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Summary: The well‐known role of antibiotics in killing sensitive organisms has been challenged by the effects they exert at subinhibitory concentrations. Unfortunately, there are very few published reports on the advantages these molecules may confer to their producers. This study describes the construction of a genetically verified deletion mutant of Streptomyces flaviscleroticus unable to synthesize chromomycin. This mutant was characterized by a rapid loss of viability in stationary phase that was correlated with high oxidative stress and altered antioxidant defences. Altered levels of key metabolites in the mutant signalled a redistribution of the glycolytic flux toward the PPP to generate NADPH to fight oxidative stress as well as reduction of ATP‐phosphofructokinase and Krebs cycle enzymes activities. These changes were correlated with a shift in the preference for carbon utilization from glucose to amino acids. Remarkably, chromomycin at subinhibitory concentration increased longevity of the non‐producer and restored most of the phenotypic features' characteristic of the wild type strain. Altogether these observations suggest that chromomycin may have antioxidant properties that would explain, at least in part, some of the phenotypes of the mutant. Our observations warrant reconsideration of the secondary metabolite definition and raise the possibility of crucial roles for their producers. [ABSTRACT FROM AUTHOR]

Factor inhibiting HIF (FIH, also known as HIF1AN) is an oxygen-dependent asparaginyl hydroxylase that regulates the hypoxiainducible factors (HIFs). Several proteins containing ankyrin repeat domains (ARDs) have been characterised as substrates of FIH, although there is little evidence for a functional consequence of hydroxylation on these substrates. This study demonstrates that the transient receptor potential vanilloid 3 (TRPV3) channel is hydroxylated by FIH on asparagine 242 within the cytoplasmic ARD. Hypoxia, FIH inhibitors and mutation of asparagine 242 all potentiated TRPV3-mediated current, without altering TRPV3 protein levels, indicating that oxygen-dependent hydroxylation inhibits TRPV3 activity. This novel mechanism of channel regulation by oxygen-dependent asparaginyl hydroxylation is likely to extend to other ion channels. [ABSTRACT FROM AUTHOR]

Biomarkers of the age of mosquitoes are required to determine the risk of transmission of various pathogens as each pathogen undergoes a period of extrinsic incubation in the mosquito host. Using the 2-D Difference Gel Electrophoresis (2-D DIGE) procedure, we investigated the abundance of up to 898 proteins from the Yellow Fever and dengue virus vector, Aedes aegypti, during ageing. By applying a mixed-effects model of protein expression, we identified five common patterns of abundance change during ageing and demonstrated an age-related decrease in variance for four of these. This supported a search for specific proteins with abundance changes that remain tightly associated with ageing for use as ageing biomarkers. Using MALDI-TOF/TOF mass spectrometry we identified ten candidate proteins that satisfied strict biomarker discovery criteria (identified in two out of three multivariate analysis procedures and in two cohorts of mosquitoes). We validated the abundances of the four most suitable candidates (Actin depolymerising factor; ADF, Eukaryotic initiation factor 5A; eIF5A, insect cuticle protein Q17LN8, and Anterior fat body protein; AFP) using semi-quantitative Western analysis of individual mosquitoes of six ages. The redox-response protein Manganese superoxide dismutase (SOD2) and electron shuttling protein Electron transfer oxidoreductase (ETO) were subject to post-translational modifications affecting their charge states with potential effects on function. For the four candidates we show remarkably consistent decreases in abundance during ageing, validating initial selections. In particular, the abundance of AFP is an ideal biomarker candidate for whether a female mosquito has lived long enough to be capable of dengue virus transmission. We have demonstrated proteins to be a suitable class of ageing biomarkers in mosquitoes and have identified candidates for epidemiological studies of dengue and the evaluation of new disease reduction projects targeting mosquito longevity. [ABSTRACT FROM AUTHOR]

Hypoxia-inducible factor (HIF) is a transcriptional activator with a central role in regulating cellular responses to hypoxia. It is also emerging as a major target for viral manipulation of the cellular environment. Under normoxic conditions, HIF is tightly suppressed by the activity of oxygen-dependent prolyl and asparaginyl hydroxylases. The asparaginyl hydroxylase active against HIF, factor inhibiting HIF (FIH), has also been shown to hydroxylate some ankyrin repeat (ANK) proteins. Using bioinformatic analysis, we identified the five ANK proteins of the parapoxvirus orf virus (ORFV) as potential substrates of FIH. Consistent with this prediction, coimmunoprecipitation of FIH was detected with each of the ORFV ANK proteins, and for one representative ORFV ANK protein, the interaction was shown to be dependent on the ANK domain. Immunofluorescence studies revealed colocalization of FIH and the viral ANK proteins. In addition, mass spectrometry confirmed that three of the five ORFV ANK proteins are efficiently hydroxylated by FIH in vitro. While FIH levels were unaffected by ORFV infection, transient expression of each of the ORFV ANK proteins resulted in derepression of HIF-1α activity in reporter gene assays. Furthermore, ORFV-infected cells showed upregulated HIF target gene expression. Our data suggest that sequestration of FIH by ORFV ANK proteins leads to derepression of HIF activity. These findings reveal a previously unknown mechanism of viral activation of HIF that may extend to other members of the poxvirus family. [ABSTRACT FROM AUTHOR]

