Patulin inhibition of specific apple microbiome members uncovers Hanseniaspora uvarum as a potential biocontrol agent.

Penicillium expansum is a major postharvest pathogen of apples, causing loss in fruits through tissue damage, as well as in apple products due to contamination with the mycotoxin patulin. During infections, patulin is a cultivar-dependent virulence factor that facilitates apple lesion development. Patulin also has characterized antimicrobial activity and is important for inhibiting other competitive phytopathogens, but the role of this inhibitory activity has not been investigated in the context of the apple microbiome. In our current study, we isolated 68 apple microbiota and characterized their susceptibility to P. expansum extracts. We found Gram-negative bacteria and Basidiomycete yeast to demonstrate largely patulin-specific growth inhibition compared to Gram-positive and Ascomycete isolates. From co-cultures, we identified a Hanseniaspora and Gluconobacter pairing that reduced P. expansum biomass and found that Hanseniaspora uvarum alone is sufficient to reduce apple disease progression in vivo. We investigated possible mechanisms of H. uvarum biocontrol activity and found modest inhibition on apple puree plates, as well as a trend toward lower patulin levels at the wound site. Active biocontrol activity required live yeast, which also were effective in controlling Botrytis cinerea apple infections. Lastly, we explored the breadth of H. uvarum biocontrol activity with over 30 H. uvarum isolates and found consistent inhibition of P. expansum apple disease.

Fate and Exposure Assessment of Pb Leachate from Hypothetical Breakage Events of Perovskite Photovoltaic Modules.

Emerging lead halide perovskite (LHP) photovoltaics are undergoing intense research and development due to their outstanding efficiency and potential for low manufacturing costs that render them competitive with existing photovoltaic (PV) technologies. While today's efforts are focused on stability and scalability of LHPs, the toxicity of lead (Pb) remains a major challenge to their large-scale commercialization. Here, we present a screening-level, EPA-compliant model of fate and transport of Pb leachate in groundwater, soil, and air, following hypothetical catastrophic breakage of LHP PV modules in conceptual utility-scale sites. We estimated exposure point concentrations of Pb in each medium and found that most of the Pb is sequestered in soil. Exposure point concentrations of Pb from the perovskite film fell well below EPA maximum permissible limits in groundwater and air even upon catastrophic release from PV modules at large scales. Background Pb levels in soil can influence soil regulatory compliance, but the highest observed concentrations of perovskite-derived Pb would not exceed EPA limits under our assumptions. Nonetheless, regulatory limits are not definitive thresholds of safety, and the potential for increased bioavailability of perovskite-derived Pb may warrant additional toxicity assessment to further characterize public health risks.

Exploring the phylogenetic diversity of Botryosphaeriaceae and Diaporthe species causing dieback and shoot blight of blueberry in Serbia.

Identifying the precise pathogens responsible for specific plant diseases is imperative for implementing targeted and efficient interventions and mitigating their spread. Dieback and shoot blight significantly diminish the lifespan and productivity of blueberries, yet the causative agents remain largely unidentified. To determine the identity and prevalence of the causal agents of branch dieback and shoot blight, we conducted multi-year and multi-site sampling of diseased highbush blueberries (Vaccinium corymbosum cv. Duke) in Serbia. Sixty-nine monosporic isolates were collected and characterized based on morphological, physiological features and multi-locus phylogenetic analysis of internal transcribed spacer (ITS), ß-tubulin (TUB2) and translation elongation factor 1-alpha (TEF1) sequence data. Five species were identified as causal agents: Diaporthe eres (36 isolates), D. foeniculina (3 isolates), Neopestalotiopsis vaccinii (9 isolates), N. rosae (6 isolates) and Neofusicoccum parvum (15 isolates). The results of the pathogenicity tests performed with the 23 representative isolates confirmed the role of these species as primary pathogens in causing dieback and shoot blight of blueberry, with N. parvum being the most aggressive and D. eres the least. Our study underscores the diversity of genera and species of ascomycetes capable of causing blueberry dieback and shoot blight. Furthermore, our findings indicate that the agents responsible for the disease in Serbia differ from those identified in other regions worldwide.

Scientific, ethical, and legal considerations for the inclusion of pregnant people in clinical trials.

