bluuebunny/crossref_metadata_2025 huggingface dataset:
When tomorrow comes too late—the silent threat: why people delay needed medical care in Türkiye
Authors: Uzun Süleyman Utku , Akın Merve
Published: December 2024

Background

Healthcare demand procrastination is a public health concern in Türkiye, with limited research available. This study examines the prevalence of healthcare procrastination and identifies associated factors among patients at Pamukkale University Hospital.

Methods

This cross-sectional study was conducted from 2 to 13 February 2023 in Denizli and involved 503 patients from 15 outpatient clinics. Data on sociodemographic characteristics, healthcare utilization and procrastination behaviors were collected via a structured questionnaire, including the Healthcare Demand Procrastination Scale (HDPS). Descriptive statistics and linear regression were used for analysis.

Results

The participants had a mean age of 41.46 years (±15.34), with 52.9% being female. A significant 77.3% reported delaying healthcare despite needing it. The primary reasons for delays were difficulty in getting appointments (51.5%), long waiting times (21.9%) and COVID-19 concerns (19.9%). The mean HDPS score was 2.29 ± 0.65, indicating moderate procrastination. Regression analysis revealed that younger age (≤35 years) and rural residence significantly predicted higher HDPS scores, whereas receiving health information from professionals was linked to lower procrastination.

Conclusions

Healthcare procrastination is common among patients in Türkiye, particularly among younger individuals and rural residents. Improving healthcare accessibility, reducing wait times and implementing targeted interventions are essential to mitigate this issue and enhance health outcomes.
Age-period-cohort modeling of oesophageal carcinoma risk in a middle eastern country: 1980–2019
Authors: Akhtar Saeed , Al-Shammari Ahmad , Al-Huraiti Mohammad , Al-Anjery Fouzan
Published: December 2024

Background

Understanding of the factors influencing oesophageal cancer trends is crucial. Therefore, this cross-sectional cohort study sought to disentangle the age, period and cohort effects on the trends of oesophageal cancer in Kuwait.

Methods

The data on incident oesophageal carcinoma cases diagnosed between January 1, 1980, through December 31, 2019, and reference population were obtained. Age-period-cohort (APC) analysis was conducted using a loglinear Poisson regression model.

Results

A total of 496 oesophageal carcinoma cases in 12.8 million person-years (i.e. squamous-cell carcinoma, 269, 54.23%), adenocarcinoma,147, 29.64% and unspecified cases, 80,16.13%) were diagnosed. The overall age-standardized incidence rate (per 105 person-years) of oesophageal carcinoma during the study period was 10.51 (95% CI: 6.62–14.41). The APC analysis results showed that the age and birth cohort effects were the significant determinants of declining, and subsequently steadying the oesophageal carcinoma incidence rates.

Conclusions

A substantial decline in oesophageal carcinoma incidence rates was recorded, which significantly varied in all three temporal dimensions. The observed birth cohort patterns suggest changing lifestyle and dietary patterns seem to be responsible for decreasing oesophageal carcinoma risk in Kuwait. Future studies may look for the component causes maintaining the endemicity of oesophageal carcinoma risk in this and similar countries in the region.
Has a Phenomenon Been Born? Causes, Consequences, and Strategies of Gendering De-democratization in Turkey
Authors: Fernandes Raquel Santos
Published: December 2024

This article examines how the de-democratization process influences gendered politics and how opposition to gender equality assisted democratic backsliding in Turkey. Drawing on a grounded theory methodology study, this article contends that anti-gender equality policy aided the Justice and Development Party in de-democratizing the political regime. At the same time, women experience different consequences of gendering de-democratization. The evidence from in-depth fieldwork, including fifty-three semistructured interviews, demonstrates that antithetical groups are more exposed to the phenomenon. Findings suggest that anti-gender equality policy influences women’s participation in the labor market, which goes along with further exposure to domestic violence.
Group II Introns in Archaeal Genomes and the Evolutionary Origin of Eukaryotic Spliceosomal Introns
Authors: Mattick J. S. A. , Malik S.-B. , Delwiche C. F.
Published: December 2024

