Integrin α5β1 Function Is Regulated by XGIPC/kermit2 Mediated Endocytosis during Xenopus laevis Gastrulation

During Xenopus gastrulation α5β1 integrin function is modulated in a temporally and spatially restricted manner, however, the regulatory mechanisms behind this regulation remain uncharacterized. Here we report that XGIPC/kermit2 binds to the cytoplasmic domain of the α5 subunit and regulates the activity of α5β1 integrin. The interaction of kermit2 with α5β1 is essential for fibronectin (FN) matrix assembly during the early stages of gastrulation. We further demonstrate that kermit2 regulates α5β1 integrin endocytosis downstream of activin signaling. Inhibition of kermit2 function impairs cell migration but not adhesion to FN substrates indicating that integrin recycling is essential for mesoderm cell migration. Furthermore, we find that the α5β1 integrin is colocalized with kermit2 and Rab 21 in embryonic and XTC cells. These data support a model where region specific mesoderm induction acts through kermit2 to regulate the temporally and spatially restricted changes in adhesive properties of the α5β1 integrin through receptor endocytosis

MiDAS 4: A global catalogue of full-length 16S rRNA gene sequences and taxonomy for studies of bacterial communities in wastewater treatment plants

Microbial communities are responsible for biological wastewater treatment, but our knowledge of their diversity and function is still poor. Here, we sequence more than 5 million high-quality, full-length 16S rRNA gene sequences from 740 wastewater treatment plants (WWTPs) across the world and use the sequences to construct the ‘MiDAS 4’ database. MiDAS 4 is an amplicon sequence variant resolved, full-length 16S rRNA gene reference database with a comprehensive taxonomy from domain to species level for all sequences. We use an independent dataset (269 WWTPs) to show that MiDAS 4, compared to commonly used universal reference databases, provides a better coverage for WWTP bacteria and an improved rate of genus and species level classification. Taking advantage of MiDAS 4, we carry out an amplicon-based, global-scale microbial community profiling of activated sludge plants using two common sets of primers targeting regions of the 16S rRNA gene, revealing how environmental conditions and biogeography shape the activated sludge microbiota. We also identify core and conditionally rare or abundant taxa, encompassing 966 genera and 1530 species that represent approximately 80% and 50% of the accumulated read abundance, respectively. Finally, we show that for well-studied functional guilds, such as nitrifiers or polyphosphate-accumulating organisms, the same genera are prevalent worldwide, with only a few abundant species in each genus

Direct RT-qPCR Assay for the Detection of SARS-CoV-2 in Saliva Samples

Since mid-2020 there have been complexities and difficulties in the standardisation and administration of nasopharyngeal swabs. Coupled with the variable and/or poor accuracy of lateral flow devices, this has led to increased societal ‘testing fatigue’ and reduced confidence in test results. Consequently, asymptomatic individuals have developed reluctance towards repeat testing, which remains the best way to monitor COVID-19 cases in the wider population. On the other hand, saliva-based PCR, a non-invasive, highly sensitive, and accurate test suitable for everyone, is gaining momentum as a straightforward and reliable means of detecting SARS-CoV-2 in symptomatic and asymptomatic individuals. Here, we provide an itemised list of the equipment and reagents involved in the process of sample submission, inactivation and analysis, as well as a detailed description of how each of these steps is performed

Performance evaluation of a non-invasive one-step multiplex RT-qPCR assay for detection of SARS-CoV-2 direct from saliva

Polymerase chain reaction (PCR) has proven to be the gold-standard for SARS-CoV-2 detection in clinical settings. The most common approaches rely on nasopharyngeal specimens obtained from swabs, followed by RNA extraction, reverse transcription and quantitative PCR. Although swab-based PCR is sensitive, swabbing is invasive and unpleasant to administer, reducing patient compliance for regular testing and resulting in an increased risk of improper sampling. To overcome these obstacles, we developed a non-invasive one-step RT-qPCR assay performed directly on saliva specimens. The University of Nottingham Asymptomatic Testing Service protocol simplifies sample collection and bypasses the need for RNA extraction, or additives, thus helping to encourage more regular testing and reducing processing time and costs. We have evaluated the assay against the performance criteria specified by the UK regulatory bodies and attained accreditation (BS EN ISO/IEC 17,025:2017) for SARS-CoV-2 diagnostic testing by the United Kingdom Accreditation Service. We observed a sensitivity of 1 viral copy per microlitre of saliva, and demonstrated a concordance of > 99.4% between our results and those of other accredited testing facilities. We concluded that saliva is a stable medium that allows for a highly precise, repeatable, and robust testing method

