bluuebunny/crossref_metadata_2025 huggingface dataset:
The rapid emergence ofSalmonellaTyphi with decreased ciprofloxacin susceptibility following an increase in ciprofloxacin prescriptions in Blantyre, Malawi
Authors: Ashton Philip M. , Chirambo Angeziwa Chunga , Meiring James E. , Patel Priyanka D. , Mbewe Maurice , Silungwe Niza , Chizani Kenneth , Banda Happy , Heyderman Robert S. , Dyson Zoe A. , MacPherson Peter , Henrion Marc Y.R. ,   STRATAA Study Group, Holt Kathryn E. , Gordon Melita A.
Published: March 2023

Ciprofloxacin is the first-line drug for treating typhoid fever in many high burden countries in Africa, but the emergence of non-susceptibility poses a grave challenge to public health programmes. Through enhanced surveillance as part of vaccine evaluation, we set out to investigate the occurrence and determinants of ciprofloxacin non-susceptibility in Blantyre, Malawi.

We performed systematic typhoid fever and antibiotic prescription surveillance in two health centres in Blantyre, Malawi between 01/10/2016 and 31/10/2019, as part of the STRATAA and TyVAC studies. Blood culture isolates from study participants underwent i) pefloxacin screening and ciprofloxacin E-tests to identify ciprofloxcain non-susceptibility and ii) whole genome sequencing (WGS) to identify drug resistance mutations and phylogenetic relationships between non-susceptible and sensitive isolates. We constructed generalised linear regression models to investigate associations between ciprofloxacin prescription rates andS.Typhi isolates with Quinolone Resistance Determining Region (QRDR) mutations.

We carried out 11295 blood cultures and microbiologically confirmed 239 cases of typhoid fever, with isolates from 193 participants sequenced (mean age of participants with sequenced genomes 12.8 years, 47% male). Between October 2016 and August 2019 2% (n=4/175) of WGS-confirmed typhoid fever cases were caused byS.Typhi with QRDR mutations, compared with 33% (n=6/18) in September and October 2019. Nine of the tenS.Typhi with QRDR mutations had a decreased ciprofloxacin susceptibility phenotype. Every additional prescription of ciprofloxacin given to study participants in the preceding month was associated with a 4.2% increase in the relative risk of isolatingS.Typhi with a QRDR mutation (95% CI, 1.8-7.0%, p=0.0008). Phylogenetic analysis showed thatS.Typhi isolates with QRDR mutations in September/October 2019 belonged to two distinct sub-clades encoding two different QRDR mutations, and were closely related (0-6 SNPs) to susceptibleS.Typhi endemic to Blantyre.

We have shown a close temporal association between empiric antimicrobial usage with an increase of fluoroquinolone non-susceptibility inS. Typhi, with two sub-clades responsible for the increase. Decreasing ciprofloxacin usage by improving typhoid diagnostics could help to limit the emergence of resistance.
Advancing Understanding of Inequities in Rare Disease Genomics
Authors: Serrano Jillian G , O’Leary Melanie , VanNoy Grace , Holm Ingrid A , Fraiman Yarden S , Rehm Heidi L , O’Donnell-Luria Anne , Wojcik Monica H
Published: March 2023

Purpose

Advances in genomic research have led to the diagnosis of rare, early-onset diseases for thousands of individuals. Unfortunately, the benefits of advanced genetic diagnostic technology are not distributed equitably among the population, as has been seen in many other healthcare contexts. Even quantifying and describing inequities in genetic diagnostic yield is challenging due to variation in referrals to clinical genetics practices and other barriers to clinical genetic testing.

Methods

The Rare Genomes Project (RGP) at the Broad Institute of MIT and Harvard offers research genome sequencing to individuals with rare disease who remain genetically undiagnosed through direct interaction with the individual or family. This presents an opportunity for diagnosis beyond the clinical context, thus eliminating many barriers to access.

Findings

An initial goal of RGP was to equalize access to genomic sequencing by decoupling testing access from proximity to a major medical center and physician referral. However, our study participants are overwhelmingly non-disadvantaged, as evidenced by their access to specialist care and genetic testing prior to RGP enrollment, and are also predominantly white.

Implications

We therefore describe our novel initiative to diversify RGP enrollment in order to advance equity in rare disease genetic diagnosis and research. In addition to the moral imperative of medical equity, this is also critical in order to fully understand the genomic underpinnings of rare disease. We utilize a mixed methods approach to understand the priorities and values of underrepresented communities, existing disparities, and the obstacles to addressing them: all of which is necessary to promote equity in future genomic medicine initiatives.
Amygdala-related electrical fingerprint is modulated with neurofeedback training and correlates with deep-brain activation: Proof-of-concept in borderline personality disorder
Authors: Zopfs Malte , Jindrová Miroslava , Gurevitch Guy , Keynan Jackob N. , Hendler Talma , Baumeister Sarah , Aggensteiner Pascal-M. , Cornelisse Sven , Brandeis Daniel , Schmahl Christian , Paret Christian
Published: March 2023

Background

The modulation of brain circuits of emotion is a promising pathway to treat Borderline Personality Disorder (BPD). Precise and scalable approaches have yet to be established. Two studies investigating the Amygdala-related Electrical Fingerprint (Amyg-EFP) in BPD are presented: One study addressing the deep-brain correlates of Amyg-EFP, and a second study investigating neurofeedback (NF) as a means to improve brain self-regulation.