Clinical trials to address the COVID-19 public health emergency have broadly excluded pregnant people from participation, illustrating a long-standing trend of clinical trial exclusion that has led to a clear knowledge gap and unmet need in the treatment and prevention of medical conditions experienced during pregnancy and of pregnancy-related conditions. Drugs (includes products such as drugs, biologics, biosimilars and vaccines) approved for a certain medical condition in adults are also approved for use in pregnant adults with the same medical condition, unless contraindicated for use in pregnancy. However, there are limited pregnancy-specific data on risks and benefits of drugs in pregnant people, despite their approval for all adults. The United States Food and Drug Administration-approved medical products are used widely by pregnant people, 90% of whom take at least 1 medication during the course of their pregnancy despite there being sparse data from clinical trials on these products in pregnancy. This overall lack of clinical data precludes informed decision-making, causing clinicians and pregnant patients to have to decide whether to pursue treatment without an adequate understanding of potential effects. Although some United States Food and Drug Administration initiatives and other federal efforts have helped to promote the inclusion of pregnant people in clinical research, broader collaboration and reforms are needed to address challenges related to the design and conduct of trials that enroll pregnant people, and to forge a culture of widespread inclusion of pregnant people in clinical research. This article summarizes the scientific, ethical, and legal considerations governing research conducted during pregnancy, as discussed during a recent subject matter expert convening held by the Duke-Margolis Center for Health Policy and the United States Food and Drug Administration on this topic. This article also recommends strategies for overcoming impediments to inclusion and trial conduct.

The child's perception on monitoring inflammatory bowel disease activity.

To determine the perception of children with inflammatory bowel disease (IBD) regarding monitoring tests, we first compared the reported discomfort and patient perspective during gastro-intestinal (GI)-endoscopy, magnetic resonance entrography (MRE), and ultrasound (US) and, in a second comparison, patient preference on non-invasive tests (venipuncture, sampling stool and US). A cross-sectional study in children 8-18 years undergoing an US, MRE, and GI-endoscopy for diagnosis or follow-up of IBD. After each procedure, the children filled out the Discomfort during research procedures questionnaire (DISCO-RC). Items of the DISCO-RC are as follows: nervousness, annoyance, pain, fright, boredom, and tiredness. Answers range from "not" (= 0 points) to "extremely" (= 4 points) (range total score: 0-24). Differences between the procedures were assessed with Friedman test, with subsequent Wilcoxon signed-rank test. The children were also asked which non-invasive test they preferred not to undergo regularly (venipuncture, stool-sampling, or US). Answers were analyzed with χ2-test. Forty-nine patients (27 (55%) female, median age 15 (range 9-17)) were included. The children reported to be most nervous, frightened, and tired after GI-endoscopy (median: 1, 1, 2 points, respectively), equally annoyed by MRE and GI-endoscopy (median 1 point), and equally bored by MRE and US. GI-endoscopy was ranked as most discomfortable, followed by MRE and US (total DISCO-RC scores: 7 vs. 5 vs. 2, p < 0.001). Most of the children preferred not to sample stool or perform venipuncture regularly (n = 20 (41%, both) (p < 0.001)).Conclusion: Our results suggest that the children with IBD report low discomfort after US, MRE, and GI-endoscopy. US is preferred as a monitoring tool, also among non-invasive monitoring tests. GI-endoscopy was most discomfortable. What is Known: • Children with inflammatory bowel disease need to be monitored frequently for disease activity. • Adult studies - including a systematic review - on acceptability of monitoring tools among IBD patients showed mixed results. What is New: • Children in our study ranked gastro-intestinal endoscopy as most discomfortable, followed by MRE and US. • With regard to non-invasive monitoring, most children preferred not to sample stool or perform venipuncture regularly, and preferred US.

Viral E3 ubiquitin ligase-mediated degradation of a cellular E3: viral mimicry of a cellular phosphorylation mark targets the RNF8 FHA domain.