Influence of socioeconomic factors on pregnancy outcome in women with structural heart disease

OBJECTIVE: Cardiac disease is the leading cause of indirect maternal mortality. The aim of this study was to analyse to what extent socioeconomic factors influence the outcome of pregnancy in women with heart disease.  METHODS: The Registry of Pregnancy and Cardiac disease is a global prospective registry. For this analysis, countries that enrolled ≥10 patients were included. A combined cardiac endpoint included maternal cardiac death, arrhythmia requiring treatment, heart failure, thromboembolic event, aortic dissection, endocarditis, acute coronary syndrome, hospitalisation for cardiac reason or intervention. Associations between patient characteristics, country characteristics (income inequality expressed as Gini coefficient, health expenditure, schooling, gross domestic product, birth rate and hospital beds) and cardiac endpoints were checked in a three-level model (patient-centre-country).  RESULTS: A total of 30 countries enrolled 2924 patients from 89 centres. At least one endpoint occurred in 645 women (22.1%). Maternal age, New York Heart Association classification and modified WHO risk classification were associated with the combined endpoint and explained 37% of variance in outcome. Gini coefficient and country-specific birth rate explained an additional 4%. There were large differences between the individual countries, but the need for multilevel modelling to account for these differences disappeared after adjustment for patient characteristics, Gini and country-specific birth rate.  CONCLUSION: While there are definite interregional differences in pregnancy outcome in women with cardiac disease, these differences seem to be mainly driven by individual patient characteristics. Adjustment for country characteristics refined the results to a limited extent, but maternal condition seems to be the main determinant of outcome

Defining murine organogenesis at single-cell resolution reveals a role for the leukotriene pathway in regulating blood progenitor formation.

During gastrulation, cell types from all three germ layers are specified and the basic body plan is established 1 . However, molecular analysis of this key developmental stage has been hampered by limited cell numbers and a paucity of markers. Single-cell RNA sequencing circumvents these problems, but has so far been limited to specific organ systems 2 . Here, we report single-cell transcriptomic characterization of >20,000 cells immediately following gastrulation at E8.25 of mouse development. We identify 20 major cell types, which frequently contain substructure, including three distinct signatures in early foregut cells. Pseudo-space ordering of somitic progenitor cells identifies dynamic waves of transcription and candidate regulators, which are validated by molecular characterization of spatially resolved regions of the embryo. Within the endothelial population, cells that transition from haemogenic endothelial to erythro-myeloid progenitors specifically express Alox5 and its co-factor Alox5ap, which control leukotriene production. Functional assays using mouse embryonic stem cells demonstrate that leukotrienes promote haematopoietic progenitor cell generation. Thus, this comprehensive single-cell map can be exploited to reveal previously unrecognized pathways that contribute to tissue development

Full-scale emission results (Ninf2/infO and CHinf4/inf)