A key attribute of eukaryotic genomes is the presence of abundant spliceosomal introns that break up many protein-coding genes into multiple exons and must be spliced out during the process of gene expression. These introns are believed to be evolutionarily derived from group II introns, which are known to be widespread in bacteria. One prominent hypothesis is that the spliceosomal intron arose after the endosymbiotic origin of the mitochondrion, as a consequence of transfer of genes containing group II introns from the organelle to nuclear genome; in this model, transfer of group II introns into the ancestral eukaryotic genome set the stage for evolution of the spliceosomal form. However, the recent discovery and sequencing of asgard archaea — the closest archaeal relatives of extant eukaryotes — has shed significant light on the composition of the early eukaryotic genome and calls that model into question. Using sequence analysis and structural modeling, we show here the presence of group II intron maturases in the genomes of Heimdallarchaeia and other asgard archaea, and demonstrate by phylogenetic inference that these are closely related to both eukaryotic mitochondrial group II intron maturases and the spliceosome protein PRP8. This suggests that the first intron-containing eukaryotic common ancestor (FIECA) inherited selfish group II introns from its ancestral archaeal genome – the progenitor of the nuclear genome – rather than from the mitochondrial endosymbiont. These observations suggest that the spread and diversification of introns may have occurred independently of the acquisition of the mitochondrion. To better understand the context for intron evolution, we investigate the broader occurrence of group II introns in archaea, identify archaeal clades enriched in group II introns, and perform structural modeling to examine the relationship between the archaeal group II intron maturase and the eukaryotic spliceosome. We propose a model of intron acquisition and expansion during early eukaryotic evolution that places the spread of introns prior to the acquisition of mitochondria, possibly facilitated by the separation of transcription and translation afforded by the nucleus.
Elevated FGF23 levels predict cardiovascular and cerebrovascular events in acute ischaemic stroke patients
Authors: Gu Lingyun , Shen Liping , Li Weizhang , Zhou Jinfeng , Jin Qi , Yang Jiangsheng , Cui Junyou , Zhu Zufu , Xu Zhuowen , Weng Yulong
Published: December 2024

BACKGROUND

There have been recent studies on fibroblast growth factor 23 (FGF23) and acute ischaemic stroke (AIS), but some of the results were conflicting. Therefore, the objective of this study was to assess whether baseline FGF23 was associated with the occurrence of major adverse cardiovascular and cerebrovascular events (MACCE) in AIS patients.

METHODS

This study enrolled 394 patients with AIS and followed for a median period of 43 months. 130 people who underwent health check-ups in our hospital were selected as the control group. The study endpoint was the first occurrence of MACCE outside the hospital. Serum FGF23 levels were quantified using an enzyme-linked immunosorbent assay.

RESULTS

Serum FGF23 levels were found to be significantly higher in the AIS group than in the control group (525.14±167.40 vs 338.62±161.71,P<0.05). Furthermore, serum FGF23 levels were observed to be elevated in the MACCE group in comparison to the no MACCE group (646.09±164.05 vs 491.87±152.54,P<0.05). According to the receiver operating characteristic curve analysis, the predictive value of the combination of the three [FGF23, National Institutes of Health Stroke Scale (NIHSS) and modified Rankin Scale (mRS)] for the occurrence of MACCE in AIS was higher than that of any of them (P<0.05), while the difference between the three was not statistically significant. The cumulative MACCE-free survival was found to be significantly higher in the group with low FGF23 levels than in the group with high FGF23 levels, as demonstrated by Kaplan-Meier analysis (P<0.05). The multifactorial Cox analysis revealed that the elevated baseline serum FGF23 level (HR: 3.731, 95% CI: 2.157-6.452) was a significant predictor of MACCE in AIS patients.