Methods

Study 1 combined EEG and simultaneous fMRI to investigate the replicability of Amyg-EFP-related brain activation found in the reference dataset (N=24 healthy subjects, 8 female; re-analysis of published data) in the replication dataset (N=16 female individuals with BPD). In the replication dataset, we additionally explored how the Amyg-EFP would map to neural circuits defined by the Research Domain Criteria. Study 2 investigated a 10-session Amyg-EFP NF training in parallel to a 12-weeks residential Dialectical Behavior Therapy (DBT) program. N=15 patients with BPD completed the training, N=15 matched patients served as DBT-only controls.

Results

Study 1 replicated previous findings and showed significant amygdala BOLD-activation in a whole-brain regression analysis with the Amyg-EFP. Neurocircuitry activation (negative affect, salience, and cognitive control) was correlated with the Amyg-EFP signal. Study 2 showed significant learning of Amyg-EFP modulation with NF training. No clinical benefits of NF beyond DBT-only were observed.

Conclusions

Recorded via scalp EEG, the Amyg-EFP picks up brain activation of high relevance for emotion. Administering Amyg-EFP NF in addition to standardized BPD treatment was shown to be feasible. Clinical utility remains to be investigated.
Lasting first impression: Pre-existing immunity restricts mucosal antibody responses during Omicron breakthrough
Authors: Selva Kevin John , Ramanathan Pradhipa , Haycroft Ebene Regina , Reynaldi Arnold , Cromer Deborah , Tan Chee Wah , Wang Lin-Fa , Wines Bruce D , Hogarth P Mark , Downie Laura E , Davis Samantha K , Purcell Ruth Amy , Kent Helen E , Juno Jennifer A , Wheatley Adam K , Davenport Miles P , Kent Stephen John , Chung Amy W
Published: March 2023

Understanding mucosal antibody responses from SARS-CoV-2 infection and/or vaccination is crucial to develop strategies for longer term immunity, especially against emerging viral variants. We profiled serial paired mucosal and plasma antibodies from: COVID-19 vaccinated only vaccinees (vaccinated, uninfected), COVID-19 recovered vaccinees (convalescent, vaccinated) and individuals with breakthrough Delta or Omicron BA.2 infections (vaccinated, infected). Saliva from COVID-19 recovered vaccinees displayed improved antibody neutralizing activity, FcγR engagement and IgA compared to COVID-19 uninfected vaccinees. Furthermore, repeated mRNA vaccination boosted SARS-CoV-2-specific IgG2 and IgG4 responses in both mucosa biofluids (saliva and tears) and plasma. IgG, but not IgA, responses to breakthrough COVID-19 variants were dampened and narrowed by increased pre-existing vaccine-induced immunity to the ancestral strain. Salivary antibodies delayed initiation of boosting following breakthrough COVID-19 infection, especially Omicron BA.2, however, rose rapidly thereafter. Our data highlight how pre-existing immunity shapes mucosal SARS-CoV-2-specific antibody responses and has implications for long-term protection from COVID-19.
Growth defect of domain III glycoprotein B mutants of human cytomegalovirus reverted by compensatory mutations co-localizing in post-fusion conformation
Authors: Madlen Mollik , Lukas Eisler , Büsra Külekci , Elisabeth Puchhammer-Stöckl , Irene Görzer
Published: March 2023

Cell entry is a crucial step for a virus to infect a host cell. Human cytomegalovirus (HCMV) utilizes the glycoprotein B (gB) to fuse the viral and host cell membrane upon receptor binding of gH/gL-containing complexes. Fusion is mediated by major conformational changes of gB from a metastable pre-fusion to a stable post-fusion whereby the central trimeric coiled-coils, formed by domain (D) III α helices, remain structurally nearly unchanged. To better understand the role of the stable core, we individually introduced three potentially helix-breaking and one disulfide bond-breaking mutation in the DIII α3 to alter the gB stability, and studied different aspects of the viral behavior upon long-term culturing. Two of the three helix-breaking mutations were lethal for the virus in either fibroblasts or epithelial cells and the third substitution led from mild to severe effects on viral replication and infection efficiency. gB_Y494P and gB_I495P suggest that the pre-fusion conformation was stabilized and the fusion process inhibited, gB_G493P on the other hand displayed a delayed replication increase and spread, more pronounced in epithelial cells, hinting at an impaired fusion. Interestingely, the disulfide bond-breaker mutation, gB_C507S, performed strikingly different in the two cell types – lethal in epithelial cells and an atypical phenotype in fibroblasts, respectively. Replication curve analyses paired with the infection efficiency and the spread morphology suggest a dysregulated fusion process which could be reverted by second-site mutations mapping predominantly to gB DV. This underlines the functional importance of a stable core for a well-regulated DV rearrangement during fusion.