Viral hijacking of cellular processes relies on the ability to mimic the structure or function of cellular proteins. Many viruses encode ubiquitin ligases to facilitate infection, although the mechanisms by which they select their substrates are often unknown. The Herpes Simplex Virus type-1-encoded E3 ubiquitin ligase, ICP0, promotes infection through degradation of cellular proteins, including the DNA damage response E3 ligases RNF8 and RNF168. Here we describe a mechanism by which this viral E3 hijacks a cellular phosphorylation-based targeting strategy to degrade RNF8. By mimicking a cellular phosphosite, ICP0 binds RNF8 via the RNF8 forkhead associated (FHA) domain. Phosphorylation of ICP0 T67 by CK1 recruits RNF8 for degradation and thereby promotes viral transcription, replication, and progeny production. We demonstrate that this mechanism may constitute a broader viral strategy to target other cellular factors, highlighting the importance of this region of the ICP0 protein in countering intrinsic antiviral defenses.

Young children experience little emotional burden during invasive procedures in asthma research.

Research in children should strike the right balance between protecting underage study subjects and advancing the medical field. This study gives insight into the emotional burden that common invasive research procedures in asthma research have on young children, both from the child and parent perspective. Puppetry was used to stimulate children (age 5-6 years) to explain their emotional burden prior to and after the research procedures. We operationalised emotional burden as willingness to participate in future research and reluctance towards participation. Parents filled out a questionnaire on this topic. Symptomatic patients as well as healthy controls were analysed. Forty-one children were included. Children's anticipatory fear for future research showed a clear decrease of 0.7 ± 1.6 on a 5-point Likert scale as a consequence of participation (p = 0.02). Sixty percent of all participating children explicitly indicated willingness to undergo identical research procedures again. Children uninformed by their parents about the venipuncture were significantly more reluctant to the venipuncture after the procedure (p < 0.01), compared to children who had been informed (4.0 ± 0.9 resp. 2.8 ± 1.2).Conclusion: This study suggests that the emotional burden of participation in asthma research for underage children can be prevented when they are properly informed and decreases as a consequence of participations. We believe increased emphasis should be placed on informing children and evaluating the emotional impact of research to help caretakers and research ethics committees make informed decisions about participation of children in medical research. What is Known: • Medical professionals and parents are likely to overestimate children's discomfort undergoing (invasive) research procedures. • Two thirds of children (age 6-18 years) participating in medical research indicated that they would participate in the same research study again. What is New: • Pre-school children experience little emotional burden during invasive procedures in asthma research. • Proper communication about (invasive) research procedures in pre-school children helps to reduce the anticipatory fear of these procedures in the future.

IMPROVING AFTER-HOURS CALLS TO ENDOCRINOLOGY FELLOWS.

OBJECTIVE: To analyze the frequency and nature of after-hours calls to endocrinology fellows and employ interventions to direct appropriate care to primary endocrinologists. METHODS: The on-call fellows logged calls that came to them during the after-hours and marked them as urgent or nonurgent. We analyzed these calls and then implemented interventions to educate patients on calls that can wait until the next business day. We also trained providers to provide script refills during clinic visits and educated fellows on how to best manage and document these after-hours calls. RESULTS: From July to August 2017, 100 calls were logged. The average number of calls per 24 hours was 1.61, and 47% were marked nonurgent. From January to March 2018, the fellows logged 0.64 calls per 24 hours, and 51% were logged as nonurgent. Most of these calls were for insulin and testing supply refills. CONCLUSION: Many after-hours calls to the fellows were nonurgent and could have waited until the next business day. Our continuing interventions aim at improving both physician and patient satisfaction, as well as patient care.

Cycloaddition Reactions of Azomethine Ylides and 1,3-Dienes on the C2v-Symmetrical Pentakisadduct of C60.

The reactivity of the C2v-symmetric pentakisadduct of C60 with azomethine ylides and conjugated dienes was studied experimentally and computationally. This derivative possesses four [6,6] double bonds, each with unique electrophilicity. The Diels-Alder reaction studied is a regiospecific, kinetically and thermodynamically guided [4 + 2] process producing [5:1]-hexaadducts with an octahedral addition pattern. The kinetically controlled Prato reaction gives a mixture of regioisomeric [5:1]-hexaadducts. The synthesis of geometrically well-defined supramolecular architectures may benefit from these new types of highly functionalized [5:1]-hexaadducts.

