed to impair subsequent responsiveness to the cross-reactive allergen 2,4-dinitrochlorobenzene. These data provide evidence that DNTB cannot be regarded as an exclusive tolerogen when applied epicutaneously and indicate that attempts to define the characteristics of tolerising chemicals from analysis of this agent may be misleading.

Guinea-pigs injected intradermally with the known respiratory sensitiser trimellitic anhydride (TMA) developed high-titre antigen-specific homocytotropic (IgG1 and IgE) antibodies. Many of the sensitised animals responded to a challenge by inhalation with either free TMA or a TMA-protein conjugate with a change in respiratory rate, reflecting the onset of bronchoconstriction. Guinea-pigs were also injected intradermally with 2,4-dinitrochlorobenzene (DNCB), which is a potent skin sensitiser in man but which has not been reported to cause respiratory allergy. These animals developed only low-titre homocytotropic antibodies and were unresponsive to an inhalation challenge with either free or conjugated hapten. The animals were, however, contact-sensitised to the chemical.

Guinea pigs could be immunologically sensitised (as shown by the development of antigen-specific homocytotropic antibodies) to toluene diisocyanate by exposing them for 3 h a day for 5 consecutive days to atmospheres containing free chemical. Pulmonary reactions could be elicited in many of the sensitised animals by challenging them with atmospheres containing protein conjugates of the chemical and then measuring changes in respiratory rate. Successful elicitation of pulmonary reactions appeared to depend upon a number of factors, including the quality of the protein conjugate used for the challenge, but possibly also the development of IgE as well as IgG1 antibodies. Antigen-specific homocytotropic antibodies were detected in guinea pigs similarly exposed by inhalation to two non-isocyanate respiratory allergens, trimellitic anhydride and a reactive dye. Although the animals were immunologically sensitised to the chemicals, challenge with atmospheres containing appropriate chemical-protein conjugates failed to stimulate changes in respiratory rate.