Importance

Human cytomegalovirus (HCMV) can establish a lifelong infection. In most people, the infection follows an asymptomatic course, however it is a major cause of morbidity and mortality in immunocompromised patients or neonates. HCMV has a very broad cell tropism, ranging from fibroblasts to epi- and endothelial cells. It uses different entry pathways utilizing the core fusion machinery consisting of glycoprotein complexes gH/gL and gB. The fusion protein gB undergoes severe rearrangements from a metastable pre-fusion to a stable post-fusion. Here, we were able to characterize the viral behavior after the introduction of four single point mutations in gBs central core. These led to various cell type-specific atypical phenotypes and the emergence of compensatory mutations, demonstrating an important interaction between domains III and V. We provide a new basis for the delevopment of recombinant stable pre-fusion gB which can further serve as a tool for the drug and vaccine development.
Untargeted metabolomics profiling in pediatric patients and adult populations indicates a connection between lipid imbalance and epilepsy
Authors: Oja Kaisa Teele , Ilisson Mihkel , Reinson Karit , Muru Kai , Reimand Tiia , Peterson Hedi , Fishman Dmytro , Esko Tõnu , Haller Toomas , Kronberg Jaanika , Wojcik Monica H. , Kennedy Adam , Michelotti Gregory , O’Donnell-Luria Anne , Õiglane-Šlik Eve , Pajusalu Sander , Õunap Katrin
Published: March 2023

Introduction

Epilepsy is a common central nervous system disorder characterized by abnormal brain electrical activity. We aimed to compare the metabolic profiles of plasma from patients with epilepsy across different etiologies, seizure frequency, seizure type, and patient age to try to identify common disrupted pathways.

Material and methods

We used data from three separate cohorts. The first cohort (PED-C) consisted of 31 pediatric patients with suspicion of a genetic disorder with unclear etiology; the second cohort (AD-C) consisted of 250 adults from the Estonian Biobank (EstBB), and the third cohort consisted of 583 adults ≥ 69 years of age from the EstBB (ELD-C). We compared untargeted metabolomics and lipidomics data between individuals with and without epilepsy in each cohort.

Results

In the PED-C, significant alterations (p-value <0.05) were detected in sixteen different glycerophosphatidylcholines (GPC), dimethylglycine and eicosanedioate (C20-DC). In the AD-C, nine significantly altered metabolites were found, mainly triacylglycerides (TAG), which are also precursors in the GPC synthesis pathway. In the ELD-C, significant changes in twenty metabolites including multiple TAGs were observed in the metabolic profile of participants with previously diagnosed epilepsy. Pathway analysis revealed that among the metabolites that differ significantly between epilepsy-positive and epilepsy-negative patients in the PED-C, the lipid superpathway (p = 3.2*10-4) and phosphatidylcholine (p = 9.3*10-8) and lysophospholipid (p = 5.9*10-3) subpathways are statistically overrepresented. Analogously, in the AD-C, the triacylglyceride subclass turned out to be statistically overrepresented (p = 8.5*10-5) with the lipid superpathway (p = 1.4*10-2). The presented p-values are FDR-corrected.

Conclusion

Our results suggest that cell membrane fluidity may have a significant role in the mechanism of epilepsy, and changes in lipid balance may indicate epilepsy. However, further studies are needed to evaluate whether untargeted metabolomics analysis could prove helpful in diagnosing epilepsy earlier.
Genome-wide polygenic risk score for muscle strength predicts lower risk for common diseases and longer life span among the Finnish population: a prospective population based cohort study of the 342 443 FinnGen participants
Authors: Herranen Päivi , Koivunen Kaisa , Palviainen Teemu ,   FinnGen, Kujala Urho M , Ripatti Samuli , Kaprio Jaakko , Sillanpää Elina
Published: March 2023

Purpose

To use a genome-wide polygenic risk score for hand grip strength (PRS HGS) to investigate whether the muscle strength genotype predicts the most common age related noncommunicable diseases, survival from acute adverse health events, and all cause mortality.

Methods

This study consisted of 342 443 Finnish biobank participants from FinnGen Data Freeze 10 (53% women) aged 40 to 108 with combined genotype and health registry data. Associations were explored with a linear or Cox proportional hazards regression models.

Results

A higher PRS HGS predicted a lower body mass index (BMI) (β = −0.112 kg/m2, standard error (SE) = 0.017,P= 1.69 × 10−11) in women but not in men (β = 0.004 kg/m2,P= 0.768, sex by PRS HGS interaction:P= 2.12 × 10−07). In all participants, a higher PRS HGS was associated with a lower risk for obesity diagnosis (hazard ratio 0.94, 95% confidence interval 0.93 to 0.95), type 2 diabetes (0.95, 0.94 to 0.96), ischemic heart diseases (0.97, 0.96 to 0.97), hypertension (0.97, 0.96 to 0.97), stroke (0.97, 0.96 to 0.98), asthma (0.94, 0.93 to 0.95), chronic obstructive pulmonary disease (0.94, 0.92 to 0.95), polyarthrosis (0.90, 0.88 to 0.92), knee arthrosis (0.98, 0.97 to 0.99), rheumatoid arthritis (0.95, 0.94 to 0.97), osteoporosis (0.95, 0.93 to 0.97), falls (0.98, 0.98 to 0.99), depression (0.95, 0.94 to 0.96), and vascular dementia (0.93, 0.89 to 0.96). In women only, a higher PRS predicted a lower hazard for any dementia (0.94, 0.92 to 0.96) and Alzheimer’s disease (0.96, 0.93 to 0.98). Participants with a higher PRS HGS had a decreased risk of cardiovascular (0.96, 0.95 to 0.98) and all cause mortality (0.97, 0.96 to 0.98). However, the predictive value of the PRS HGS for mortality was not pronounced after adverse acute health events compared to the non-diseased period.