Influence of Scale on Biomass Growth and Nutrient Removal in an Algal-Bacterial Leachate Treatment System.

Data collected from experiments conducted at a flask scale are regularly used as input data for life cycle assessments and techno-economic analyses for predicting the potential productivities of large-scale commercial facilities. This study measures and compares nitrogen removal and biomass growth rates in treatment systems that utilize an algae-bacteria consortium to remediate landfill leachate at three scales: small (0.25 L), medium (100 L), and large (1000 L). The medium- and large-scale vessels were run for 52 consecutive weeks as semibatch reactors under variable environmental conditions. The small-scale experiments were conducted in flasks as batch experiments under controlled environmental conditions. Kolomogov-Smirnov statistical tests, which compare the distributions of entire data sets, were used to determine if the ammonia removal, total nitrogen removal, and biomass growth rates at each scale were statistically different. Results from the Kolmogov-Smirnov comparison indicate that there is a significant difference between all rates determined in the large-scale vessels compared to those in the small-scale vessels. These results suggest that small-scale experiments may not be appropriate as input data in predictive analyses of full scale algal processes. The accumulation of nitrite and nitrate within the reactor, observed midway through the experimental process, is attributed to high relative abundances of ammonia- and nitrite-oxidizing bacteria, identified via metagenomic analysis.

Are positive experiences of children in non-therapeutic research justifiable research benefits?

BACKGROUND: Conducting non-therapeutic research is ethically challenging because participation conveys risks and burden and no health benefit. In this paper, we report the positive experiences of a diverse group of healthy and ill children (6-18 years) who participated in non-therapeutic research studies and discuss whether these positive experiences can justifiably be viewed as benefits. METHODS: We used semistructured interviews from an earlier study about children's experiences in clinical research and did a secondary analysis on the positive experiences of the children in the non-therapeutic studies (N=30). Interviews were analysed using 'thematic' analysis. RESULTS: The interviewed children most frequently mentioned as positive experiences of non-therapeutic research participation helping others and the gratification that comes with it, possible health benefits in the future, having fun and new/increased knowledge about the human body, hospitals and doing research. Less frequently mentioned were getting a present, not having to go to school and getting extra attention from healthcare staff. CONCLUSIONS: Our study shows that children participating in non-therapeutic research have various positive experiences while taking part. We argue that some of these justifiably could be taken into the risk-benefit analysis in certain situations or maybe even as a standard part of this analysis. This may help to increase the number of (crucial) non-therapeutic studies with children.

The development of the DISCO-RC for measuring children's discomfort during research procedures.

BACKGROUND: There is a need for data on children's self-reported discomfort in clinical research, helping ethics committees to make their evaluation of discomfort described in study protocols evidence-based. Since there is no appropriate instrument to measure children's discomfort during medical research procedures, we aimed to develop a generic, short and child-friendly instrument: the DISCO-RC questionnaire (DISCOmfort in Research with Children). METHODS: This article describes the six steps of the development of the DISCO-RC. First, we updated a literature search on children's self-reported discomfort in clinical research to get insight in what words are used to measure discomfort (step 1). Subsequently, we interviewed 46 children (6-18 years) participating in research to get insight into important forms of discomfort for children (step 2), and asked them about their preferred response option for measuring discomfort (step 3). Next, we consulted nine paediatric research professionals from various backgrounds for input on the content and feasibility of the DISCO-RC (step 4). Based on the previous steps, we developed a draft version of the DISCO-RC, which we discussed with the professionals. The DISCO-RC was then pretested in 25 children to ensure face-validity from the child's perspective and feasibility (step 5). Finally, validity, reliability and internal consistency were tested (step 6). RESULTS: The search-update revealed several words used for measuring discomfort in research (e.g. 'worries', 'unpleasantness'). The interviews gave insight into important forms of discomfort for children in research (e.g. 'pain', 'boredom'). Children preferred a 5-point Likert scale as response option for the DISCO-RC. The experts recommended a short, digital instrument involving different forms of discomfort, and measuring discomfort of individual research procedures. Pretesting of the DISCO-RC resulted in a few layout changes, and feedback from the children confirmed the feasibility of the DISCO-RC. Convergent validity and test-retest reliability were acceptable. Internal consistency based on item-rest correlations and Cronbach's alpha were low, as expected. CONCLUSIONS: The DISCO-RC is a generic, practical and psychometrically sound instrument for measuring children's discomfort during research procedures. It contributes to make the evaluation of discomfort in paediatric research evidence-based. Therefore, we recommend including the DISCO-RC as standard component of paediatric research studies.