CONCLUSIONS

Elevated baseline serum FGF23 levels were considered to be a valid predictor of MACCE in patients with AIS.
Subjective experience of women with childbirth in hospitals after COVID-19 outbreak: Insights from Babies Born Better survey
Authors: Greškovičová Katarína , Němcová Mariana , Šiková Tereza
Published: December 2024

After COVID-19 outbreak, there has been changes in maternity care suggesting that childbirth experience was also change. This study thus investigates the impact of the COVID-19 pandemic on childbirth experiences in Slovak hospitals, focusing on women’s perspectives to childbirth experience. Utilizing data from the Babies Born Better survey, we analyzed responses from 810 women who gave birth in Slovakia between March 2020 and June 2022. Via inductive content analysis we identified 4 main themes: Compassionate and Supportive Care, Medical Expertise and Care, Autonomy and Empowerment, and External Conditions. Our research has provided further evidence of the multifaceted nature of childbirth experience. The childbirth experience in a period of general adversity (such as pandemics) may have not been captured in this research or it may not significantly differ from periods of non-adversity. What is important for women remain unchanged - compassionate and supportive care from healthcare providers. Based on our findings we propose improvements in maternal healthcare during childbirth. These improvements aim not only to improve womeńs childbirth experience but also foster better healthcare outcomes for professionals and hospitals.
PREVALENCE OF ANEMIA, IRON DEFICIENCY ANEMIA, AND ASSOCIATED FACTORS AMONG DIABETICS IN THE WEST REGION of CAMEROON
Authors: Tedong Leonard , Mekieje Maï-Pamela Tumchou , Djeukamgang Melisa Mene , De Manfouo Romaric Tuono , Tchoumke Marius Mbiandjeu ,   Pascal Blaise Well a Well a Koul, Simo Josué Louokdom , Chuisseu Djamen Pascal Dieudonne
Published: December 2024

Aims and objective

Anemias are a burden for management because they are associated with the worsening of the pathophysiology of diseases. They can be multifactorial, including micronutrient deficiencies including iron deficiencies. This study aimed to biologically characterize anemias and iron deficiency anemias during diabetes and to identify the associated factors.

Methods

Over 4 months, we conducted an analytical cross-sectional study from August to November 2024. 210 diabetic patients, attending the diabetology unit of the regional hospital in West Cameroon were studied, their age range between <20 and > 70 years. Blood samples were taken for biological tests including blood count, glycemia, glycated hemoglobin, ferritin, and serum following standard assay protocols. The results obtained were analyzed using the R statistical tool version 4.1.1.

Results

The sex ratio was 0.89 and the mean age was 48.32±18.19 years. 48.6% of patients were overweight. The frequency of anemia was observed in 72 females (53%) and 63 males (46.7%) with a total frequency of 64.3%. 11.4% were found to have iron deficiency anemias. The blood count revealed hematological abnormalities characterized by leukopenia (41.4%), granulocytopenia (35.7%), and thrombocytopenia (2.86%) with macroplateletosis (40.0%). Anemias were characterized by microcytosis (11.1% in anemic patients versus none in non-anemic patients (p<0.001), hypochromia (88.9% in anemic patients versus 72% in non-anemic patients (p<0.001)) and greater regeneration in anemic patients versus non-anemic patients (p<0.001). Macroplateletosis and high Hb1AC were identified as predictive factors for iron deficiency anemia in diabetes with (Or=2.83, 95% CI= [1.17;6.80];p=0.021) and Or=2.38, 95% CI= [0.14;3.09],p=0.045, respectively.

Conclusion

This study reports a significant frequency of anemia and iron deficiency anemia in diabetic patients; the latter is associated with worsening clinical manifestations in patients. This study raises the need for regular exploration of iron status in diabetics to prevent complications related to iron deficiency.
Leucine Aminopeptidase LyLAP enables lysosomal degradation of membrane proteins
Authors: Jain Aakriti , Heremans Isaac , Rademaker Gilles , Detomasi Tyler C. , Hernandez Grace A. , Zhang Justin , Gupta Suprit , Linde Teresa von , Lange Mike , Spacci Martina , Rohweder Peter , Anderson Dashiell , Citron Y. Rose , Olzmann James A. , Dawson David W. , Craik Charles S. , Bommer Guido , Perera Rushika M. , Zoncu Roberto
Published: December 2024