Conclusions

The genotype that supports higher muscle strength protects against many future health adversities. Further research is needed to investigate whether or how a favourable lifestyle modifies this intrinsic capacity to resist diseases, and if the impacts of lifestyle behaviour on health differ due to polygenic risk.
Transcription decouples estrogen-dependent changes in enhancer-promoter contact frequencies and spatial proximity
Authors: Acuña Luciana I. Gómez , Flyamer Ilya , Boyle Shelagh , Friman Elias , Bickmore Wendy A.
Published: March 2023

How enhancers regulate their target genes in the context of 3D chromatin organization is extensively studied and models which do not require direct enhancer-promoter contact have recently emerged. Here, we use the activation of estrogen receptor-dependent enhancers in a breast cancer cell line to study enhancer-promoter communication. This allows high temporal resolution tracking of molecular events from hormone stimulation to efficient gene activation. We examine how both enhancer-promoter spatial proximity assayed by DNA fluorescence in situ hybridization, and contact frequencies resulting from chromatin in situ fragmentation and proximity ligation by Capture-C, change dynamically during enhancer-driven gene activation. These orthogonal methods produce seemingly paradoxical results: upon enhancer activation enhancer-promoter contact frequencies increase while spatial proximity decreases. We explore this apparent discrepancy using different estrogen receptor ligands and transcription inhibitors. Our data demonstrate that enhancer-promoter contact frequencies are transcription independent but are influenced by enhancer-bound protein complexes whereas altered enhancer-promoter proximity depends on transcription. Our results emphasize that the relationship between contact frequencies and physical distance in the nucleus, especially over short genomic distances, is not always a simple one.
Short Term Impact of Recycling-Derived Fertilizers on Their P Supply for Perennial Ryegrass (Lolium perenne)
Authors: Deinert Lea , Egeter Bastian , Ikoyi Israel , Forrestal Patrick , Schmalenberger Achim
Published: March 2023

Phosphorus is a finite, essential macronutrient for agriculture. Various nutrient recycling technologies in waste streams management are currently under development in many European countries in order to alleviate the dependency of the EU on imports of non-renewable raw material for the production of mineral phosphorus fertilizers commonly used in agriculture. The resulting products such as struvites and ashes need to be assessed for their application as so-called recycling-derived fertilisers (RDF) in the agricultural sector prior to commercialisation. Albeit high phosphorus abundance in most soils, the phosphorus availability for plant growth promotion in the soil solution is usually low due to strong P sorption in soil and depends vastly on the microbial mobilisation capability of the soil.

To investigate the impact of different phosphorus fertilizers on plant growth and the soil P cycling microbiota, a short-term pot trial was conducted over the period of 54 days.Lolium perenne(var. AberGreen) was grown with application of superphosphate (SP) as inorganic fertiliser, two ashes (poultry litter ash (PLA) and sewage sludge ash (SSA) and two struvites (municipal wastewater struvite (MWS) and commercial CrystalGreen®(CGS) in rates of 20 and 60 kg P ha-1in four replicates. A P-free control (SP0) was also included in the trial.

Post-harvest, a positive correlation between dry weight yield and struvite application was detected, struvite P also was higher readily available and ACP activity was significantly improved for struvites at the high P application rate. The ash RDFs showed a liming effect at 60 kg P ha-1, and PLA60 negatively affected ACP activity, while PLA20 had significantly lowerphoDcopy numbers. P mobilization from phosphonates and phytates was not affected, TCP solubilization was negatively affected by mineral SP fertilizer application at both P concentrations. Overall, the bacterial andphoDharbouring community were not strongly affected by the P fertilization in this study.
Utilizing a nanobody recruitment approach for assessing serine palmitoyltransferase activity in ER sub-compartments of yeast
Authors: Esch Bianca M. , Walter Stefan , Schmidt Oliver , Fröhlich Florian
Published: March 2023

Sphingolipids (SP) are one of the three major lipid classes in eukaryotic cells and serve as structural components of the plasma membrane. The rate-limiting step in SP biosynthesis is catalyzed by serine palmitoyltransferase (SPT). In yeast, SPT consists of two catalytic subunits (Lcb1 and Lcb2), a regulatory subunit (Tsc3), negative regulators (Orm1 and Orm2), and the phosphatidylinositol-4-phosphate (PI4P) phosphatase Sac1, collectively known as the SPOTS complex. Regulating SPT activity enables cells to adapt SP metabolism to changing environmental conditions. Therefore, the Orm proteins are phosphorylated by two signaling pathways originating from either the plasma membrane localized target of rapamycin (TOR) complex 2 or the lysosomal/vacuolar TOR complex 1. Moreover, uptake of exogenous serine is necessary for the regulation of SP biosynthesis, which suggests the existence of differentially regulated SPT pools based on their intracellular localization. However, tools for measuring lipid metabolic enzyme activity in different cellular compartments are currently not available. We have developed a nanobody recruitment system that enables the re-localization of the SPOTS complex to the nuclear or peripheral ER. By combining this system with sphingolipid flux analysis, we have identified two distinct active SPT pools in cells. Our method thus serves as a new and versatile tool to measure lipid metabolism with sub-cellular resolution.
Versatile Product Detection via Coupled Assays for Ultra-high-throughput Screening of Carbohydrate-Active-Enzymes in Microfluidic Droplets
Authors: Ladeveze Simon , Zurek Paul J. , Kaminski Tomasz S. , Emond Stephane , Hollfelder Florian
Published: March 2023