Nitrogen removal from raw landfill leachate by an algae-bacteria consortium.

A remediation system for the removal of nitrogen from landfill leachate by a mixed algae-bacteria culture was investigated. This system was designed to treat leachate with minimal inputs and maintenance requirements, and was operated as an open semi-batch reactor in an urban greenhouse. The results of this study showed a maximum nitrogen removal rate of 9.18 mg N/(L·day) and maximum biomass density of 480 mg biomass/L. The ammonia removal rates of this culture increased with increasing initial ammonia concentration; maximum nitrogen removal occurred at an ammonia concentration of 80 mg N-NH3/L. At starting ammonia concentrations above 80 mg N-NH3/L a reduction in nitrogen removal was seen; this inhibition is hypothesized to be caused by ammonia toxicity. This inhibiting concentration is considerably higher than that of many other published studies.

Genomes in conflict: maintaining genome integrity during virus infection.

The cellular surveillance network for sensing and repairing damaged DNA prevents an array of human diseases, and when compromised it can lead to genomic instability and cancer. The carefully maintained cellular response to DNA damage is challenged during viral infection, when foreign DNA is introduced into the cell. The battle between virus and host generates a genomic conflict. The host attempts to limit viral infection and protect its genome, while the virus deploys tactics to eliminate, evade, or exploit aspects of the cellular defense. Studying this conflict has revealed that the cellular DNA damage response machinery comprises part of the intrinsic cellular defense against viral infection. In this review we examine recent advances in this emerging field. We identify common themes used by viruses in their attempts to commandeer or circumvent the host cell's DNA repair machinery, and highlight potential outcomes of the conflict for both virus and host.

A viral E3 ligase targets RNF8 and RNF168 to control histone ubiquitination and DNA damage responses.

The ICP0 protein of herpes simplex virus type 1 is an E3 ubiquitin ligase and transactivator required for the efficient switch between latent and lytic infection. As DNA damaging treatments are known to reactivate latent virus, we wished to explore whether ICP0 modulates the cellular response to DNA damage. We report that ICP0 prevents accumulation of repair factors at cellular damage sites, acting between recruitment of the mediator proteins Mdc1 and 53BP1. We identify RNF8 and RNF168, cellular histone ubiquitin ligases responsible for anchoring repair factors at sites of damage, as new targets for ICP0-mediated degradation. By targeting these ligases, ICP0 expression results in loss of ubiquitinated forms of H2A, mobilization of DNA repair proteins and enhanced viral fitness. Our study raises the possibility that the ICP0-mediated control of histone ubiquitination may link DNA repair, relief of transcriptional repression, and activation of latent viral genomes.

The intrinsic antiviral defense to incoming HSV-1 genomes includes specific DNA repair proteins and is counteracted by the viral protein ICP0.

Cellular restriction factors responding to herpesvirus infection include the ND10 components PML, Sp100 and hDaxx. During the initial stages of HSV-1 infection, novel sub-nuclear structures containing these ND10 proteins form in association with incoming viral genomes. We report that several cellular DNA damage response proteins also relocate to sites associated with incoming viral genomes where they contribute to the cellular front line defense. We show that recruitment of DNA repair proteins to these sites is independent of ND10 components, and instead is coordinated by the cellular ubiquitin ligases RNF8 and RNF168. The viral protein ICP0 targets RNF8 and RNF168 for degradation, thereby preventing the deposition of repressive ubiquitin marks and counteracting this repair protein recruitment. This study highlights important parallels between recognition of cellular DNA damage and recognition of viral genomes, and adds RNF8 and RNF168 to the list of factors contributing to the intrinsic antiviral defense against herpesvirus infection.

Development of failure scenarios for biosolids land application risk assessment.