Proteolysis of hydrophobic helices is required for complete breakdown of every transmembrane protein trafficked to the lysosome and sustains high rates of endocytosis. However, the lysosomal mechanisms for degrading hydrophobic domains remain unknown. Combining lysosomal proteomics with functional genomic data mining, we identify Lysosomal Leucine Aminopeptidase (LyLAP; formerly Phospholipase B Domain-Containing 1) as the hydrolase most tightly associated with elevated endocytic activity. Untargeted metabolomics and biochemical reconstitution demonstrate that LyLAP is not a phospholipase, but a processive monoaminopeptidase with strong preference for N-terminal leucine – an activity necessary and sufficient for breakdown of hydrophobic transmembrane domains. LyLAP is upregulated in pancreatic ductal adenocarcinoma (PDA), which relies on macropinocytosis for nutrient uptake, and its ablation led to buildup of undigested hydrophobic peptides, which compromised lysosomal membrane integrity and inhibited PDA cell growth. Thus, LyLAP enables lysosomal degradation of membrane proteins, and may represent a vulnerability in highly endocytic cancer cells.

One sentence summary

LyLAP degrades transmembrane proteins to sustain high endocytosis and lysosomal membrane stability in pancreatic cancer.
Dynamics of the CD9 interactome during bacterial infection of epithelial cells by proximity labelling proteomics
Authors: Wolverson PA , Fernandes Parreira I , Collins MO , Shaw JG , Green LR
Published: December 2024

Epithelial colonisation is often a critical first step in bacterial pathogenesis, however, different bacterial species utilise several different receptors at the cell membrane to adhere to cells. We have previously demonstrated that interference of the human tetraspanin, CD9, can reduce adherence of multiple species of bacteria to epithelial cells by approximately 50%. However, CD9 does not act as a receptor and is responsible for organising and clustering partner proteins commandeered by bacteria for efficient adherence. CD9 can organise numerous host proteins at the cell membrane but the full interactome has not been delineated. Here, using a novel CD9 proximity-labelling model, we demonstrate a vast and diverse CD9 interactome with 845 significantly enriched proteins associated with CD9 over four hours. These putative proximal proteins were associated with various cellular pathways including cell adhesion, ECM-receptor interactions, endocytosis, SNARE interactions and adherens and tight junctions. Significant and known interactors of CD9 were enriched including β1 integrins and major immunoglobulin superfamily members but also included several known bacterial adherence receptors including CD44, CD46 and CD147. We further demonstrate dynamism of the interactome during infection at three separate time points with two different bacterial species,Neisseria meningitidisandStaphylococcus aureus. During meningococcal infection, 13 unique proximal proteins associated with CD9 were significantly enriched across four hours compared to uninfected cells. However, upon staphylococcal infection far fewer enriched proximal proteins were identified demonstrating that different bacteria require different host factors during CD9-mediated bacterial adherence. Transient knockdown of CD44 and CD147, candidate receptor proteins identified in our screen, significantly reduced staphylococcal and meningococcal adherence respectively. This effect was ablated in the absence of CD9 or if epithelial cells were treated with a CD9-derived peptide demonstrating the association of these proteins during staphylococcal and meningococcal adherence. We demonstrate for the first time the CD9 interactome of epithelial cells and that bacteria hijack these interactions to efficiently adhere to epithelial cells. This process is bacterial species specific, recruiting several different proteins during infection but a host-derived peptide is able to interfere with this process. We have therefore developed a tool that can measure changes within the CD9 interactome after cellular challenge, established a mechanism in which CD9 is used as a universal organiser of bacterial adhesion platforms and demonstrated that this process can be stopped using a CD9-derived peptide.
Typhoid toxin ofSalmonellaTyphi elicits host antimicrobial response during acute typhoid fever
Authors: Srour Salma , ElGhazaly Mohamed , O’Connor Daniel , Gibani Malick M , Darton Thomas C , Pollard Andrew J , Collins Mark O , Humphreys Daniel
Published: December 2024