Enzyme discovery and directed evolution are the two major contemporary approaches for the improvement of industrial processes by biocatalysis in various fields. Customization of catalysts for improvement of single enzyme reactions or de novo reaction development is often complex and tedious. The success of screening campaigns relies on the fraction of sequence space that can be sampled, whether for evolving a particular enzyme or screening metagenomes. Ultrahigh-throughput screening (uHTS) based on in-vitro compartmentalization in water-in-oil emulsion of picolitre droplets generated in microfluidic systems allows screening rates >1 kHz (or >107 per day). Screening for Carbohydrate Active Enzymes (CAZymes) catalysing biotechnologically valuable reactions in this format presents an additional challenge, because the released carbohydrates are difficult to monitor in high throughput. Activated substrates with large optically active hydrophobic leaving groups provide a generic optical readout, but the molecular recognition properties of sugars will be altered by incorporation of such fluoro- or chromophores and their typically higher reactivity, as leaving groups with lowered pKa values compared to native substrates make observation of promiscuous reactions more likely. To overcome these issues, we designed microdroplet assays in which optically inactive carbohydrate products are made visible by specific cascades: the primary reaction of an unlabelled substrate leads to an optical signal downstream. Successfully implementing such assays at the picoliter droplet scale allowed us to detect glucose, xylose, glucuronic acid and arabinose as final products of complex oligosaccharide degradation by glycoside hydrolases by absorbance measurements. Enabling the use of uHTS for screening CAZyme reactions that have been thus far elusive will chart a route towards faster and easier development of specific and efficient biocatalysts for biovalorisation, directing enzyme discovery towards catalysts for their natural rather than model substrates.

Graphical abstract / TOC
Altered neurological and neurobehavioral phenotypes in a mouse model of the recurrentKCNB1-p.R306C voltage-sensor variant
Authors: Kang Seok Kyu , Hawkins Nicole A. , Echevarria-Cooper Dennis M. , Baker Erin M. , Dixon Conor J. , Speakes Nathan , Kearney Jennifer A.
Published: March 2023

Pathogenic variants inKCNB1are associated with a neurodevelopmental disorder spectrum that includes global developmental delays, cognitive impairment, abnormal electroencephalogram (EEG) patterns, and epilepsy with variable age of onset and severity. Additionally, there are prominent behavioral disturbances, including hyperactivity, aggression, and features of autism spectrum disorder. The most frequently identified recurrent variant isKCNB1-p.R306C, a missense variant located within the S4 voltage-sensing transmembrane domain. Individuals with the R306C variant exhibit mild to severe developmental delays, behavioral disorders, and a diverse spectrum of seizures. Previousin vitrocharacterization of R306C described loss of voltage sensitivity and cooperativity of the sensor and inhibition of repetitive firing. ExistingKcnb1mouse models include dominant negative missense variants, as well as knockout and frameshifts alleles. While all models recapitulate key features ofKCNB1encephalopathy, mice with dominant negative alleles were more severely affected. In contrast to existing loss-of-function and dominant-negative variants,KCNB1-p.R306C does not affect channel expression, but rather affects voltage-sensing. Thus, modeling R306C in mice provides a novel opportunity to explore impacts of a voltage-sensing mutation inKcnb1. Using CRISPR/Cas9 genome editing, we generated theKcnb1R306Cmouse model and characterized the molecular and phenotypic effects. Heterozygous and homozygous R306C mice exhibited pronounced hyperactivity, altered susceptibility to flurothyl and kainic acid induced-seizures, and frequent, long runs of spike wave discharges on EEG. This novel model of channel dysfunction inKcnb1provides an additional, valuable tool to studyKCNB1encephalopathies. Furthermore, this allelic series ofKcnb1mouse models will provide a unique platform to evaluate targeted therapies.
CsrA Shows Selective Regulation of sRNA-mRNA Networks
Authors: Rojano-Nisimura Alejandra Matsuri , Simmons Trevor R. , Leistra Abigail N. , Mihailovic Mia K. , Buchser Ryan , Ekdahl Alyssa M. , Joseph Isabella , Curtis Nicholas C. , Contreras Lydia M.
Published: March 2023

Post-transcriptional regulation, by small RNAs (sRNAs) as well as the global Carbon Storage Regulator A (CsrA) protein, play critical roles in bacterial metabolic control and stress responses. The CsrA protein affects selective sRNA-mRNA networks, in addition to regulating transcription factors and sigma factors, providing additional avenues of cross talk between other stress-response regulators. Here, we expand the known set of sRNA-CsrA interactions and study their regulatory effects.In vitrobinding assays confirm novel CsrA interactions with ten sRNAs, many of which are previously recognized as key regulatory nodes. Of those 10 sRNA, we identify that McaS, FnrS, SgrS, MicL, and Spot42 interact with CsrAin vivo. We find that the presence of CsrA impacts the downstream regulation of mRNA targets of the respective sRNA.In vivoevidence supports enhanced CsrA McaS-csgDmRNA repression and showcase CsrA-dependent repression of thefucPmRNA via the Spot42 sRNA. We additionally identify SgrS and FnrS as potential new sRNA sponges of CsrA. Overall, our results further support the expanding impact of the Csr system on cellular physiology via CsrA impact on the regulatory roles of these sRNAs.
Anti-CRISPR protein mediated degradation of Cas9 in human cells
Authors: de Tacca Luisa Arake , Bondy-Denomy Joseph , Rabuka David , Schelle Michael
Published: March 2023