Although deviations from standard guidance for land application of biosolids occur in practice, their importance is largely unknown. A list of such deviations (plausible failure scenarios) were identified at a workshop of industry, regulators, and academic professionals. Next, a survey of similar professionals was conducted to rank the plausible failure scenarios according to their severity, frequency, incentive to ignore control measures, gaps in existing control processes, public concern, and overall concern. Survey participants rated intentional dumping (unpermitted disposal) as the most severe of the failure scenarios, lack of worker protection as the most frequent scenario, and application of Class A biosolids that have failed to meet treatment standards as the scenario for which incentives to ignore control measures are highest. Failure of public access restrictions to application sites was the scenario for which existing controls were judged the weakest; application of biosolids too close to wells was ranked highest for public concern and for overall concern. Two scenarios for which existing controls were considered weaker, site restriction violations and animal contact leading to human exposure, were also rated as frequently occurring. Both scenarios are related in that they (1) involve inappropriate access to a site before the required time has elapsed, and (2) could be addressed through similar biosolids management measures.

Management and control of patients with type 2 diabetes mellitus in Lebanon: results from the International Diabetes Management Practices Study (IDMPS).

BACKGROUND: The IDMPS is a study to identify changes in diabetes treatment practice in several developing countries. This paper focuses on diabetes management and compliance with guidelines in a Middle Eastern country like Lebanon. METHODS: The cross-sectional data from the 2006 wave of two weeks duration on the Lebanese population along with the longitudinal data of a 9-month follow-up study were collected. RESULTS: A large proportion of Lebanese patients were not adequately controlled or followed up. A slight proportion was managed by diet and exercise alone while most patients were on two or more oral anti-hyperglycemics. Metformin was the most common monotherapy followed by sulfonylureas. 22.6% of Lebanese patients were on insulin, most commonly basal insulin alone followed by premix insulin alone. Blood glucose self-monitoring was more frequently done by insulinized patients and was associated with better glycemic control. Glycemic control was reached in 29.6% of type 2 patients (HbA1c < 7%) with poorest outcome for patients on insulin and was more frequently achieved in patients who had more frequent monitoring of HbA1c levels. CONCLUSION: For a proper assessment of diabetic control, maintaining adherence to international guidelines needs to be evaluated. Promoting patient education, improving physician knowledge with better implementation of guidelines is recommended.

Chromatin at the intersection of viral infection and DNA damage.

During infection, viruses cause global disruption to nuclear architecture in their attempt to take over the cell. In turn, the host responds with various defenses, which include chromatin-mediated silencing of the viral genome and activation of DNA damage signaling pathways. Dynamic exchanges at chromatin, and specific post-translational modifications on histones have recently emerged as master controllers of DNA damage signaling and repair. Studying viral control of chromatin modifications is identifying histones as important players in the battle between host and virus for control of cell cycle and gene expression. These studies are revealing new complexities of the virus-host interaction, uncovering the potential of chromatin as an anti-viral defense mechanism, and also providing unique insights into the role of chromatin in DNA repair.

Anti-Cholinesterase Activity of Chalcone Derivatives: Synthesis, In Vitro Assay and Molecular Docking Study.

BACKGROUND: Chalcones, originated from natural product, have been broadly studied their biological activity against various proteins which at the molecular level, are responsible for the progress of the diseases in cancer (e.g. kinases), inflammation (oxidoreductases), atherosclerosis (cathepsins receptor), and diabetes (e.g. α-glucosidase). OBJECTIVE: Here we synthesize 10 chalcone derivatives to be evaluated their in vitro enzymatic inhibition activity against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). METHODS: The synthesis was carried out using Claissen-Schimdt condensation and the in vitro assay was conducted using Ellman Method. RESULTS: Compounds 2b and 4b demonstrated as the best IC50 of 9.3 µM and 68.7 µM respectively, towards AChE and BChE inhibition. Molecular docking studies predicted that this activity might be due to the interaction of the chalcones with important amino acid residues in the binding site of AChE such as SER200 and in that of BChE, such as TRP82, SER198, TRP430, TYR440, LEU286 and VAL288. CONCLUSION: Chalcone can be used as the scaffold for cholinesterase inhibitor, in particularly either fluorine or nitro group to be augmented at the para-position of Ring B, whereas the hydrophobic chain is necessary at the meta-position of Ring B.