SalmonellaTyphi secretes typhoid toxin that activates cellular DNA damage responses (DDR) during acute typhoid fever. Human infection challenge studies revealed that the toxin suppresses bacteraemia via unknown mechanisms. Using quantitative proteomic analysis on the plasma of bacteraemic participants, we demonstrate that wild-type toxigenicSalmonellainduced secretion of lysozyme (LYZ) and apolipoprotein C3 (APOC3). Recombinant typhoid toxin or infection with toxigenicSalmonellarecapitulate LYZ and APOC3 secretion in cultured cells, which involved ATM/ATR-dependent DDRs and confirmed observations in typhoid fever. LYZ alone inhibited secretion of virulence effector proteins SipB and SopE. LYZ caused loss ofSalmonellamorphology characterised by spheroplast formation. Spheroplast formation was mediated by LYZ and enhanced by lactoferrin, which was identified by proteomics in participants with typhoid fever. Our findings may indicate that toxin-induced DDRs elicit antimicrobial responses, which suppressSalmonellabacteraemia during typhoid fever.
Redox-dependent binding and conformational equilibria govern the fluorescence decay of NAD(P)H in living cells
Authors: Blacker Thomas S. , Mistry Nimit , Plotegher Nicoletta , Westbrook Elizabeth R. , Sewell Michael D. E. , Carroll John , Szabadkai Gyorgy , Bain Angus J. , Duchen Michael R.
Published: December 2024

When probed using fluorescence lifetime imaging microscopy (FLIM), the emission from reduced nicotinamide adenine dinucleotide (NADH) and its phosphorylated form NADPH have shown promise as sensitive intrinsic reporters of metabolism in living systems. However, an incomplete understanding of the biochemical processes controlling their fluorescence decay makes it difficult to draw unambiguous conclusions from NAD(P)H FLIM data. Here we utilised time-resolved fluorescence anisotropy imaging to identify multiple enzyme binding configurations in live cells associated with lifetimes both longer and shorter than unbound NAD(P)H. FLIM, combined with mathematical and computational modelling, revealed that the redox states of the NAD and NADP pools control the steady-state equilibrium of binding configurations, which in-turn determines the observed fluorescence decay. This knowledge will be foundational to developing the accurate interpretation of NAD(P)H FLIM.
Sp140L Is a Novel Herpesvirus Restriction Factor
Authors: Cable Jana M. , Wongwiwat Wiyada , Grabowski Jenna C. , White Robert E. , Luftig Micah A.
Published: December 2024

Herpesviruses, including the oncogenic Epstein-Barr Virus (EBV), must bypass host DNA sensing mechanisms to establish infection. The first viral latency protein expressed, EBNA-LP, is essential for transformation of naïve B cells, yet its role in evading host defenses remains unclear. Using single-cell RNA sequencing of EBNA-LP-Knockout (LPKO)-infected B cells, we reveal an antiviral response landscape implicating the ‘speckled proteins’ as key restriction factors countered by EBNA-LP. Specifically, loss ofSP100or the primate-specificSP140Lreverses the restriction of LPKO, suppresses a subset of canonically interferon-stimulated genes, and restores viral gene transcription and cellular proliferation. Notably, we also identify Sp140L as a restriction target of the herpesvirus saimiri ORF3 protein, implying a role in immunity to other DNA viruses. This study reveals Sp140L as a restriction factor that we propose links sensing and transcriptional suppression of viral DNA to an IFN-independent innate immune response, likely relevant to all nuclear DNA viruses.
Engineering prolyl hydroxylase-dependent proteolysis enables the orthogonal control of hypoxia responses in plants
Authors: Shukla Vinay , Iacopino Sergio , Dalle Carbonare Laura , He Yuming , Chiaro Alessia Del , Papachristodoulou Antonis , Giuntoli Beatrice , Licausi Francesco
Published: December 2024