Bacteriophages encode anti-CRISPR (Acr) proteins that inactivate CRISPR-Cas bacterial immune systems, allowing successful invasion, replication, and prophage integration. Acr proteins inhibit CRISPR-Cas systems using a wide variety of mechanisms. AcrIIA1 is encoded by numerous phages and plasmids, binds specifically to the Cas9 HNH domain, and was the first Acr discovered to inhibit SpyCas9. Here we report the observation of AcrIIA1-induced degradation of SpyCas9 and SauCas9 in human cell culture, the first example of Acr-induced degradation of CRISPR-Cas nucleases in human cells. Optimized expression of AcrIIA1 in human cells provided robust inhibition of SpyCas9 editing but had no impact on Type V CRIPSR-Cas12a, consistent with its binding site on the HNH domain. Targeted Cas9 protein degradation by AcrIIA1 could modulate Cas9 nuclease activity in human therapies. The small size and specificity of AcrIIA1 could be used in a CRISPR-Cas proteolysis-targeting chimera (PROTAC), providing a tool for developing safe and precise gene editing applications.
Cell wall proteomics in liveMycobacterium tuberculosisuncovers exposure of ESX substrates to the periplasm
Authors: Jaisinghani Neetika , Previti Mary L. , Andrade Joshua , Askenazi Manor , Ueberheide Beatrix , Seeliger Jessica C.
Published: March 2023

The cell wall of mycobacteria plays a key role in interactions with the environment and its ability to act as a selective filter is crucial to bacterial survival. Proteins in the cell wall enable this function by mediating the import and export of diverse metabolites from ions to lipids to proteins. Accurately identifying cell wall proteins is an important step in assigning function, especially as many mycobacterial proteins lack functionally characterized homologues. Current methods for protein localization have inherent limitations that reduce accuracy. Here we showed that protein tagging by the engineered peroxidase APEX2 within liveMycobacterium tuberculosisenabled the accurate identification of the cytosolic and cell wall proteomes. Our data indicate that substrates of the virulence-associated Type VII ESX secretion system are exposed to theMtbperiplasm, providing insight into the currently unknown mechanism by which these proteins cross the mycobacterial cell envelope.
Hydrogel Capsule-Based Digital Quantitative Polymerase Chain Reaction
Authors: Tan Zheng Lin , Yasuura Masato , Horiguchi Yukichi , Ashiba Hiroki , Fukuda Takashi
Published: March 2023

Droplet digital PCR (ddPCR) is accurate in nucleic acid quantification owing to its linearity and high sensitivity. Amplification of nucleic acid in droplets, however, is limited by the stability of droplets against thermal cycling. While the use of fluorinated oil or supplementation of surfactant could improve the stability of droplets, this process has also greatly increased the cost of ddPCR and limited post-PCR analysis. Here, we report a novel method known as gel capsule-based digital PCR (gc-dPCR) which includes method to prepare hydrogel capsules encapsulating PCR reaction mix, conducting PCR reaction, and readout by either qPCR system or fluorescence microplate reader. We have compared our method to existing methods, i.e., quantitative PCR or vortex ddPCR. Our approach results in higher fluorescence intensity compared to ddPCR suggesting higher sensitivity of the system. As hydrogel capsules are more stable than droplets in fluorinated oil throughout thermal cycling, all partitions can be quantified, thus preventing loss of information from low-concentration samples. Our approach should extend to all droplet-based PCR methods. Generally, our approach has greatly improved ddPCR by increasing droplets stability and sensitivity, and reducing the cost of ddPCR, which help to remove the barrier of ddPCR in settings with limited resources.

Abstract Figure
Improving mRNA vaccine safety and efficiency with cationized lipid nanoparticle formula
Authors: Peng Xu , Liao Guangneng , Ren Dongsheng , Zhou Yucheng , Wu Xiujin , Lei Yingxue , Zhang Yan , Chen Liang , He Chen , Zhang Yaoyi , Yin Hailin , Yang Guang , Xu Kai
Published: March 2023

The widespread use of Covid-19 mRNA vaccines has highlighted the need to address rare but concerning side effects. Systemic off-target gene expression has been identified as a primary cause of acute adverse reactions and side effects associated with nucleoside-modified mRNA vaccines. In this study, we incorporated the permanent cationic lipid Dotap component into the mRNA-LNP formula associated with the FDA-approved mRNA vaccine Comirnaty to create a novel positively charged LNP carrier for mRNA vaccine delivery. Using the optimized LNP formula to prepare SARS-Cov-2 Spike mRNA vaccines for immunogenicity testing, Balb/c mice exhibited improved immunogenicity kinetics with initial antibody titers being lower but showing a continuous upward trend, ultimately reaching levels comparable to those of control mRNA vaccines 8 weeks after boost immunization. The mRNA vaccines encapsulated in the modified LNPs have demonstrated a superior safety profile in respect to systemic delivery of LNP constituents, off-target gene expression, and the systemic pro-inflammatory stimulation. Consequently, it may represent a safer alternative of conventional mRNA-LNP vaccines.
ChatGPT in Healthcare: A Taxonomy and Systematic Review
Authors: Li Jianning , Dada Amin , Kleesiek Jens , Egger Jan
Published: March 2023