The stability of drugs can affect drug tests and interpretations. A comprehensive study to verify drug stability in Quantisal ® oral fluid (OF) collection device was undertaken in accordance with Australian standard, AS/NZS 4760:2019 (SAI-Global, 2019). The evaluation was performed for the following drugs: (±) amphetamine, (±) methylamphetamine, (±) 3,4-methylenedioxymethylamphetamine (MDMA), (-)Δ9-tetrahydrocannabinol (THC), cocaine, benzoylecgonine, morphine, codeine, and oxycodone. Stability was assessed at four different storage temperatures over seven time points at ±50% cut-off concentrations (Appendix A, Para A4-4.1, AS/NZS 4760:2019) (SAI-Global, 2019). All drugs were found to be significantly more stable at 4 and -20°C, with stability spanning at least 14 days with percentage change within ±20% from the cut-off concentrations (SAI-Global, 2019). In addition, we report a variation trend with cocaine and benzoylecgonine at elevated temperatures, suggesting hydrolytic decomposition of cocaine and a concomitant increase in benzoylecgonine quantitative values. We confirm the cross-talk by showing that the percentage change in the profile of average cocaine-benzoylecgonine measurement is within the acceptance concentration range of ±20%. This finding highlights the importance of precaution during storage and careful considerations during subsequent interpretation of liquid chromatography-mass spectrometry (LCMS) measurements., Competing Interests: The authors were employed by company Queensland Medical Laboratory (QML) Pathology, under the Healius Pathology Pty Ltd network. QML Pathology has a commercial supplier/customer relationship with Abbott Diagnostics for supply of laboratory reagents, analysers, etc as the laboratory does with many other suppliers. Abbott Diagnostics assisted by providing collection and transportation devices and by recompensing the laboratory for cost of consumables required to perform the validation. This had no influence on the performance of the investigation or the conclusions reached., (Copyright © 2021 Riggio, Dave, Koscak, Blakey and Appleton.)

COVID-19 has emerged as a pandemic of significance with potential to cause significant morbidity and mortality worldwide. Elderly with or without following comorbidities i.e Diabetes, hypertension, cardiac disease, chronic respiratory illnesses, chronic liver disease, CKD, malignancy and immunocompromised hosts are at increased risk of developing complicated course. Hemodialysis population hence are at increased risk for contracting the infection due to patient characteristics, environmental characteristics and procedural lapses. The current study was aimed at describing prevalence and characteristics of COVID19 in hemodialysis population across different HD centers across Mumbai. We found a prevalence rate of COVID19 in 6.4%, with 9 patients (12%) died during the study period. A fair proportion of Non covid HD patients (1.5%) also died due to lack of access to dialysis. At baseline, mean age of presentation was 54.5 years. On routine test 80% were asymptomatic at presentation. Patients with COPD, requiring ICU care and those on ventilation faired poorly. Contrary to assumption patients with underlying cardiovascular disease didn't show poor outcome. Total of 4.1% health care workers turned positive during the study period with mean age of 31 years and median of 28years. Out of them 5 (45.4%) were symptomatic. All recovered from the illness without any sequelae. Seventy two percent of healthcare workers were on Hydroxy-chloroquine chemoprophylaxis didn't reach statistical significance in preventing the infection. In our study elderly age with comorbidities had poor prognosis. We proposed extra healthcare measures to be taken in the dialysis unit presuming all as COVID suspect in the resource limited settings., (© Journal of the Association of Physicians of India 2011.)

Hypoxia-inducible factor (HIF) is a transcriptional activator with a central role in regulating cellular responses to hypoxia. It is also emerging as a major target for viral manipulation of the cellular environment. Under normoxic conditions, HIF is tightly suppressed by the activity of oxygen-dependent prolyl and asparaginyl hydroxylases. The asparaginyl hydroxylase active against HIF, factor inhibiting HIF (FIH), has also been shown to hydroxylate some ankyrin repeat (ANK) proteins. Using bioinformatic analysis, we identified the five ANK proteins of the parapoxvirus orf virus (ORFV) as potential substrates of FIH. Consistent with this prediction, coimmunoprecipitation of FIH was detected with each of the ORFV ANK proteins, and for one representative ORFV ANK protein, the interaction was shown to be dependent on the ANK domain. Immunofluorescence studies revealed colocalization of FIH and the viral ANK proteins. In addition, mass spectrometry confirmed that three of the five ORFV ANK proteins are efficiently hydroxylated by FIH in vitro While FIH levels were unaffected by ORFV infection, transient expression of each of the ORFV ANK proteins resulted in derepression of HIF-1α activity in reporter gene assays. Furthermore, ORFV-infected cells showed upregulated HIF target gene expression. Our data suggest that sequestration of FIH by ORFV ANK proteins leads to derepression of HIF activity. These findings reveal a previously unknown mechanism of viral activation of HIF that may extend to other members of the poxvirus family., Importance: The protein-protein binding motif formed from multiple repeats of the ankyrin motif is common among chordopoxviruses. However, information on the roles of these poxviral ankyrin repeat (ANK) proteins remains limited. Our data indicate that the parapoxvirus orf virus (ORFV) is able to upregulate hypoxia-inducible factor (HIF) target gene expression. This response is mediated by the viral ANK proteins, which sequester the HIF regulator FIH (factor inhibiting HIF). This is the first demonstration of any viral protein interacting directly with FIH. Our data reveal a new mechanism by which viruses reprogram HIF, a master regulator of cellular metabolism, and also show a new role for the ANK family of poxvirus proteins., (Copyright © 2016 American Society for Microbiology.)