Vascular plants and metazoans use selective proteolysis of transcription factors to control the adaptive responses to hypoxia, although through distinct biochemical mechanisms. The reason for this divergence is puzzling, especially when considering that the molecular components necessary to establish both strategies are conserved across the two kingdoms. To explore an alternative evolutionary scenario where plants sense hypoxia as animals do, we engineered a three-components system aimed to target proteins for degradation in an oxygen dependent mannerin Arabidopsis thaliana. Applying the synthetic biology framework, we produced a hypoxia-responsive switch independent of endogenous pathways. When applied to control transcription, the synthetic system partially restored hypoxia responsiveness in oxygen-insensitive mutants. Additionally, we demonstrated its potential to regulate growth under flood-induced hypoxia. Our work highlights the use of synthetic biology to reprogram signalling pathways in plants, providing insights into the evolution of oxygen sensing and ofering tools for crop improvement under stress conditions.
YibN, a bona fide interactor of YidC with implications in membrane protein insertion and membrane lipid production
Authors: Zhao Zhiyu , Yamamoto Nachi , Young John W. , Solis Nestor , Fong Amos , Al-Seragi Mohammed , Kim Sungyoung , Aoki Hiroyuki , Phanse Sadhna , Le Hai-Tuong , Overall Christopher M. , Nishikawa Hanako , Babu Mohan , Nishiyama Ken-ichi , Duong van Hoa Franck
Published: December 2024

YidC, a prominent member of the Oxa1 superfamily, is essential for the biogenesis of the bacterial inner membrane, significantly influencing its protein composition and lipid organization. It interacts with the Sec translocon, aiding the proper folding of multi-pass membrane proteins. It also functions independently, serving as an insertase and lipid scramblase, augmenting the insertion of smaller membrane proteins while contributing to the organization of the bilayer. Despite the wealth of structural and biochemical data available, how YidC operates remains unclear. To investigate this, we employed proximity-dependent biotin labeling (BioID), leading to the identification of YibN as a crucial component within the YidC protein environment. We then demonstrated the association between YidC and YibN by affinity purification-mass spectrometry assays conducted on native membranes, with further confirmation using on-gel binding assays with purified proteins. Co-expression studies andin vitroassays indicated that YibN enhances the production and membrane insertion of YidC substrates, such as M13 and Pf3 phage coat proteins, ATP synthase subunit c, and various small membrane proteins like SecG. Additionally, the overproduction of YibN was found to stimulate membrane lipid production and promote inner membrane proliferation, perhaps by interfering with YidC lipid scramblase activity. Consequently, YibN emerges as a significant physical and functional interactor of YidC, influencing membrane protein insertion and lipid organization.
The CarSR Two-Component System Directly ControlsradDExpression as a Global Regulator that Senses Bacterial Coaggregation inFusobacterium nucleatum
Authors: Bibek G C , Wu Chenggang
Published: December 2024

Two-component systems (TCS) enable bacteria to sense and respond to environmental signals, facilitating rapid adaptation.Fusobacterium nucleatum, a key oral pathobiont, employs the CarSR TCS to modulate coaggregation with various Gram-positive partners by regulating the expression ofradD, encoding a surface adhesion protein, as revealed by RNA-Seq analysis. However, the direct regulation of theradD-containing operon (radABCD) by the response regulator CarR, the broader CarR regulon, and the signals sensed by this system remain unclear. In this study, chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) identified approximately 161 CarR-enriched loci across the genome and a 17-bp consensus motif that likely serves as the CarR binding site. Notably, one such binding motif was found in the promoter region of theradABCDoperon. The interaction of CarR with this binding motif was further validated using electrophoretic mobility shift assays (EMSA), mutagenesis, and DNase I footprinting analyses. Beyond regulatingradABCD, CarR directly controls genes involved in fructose and amino acid (cysteine, glutamate, lysine) utilization, underscoring its role as a global regulator inF. nucleatum. Lastly, we discovered that RadD-mediated coaggregation enhancesradDexpression, and deletion ofcarSabolished this enhancement, suggesting that coaggregation itself serves as a signal sensed by this TCS. These findings provide new insights into the CarR regulon and the regulation of RadD, elucidating the ecological and pathogenic roles ofF. nucleatumin dental plaque formation and disease processes.