Chapter six of the Open Access book, 'Quantification and Modelling of Fugitive Greenhouse Gas Emissions from Urban Water Systems', published by IWA Publishing, is available online at https://iwaponline.com/ebooks/book/844/Quantification-and-Modelling-of-Fugitive .Copyright © 2022 The Authors and Editors. This chapter reviews the studies from N2 O and CH4 monitoring campaigns in full-scale wastewater treatment plants (WWTPs) and sewer networks. The focus is on greenhouse gas (GHG) emissions from WWTPs as more literature is available. The analysis classifies quantified N2 O and CH4 emission factors (EFs), triggering operational conditions and formation pathways for different configurations. Control strategies to minimize N2 O emissions are proposed for different process groups. The main reasons for EF discrepancies are discussed. Overall, N2 O emission factors for processes treating lowstrength wastewater streams range between 0.003 and 5.6% of the N-load (average equal to 0.9% of the N-load). Emissions higher than mainstream process average emissions have been reported in sequencing batch reactors (average equal to 3.6% of the influent N-load) and step-fed plug flow reactors. In full-scale sidestream processes, less than 15 monitoring campaigns have reported EFs (average equal to 2.5% of the N-load). Differences in the EFs among the process groups are partially attributed to disparities in the control strategies (i.e. aeration control), configuration, and operational and environmental conditions that favour the preferred enzymatic pathways. Overall, triggering operational conditions for elevated N2 O emissions in full-scale wastewater treatment processes include (i) increased NH4+ concentrations leading to a high ammonia oxidation rate (AOR) and increased production of intermediates (e.g. NH2 OH, NO−, etc.), (ii) improper aeration control (i.e. inadequate aeration and non-aeration duration, over-aeration, under-aeration), (iii) NO2− accumulation triggering the nitrifier denitrification pathway, and (iv) sudden shifts in incomplete heterotrophic denitrification (i.e. due to excess dissolved oxygen (DO), chemical oxygen demand (COD) limitation etc.). The N2 O monitoring strategies can also influence the reliability of the quantified EFs. Due to temporal variation of N2 O emissions, short-term studies are not sufficient to quantify annual EFs. The analysis showed that the average EF for processes treating low-strength streams monitored for less than a week is 0.66% of the influent N-load. On the other hand, processes monitored over 6 months have an average EF equal to 1.74%. Compared with N2 O, CH4 quantification from full-scale WWTPs is less investigated, while it also contributes significantly to the overall plant carbon footprint. The results of full-scale CH4 quantification studies are summarized in this chapter. Emissions of CH4 in WWTPs mainly originate from the influent, anaerobic wastewater treatment and anaerobic sludge handling processes. The amount of CH4 emissions varies greatly with different configurations of WWTPs. For WWTPs without anaerobic sludge handling processes, the CH4 emissions can mainly be traced back to the CH4 dissolved in the influent. When anaerobic treatment is applied in WWTPs for wastewater COD removal, its CH4 emissions might substantially increase the overall plant carbon footprint. GHG monitoring campaigns carried out in WWTPs should include the monitoring of fugitive CH4 emissions. Finally, CH4 and N2 O emissions reported from sewer networks are also summarized in this chapter. The last part of the chapter summarizes some mitigation strategies applied at full-scale to control fugitive CHG emissions from WWTPs and sewers.Maite Pijuan acknowledges the support from the Economy and Knowledge Department of the Catalan Government through a Consolidated Research Group (ICRA-TECH – 2017 SGR 1318) – Catalan Institute for Water Research and the Spanish Government through the Salvador de Madariaga mobility program (PRX19/00051). Vasileia Vasilaki and Evina Katsou would like to acknowledge the Horizon 2020 research and innovation program, SMART-Plant under grant agreement No 690323. Haoran Duan acknowledges the support of the Australian Research Council (ARC) through project DP180103369

Rapid stimulation of Ins (1,4,5)P3 production in rat aorta by NE: correlation with contractile state

Rapid stimulation of Ins(1,4,5)P3 production in rat aorta by NE: correlation with contractile state. Am. J. Physiol. 264 (Heart Circ. Physiol. 33): H126-H132, 1993.--The isomeric composition of inositol phosphates generated in response to norepinephrine (NE) stimulation and the relationship of inositol phosphate production to release of intracellular Ca2+ as measured by contraction were characterized in rat aorta prelabeled with [3H]inositol. NE stimulated a rapid and transient increase in labeled D-myo-inositol 1,4,5-trisphosphate [Ins-(1,4,5)P3] levels. A maximal increase in labeled Ins(1,4,5)P3 occurred within 15 s of stimulation followed by a decline to control levels at 5 min. D-Myo-inositol 1,3,4-trisphosphate [Ins-(1,3,4)P3] and D-myo-inositol 1-monophosphate [Ins(1)P] levels also increased rapidly in response to NE. In contrast to the transient production of Ins(1,4,5)P3, Ins(1,3,4)P3 and Ins(1)P production was maintained in the presence of NE. Half-maximal stimulation of Ins(1,4,5)P3 production and Ca2+ release occurred at 0.3 microM NE, and maximal effects were obtained with 10 microM NE. The concentration-response curve and time course for production of Ins(1,4,5)P3 correlated with the neurotransmitter-induced Ca2+ release from intracellular stores, indicating that the level of Ins(1,4,5)P3 regulated the Ca(2+)-release mechanism. In the continued presence of NE, the intracellular pools did not completely refill with Ca2+ despite the return of Ins-(1,4,5)P3 levels to basal at 5 min. These results demonstrate that NE stimulates a rapid increase in Ins(1,4,5)P3 that correlates with contraction in Ca(2+)-free buffer. The reuptake of Ca2+ into intracellular stores is regulated by a mechanism that may not involve Ins(1,4,5)P3