The recent release of ChatGPT, a chat bot research project/product of natural language processing (NLP) by OpenAI, stirs up a sensation among both the general public and medical professionals, amassing a phenomenally large user base in a short time. This is a typical example of the ‘productization’ of cutting-edge technologies, which allows the general public without a technical background to gain firsthand experience in artificial intelligence (AI), similar to the AI hype created by AlphaGo (DeepMind Technologies, UK) and self-driving cars (Google, Tesla, etc.). However, it is crucial, especially for healthcare researchers, to remain prudent amidst the hype. This work provides a systematic review of existing publications on the use of ChatGPT in healthcare, elucidating the ‘status quo’ of ChatGPT in medical applications, for general readers, healthcare professionals as well as NLP scientists. The large biomedical literature databasePubMedis used to retrieve published works on this topic using the keyword ‘ChatGPT’. An inclusion criterion and a taxonomy are further proposed to filter the search results and categorize the selected publications, respectively. It is found through the review that the current release of ChatGPT has achieved only moderate or ‘passing’ performance in a variety of tests, and is unreliable for actual clinical deployment, since it is not intended for clinical applications by design. We conclude that specialized NLP models trained on (bio)medical datasets still represent the right direction to pursue for critical clinical applications.
WITHDRAWN: Assessing the Causal Effects of Environmental Tobacco Smoke Exposure: A meta-analytic Mendelian randomisation study
Authors: Woolf Benjamin , Rajasundaram Skanda , Gill Dipender , Sallis Hannah M. , Munafò Marcus R.
Published: March 2023

Withdrawal Statement

The authors have withdrawn their manuscript owing to an issue, which they were recently alerted to, with GWAS of paternal smoking used here. They hope to update the manuscript with an alternative GWAS as soon as possible. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.
Validation of the Strengths and Difficulties Questionnaire (SDQ) emotional subscale in assessing depression and anxiety across development
Authors: Armitage Jessica M , Tseliou Foteini , Riglin Lucy , Dennison Charlotte , Eyre Olga , Bevan Jones Rhys , Rice Frances , Thapar Ajay K , Thapar Anita , Collishaw Stephan
Published: March 2023

Emotional disorders are common in childhood, and their prevalence sharply increases during adolescence. The Strengths and Difficulties Questionnaire (SDQ) is widely used for screening emotional and behavioural difficulties in children and young people, but little is known about the accuracy of the emotional subscale (SDQ-E) in detecting emotional disorders, and whether this changes over development. Such knowledge is important in determining whether symptom changes across age are due to developmental or measurement differences. This study assessed the validity of the SDQ-E and two individual items (low mood and general worry) in differentiating between cases and non-cases of Major Depressive Disorder (MDD), Generalised Anxiety Disorder (GAD), and other anxiety disorders across ages 7, 10, 13, 15, and 25 years in a UK population cohort. Analyses showed moderate accuracy of the subscale in discriminating cases of MDD (AUC=0.67-0.85), and high accuracy for discriminating cases of GAD (AUC=0.80-0.93) and any anxiety disorder (AUC=0.74-0.83) compared to non-cases. The SDQ-E performed well across ages and sex, and generally performed better than the two individual items. Together our findings validate the SDQ-E as a screen for emotional disorders during childhood, adolescence, and early adulthood, and as a tool for longitudinal research on depression and anxiety disorders.
Progressive metastatic infantile fibrosarcoma with multiple acquired mutations
Authors: Furtado Larissa V. , Kacar Marija , Mostafavi Roya , Shi Zonggao , Ruiz Robert , Koo Selene C. , Santiago Teresa , Segers Blair , Krasin Matthew J. , Abramson Zachary R. , Shulkin Barry , Talbot Lindsay J. , Pappo Alberto , Gartrell Jessica
Published: March 2023

Infantile fibrosarcoma is the most common soft-tissue sarcoma in children under the age of 1 yr and is defined molecularly byNTRKfusion proteins. This tumor is known to be locally invasive; however, although rare, metastases can occur. TheNTRKfusion acts as a driver for tumor formation, which can be targeted by first- and second-generationTRKinhibitors. AlthoughNTRKgatekeeper mutations have been well-described as mechanisms of resistance to these agents, alternative pathway mutations are rare. Here, we report the case of a patient with infantile fibrosarcoma treated with chemotherapy andTRKinhibition that developed metastatic, progressive disease with multiple acquired mutations, includingTP53,SUFU, and anNTRKF617L gatekeeper mutation. Alterations in pathways ofSUFUandTP53have been widely described in the literature in other tumors; however, not yet in infantile fibrosarcoma. Although most patients have a sustained response toTRKinhibitors, a subset will go on to develop mechanisms of resistance that have implications for clinical management, such as in our patient. We hypothesize this constellation of mutations contributed to the patient's aggressive clinical course. Taken together, we report the first case of infantile fibrosarcoma withETV6::NTRK3and acquiredSUFU,TP53, andNTRKF617L gatekeeper mutation along with detailed clinical course and management. Our report highlights the importance of genomic profiling in recurrent infantile fibrosarcoma to reveal actionable mutations, such as gatekeeper mutations, that can improve patient outcomes.
Evaluation of factors determining the development and progress of library and information science in different Pakistani universities
Authors: Shah Suhaib Hussain , Pei Lei , Chen Tianyu
Published: March 2023

Purpose

The field of library and information sciences (LIS) is crucial to our educational system. Across the globe, the LIS systems operate at varying levels and rates of efficiency. One of the developing nations is Pakistan, which has LIS systems in all of its colleges, universities and schools. This multimethod study aims to identify and quantify elements that are detrimental to LIS progress as well as evaluate the faculty and infrastructure profiles of universities that offer LIS undergraduate and graduate level programmes.