IMPORTANCE

Fusobacterium nucleatumis an essential member of oral biofilms, acting as a bridging organism that connects early and late colonizers, thus driving dental plaque formation. Its remarkable ability to aggregate with diverse bacterial partners is central to its ecological success, yet the mechanisms it senses and responds to these interactions remain poorly understood. This study identifies the CarSR two-component system as a direct regulator of RadD, the primary adhesin mediating coaggregation, and reveals its role in sensing coaggregation as a signal. These findings uncover a novel mechanism by whichF. nucleatumdynamically adapts to polymicrobial environments, offering new perspectives on biofilm formation and bacterial communication in complex oral microbial ecosystems.
A mutualistic model bacterium is lethal to non-symbiotic hosts via the type VI secretion system
Authors: Gaddy Keegan E. , Septer Alecia N. , Mruk Karen , Milton Morgan E.
Published: December 2024

What makes a bacterium pathogenic? Since the early days of germ theory, researchers have categorized bacteria as pathogens or non-pathogens, those that cause harm and those that do not, but this binary view is not always accurate.Vibrio fischeriis an exclusive mutualistic symbiont found within the light organs of Hawaiian bobtail squid. This symbiotic interaction requiresV. fischerito utilize a range of behaviors and produce molecules that are often associated with pathogenicity. This juxtaposition of employing “pathogenic” behaviors for a symbiotic relationship led the field to focus on howV. fischeriestablishes a beneficial association with its host. In this study, we observe thatV. fischeriinduces mortality in zebrafish embryos andArtemianauplii. Non-lethal doses ofV. fischerileads to zebrafish growth delays and phenotypes indicative of disease. Our data also provide evidence that the conserved type VI secretion system on chromosome I (T6SS1) plays a role in theV. fischeri-induced mortality of zebrafish embryos andArtemianauplii. These results support the hypothesis that theV. fischeriT6SS1 is involved in eukaryotic cell interactions. Despite its traditional view as a beneficial symbiont, we provide evidence thatV. fischeriis capable of harming aquatic organisms, indicating its potential to be pathogenic toward non-symbiotic hosts.
The hit-and-run of cell wall synthesis: LpoB transiently binds and activates PBP1b through a conserved allosteric switch
Authors: Shlosman Irina , Vettiger Andrea , Bernhardt Thomas G. , Kruse Andrew C. , Loparo Joseph J.
Published: December 2024

The peptidoglycan (PG) cell wall is the primary protective layer of bacteria, making the process of PG synthesis a key antibiotic target. Class A penicillin-binding proteins (aPBPs) are a family of conserved and ubiquitous PG synthases that fortify and repair the PG matrix. In gram-negative bacteria, these enzymes are regulated by outer-membrane tethered lipoproteins. However, the molecular mechanism by which lipoproteins coordinate the spatial recruitment and enzymatic activation of aPBPs remains unclear. Here we use single-molecule FRET and single-particle tracking in E. coli to show that a prototypical lipoprotein activator LpoB triggers site-specific PG synthesis by PBP1b through conformational rearrangements. Once synthesis is initiated, LpoB affinity for PBP1b dramatically decreases and it dissociates from the synthesizing enzyme. Our results suggest that transient allosteric coupling between PBP1b and LpoB directs PG synthesis to areas of low peptidoglycan density, while simultaneously facilitating efficient lipoprotein redistribution to other sites in need of fortification.
MMETHANE: interpretable AI for predicting host status from microbial composition and metabolomics data
Authors: Dawkins Jennifer J. , Gerber Georg K.
Published: December 2024

Metabolite production, consumption, and exchange are intimately involved with host health and disease, as well as being key drivers of host-microbiome interactions. Despite the increasing prevalence of datasets that jointly measure microbiome composition and metabolites, computational tools for linking these data to the status of the host remain limited. To address these limitations, we developed MMETHANE, an open-source software package that implements a purpose-built deep learning model for predicting host status from paired microbial sequencing and metabolomic data. MMETHANE incorporates prior biological knowledge, including phylogenetic and chemical relationships, and is intrinsically interpretable, outputting an English-language set of rules that explains its decisions. Using a compendium of six datasets with paired microbial composition and metabolomics measurements, we showed that MMETHANE always performed at least on par with existing methods, including blackbox machine learning techniques, and outperformed other methods on >80% of the datasets evaluated. We additionally demonstrated through two cases studies analyzing inflammatory bowel disease gut microbiome datasets that MMETHANE uncovers biologically meaningful links between microbes, metabolites, and disease status.
MaAsLin 3: Refining and extending generalized multivariable linear models for meta-omic association discovery
Authors: Nickols William A. , Kuntz Thomas , Shen Jiaxian , Maharjan Sagun , Mallick Himel , Franzosa Eric A. , Thompson Kelsey N. , Nearing Jacob T. , Huttenhower Curtis
Published: December 2024