Design/methodology/approach

Data was collected from the study's participants, who were mainly LIS professionals and faculty at 17 different universities, using survey questionnaires and in-person interviews. This study used a descriptive survey methodology, gathering information through a Google Survey and filling it out with a premade survey proforma. The survey responses were examined using content analysis. The development of LIS instructional and scholarly output is influenced by a variety of factors, which were investigated using a generalized linear model (GLM). To determine whether there was a statistically significant difference in opinion between faculty members and working professionals, as well as between men and women, the outcomes of an independent sample t-test were examined.

Findings

According to our data, the factors that have the biggest impact on the caliber and output of LIS research are “poor writing skills” (3.43), “lack of journal publication fees” (3.51) and “lack of research skills” (3.78). The top GLM model identified poor writing skills, a lack of publication fees and a lack of research expertise as bottleneck characteristics for producing high-quality LIS research. The aforementioned factors were 3.62, 2.41 and 2.19 times more significant than the average factor, respectively, to put it another way.

Originality/value

This study’s analysis revealed that there is no real distinction between the two groups' viewpoints. The results of this study can be applied to problems and challenges associated with LIS education in Pakistani educational institutions.
Explore the relationship between leadership skills, organizational citizenship behavior and organizational culture in Vietnamese public organizations
Authors: Tran Quan Hoang Nguyen
Published: March 2023

Purpose

The purpose of this study is to look into the relationship between leadership skills, especially technical, human and conceptual skills, and organizational citizenship behavior (OCB), with organizational culture acting as a mediator.

Design/methodology/approach

For the purpose of this study, a quantitative strategy was adopted. To collect data, paper/online surveys were administered to 294 individuals in various Vietnamese public organizations. Stata 15.1 was used to assess the reliability, validity and consistency of the measurements. Regression analysis was used to evaluate the proposed hypotheses.

Findings

It was discovered that leadership skills have a significant effect on OCB, which partially supported the hypotheses. However, only conceptual skill was found to have a significant impact on individual OCB (OCBI). Conceptual skill was also found to have a significant impact on supportive culture. In addition, only supportive culture had a significant effect on OCBI. Finally, supportive culture was confirmed to mediate the relationship between conceptual skill and OCBI.

Originality/value

This study adds to the social exchange theory by filling three significant gaps in previous research on leadership skills. First, it investigates the impact of three leadership skills on OCB in public organizations. Second, it examines the role of leadership skills in fostering an organizational culture. Third, the paper concludes by investigating the underlying mechanism through which leadership skills contribute to OCB via the mediation of organizational culture. The implications of these findings for practice are discussed.
Applications of six sigma for service quality enhancement in the UAE: a multiple case study analysis and lessons learned
Authors: Bhat Shreeranga , Antony Jiju , Maalouf Maher , E.V. Gijo , Salah Souraj
Published: March 2023

Purpose

This paper aims to unearth the essential components of Six Sigma for successful deployment and sustainment of service quality in four different organizations in the United Arab Emirates (UAE). More specifically, the paper is intended to determine the motivation to apply Six Sigma, Voice of Customer, Key Performance Indicators (KPIs), Critical-to-Quality (CTQ), readiness factors, Critical Success Factors, sustainment measures, tools and techniques used, challenges/barriers and performance impact on the company.

Design/methodology/approach

An exploratory research methodology with multiple case study analyses was adopted to determine the answers to the research objectives. Four case studies from different service processes of four companies were analyzed. The case studies were collated from these companies via a case study protocol with pre-defined criteria.

Findings

The analysis shows that service operation improvement projects are primarily dependent on the voice of the internal customer, with return on investment in savings as the KPI of the process. Most organizations prefer cycle time and errors as the CTQs in the Six Sigma projects. Even novice users can effectively apply the Six Sigma methodology with external experts’ assistance, mentoring and interventions. Across the case studies, it is observed that the projects were successfully deployed due to the support of top management leadership, effective communication and cross-functional teams. Employee resistance to change is the common barrier observed during the case study analysis. Eventually, in all the four case studies, Six Sigma is executed with standard tools and techniques within the define, measure, analyze, improve, control (DMAIC) approach.

Research limitations/implications

The present study’s findings cannot be generalized due to the limited number of case study analyses in different ecosystems in the UAE. The authors would like to analyse and report more case studies in service quality improvement through the Six Sigma methodology to comprehend and develop a generic roadmap for the deployment of Six Sigma in the UAE service industry.

Practical implications

The study’s findings provide insights into commonalities and differences between the essential factors of Six Sigma deployment and sustainability in UAE companies.

Originality/value

The study results might help the policymakers and key decision makers in UAE and other countries understand the effectiveness of Six Sigma in service quality improvement with its essential factors for deployment.