A key question in microbial community analysis is determining which microbial features are associated with community properties such as environmental or health phenotypes. This statistical task is impeded by characteristics of typical microbial community profiling technologies, including sparsity (which can be either technical or biological) and the compositionality imposed by most nucleotide sequencing approaches. Many models have been proposed that focus on how the relative abundance of a feature (e.g. taxon or pathway) relates to one or more covariates. Few of these, however, simultaneously control false discovery rates, achieve reasonable power, incorporate complex modeling terms such as random effects, and also permit assessment of prevalence (presence/absence) associations and absolute abundance associations (when appropriate measurements are available, e.g. qPCR or spike-ins). Here, we introduce MaAsLin 3 (Microbiome Multivariable Associations with Linear Models), a modeling framework that simultaneously identifies both abundance and prevalence relationships in microbiome studies with modern, potentially complex designs. MaAsLin 3 also newly accounts for compositionality with experimental (spike-ins and total microbial load estimation) or computational techniques, and it expands the space of biological hypotheses that can be tested with inference for new covariate types. On a variety of synthetic and real datasets, MaAsLin 3 outperformed current state-of-the-art differential abundance methods in testing and inferring associations from compositional data. When applied to the Inflammatory Bowel Disease Multi-omics Database, MaAsLin 3 corroborated many previously reported microbial associations with the inflammatory bowel diseases, but notably 77% of associations were with feature prevalence rather than abundance. In summary, MaAsLin 3 enables researchers to identify microbiome associations with higher accuracy and more specific association types, especially in complex datasets with multiple covariates and repeated measures.
Structural basis of transcription-coupled H3K36 trimethylation by Set2 and RNAPII elongation complex in the nucleosome
Authors: Kujirai Tomoya , Ehara Haruhiko , Ito Tomoko , Henmi Masami , Sekine Shun-ichi , Kurumizaka Hitoshi
Published: December 2024

Trimethylation of the histone H3K36 residue (H3K36me3) plays an indispensable role in ensuring transcription fidelity by suppressing undesired cryptic transcription in chromatin. The H3K36me3 modification is accomplished by Set2/SETD2 during transcription elongation by the RNA polymerase II elongation complex (EC). Here we found that the Set2-mediated H3K36me3 deposition occurs primarily on the nucleosome reassembling behind the EC. Cryo-electron microscopy structures of the transcribing EC complexed with Set2 and the reassembled nucleosome revealed that Set2 is anchored by the Spt6 subunit of the EC and captures an H3 N-terminal tail of the nucleosome. Abrogation of the Set2-Spt6 interaction leads to defective transcription-coupled H3K36me3 deposition. These insights elucidate the structure-based mechanism of transcription-coupled H3K36me3 deposition in chromatin.

One-Sentence Summary

Cryo-EM structures of the EC-Set2-nucleosome complex reveal the mechanism of H3 Lys36 trimethylation in chromatin.
Rapid Sterilization of Clinical Apheresis Blood Products using Ultra-High Dose Rate Radiation
Authors: Melemenidis Stavros , Nguyen Khoa D. , Baraceros-Pineda Rosella , Barclay Cherie K. , Bautista Joanne , Lau Hubert , Ashraf M. Ramish , Manjappa Rakesh , Dutt Suparna , Soto Luis Armando , Katila Nikita , Lau Brianna , Visvanathan Vignesh , Yu Amy S. , Surucu Murat , Skinner Lawrie B. , Engleman Edgar G. , Loo Billy W. , Pham Tho D.
Published: December 2024

BACKGROUND AND OBJECTIVES

Apheresis platelets products and plasma are essential for medical interventions, but both still have inherent risks associated with contamination and viral transmission. Platelet products are vulnerable to bacterial contamination due to storage conditions, while plasma requires extensive screening to minimize virus transmission risks. Here we investigate rapid irradiation to sterilizing doses for bacteria and viruses as an innovative pathogen reduction technology.

MATERIALS AND METHODS

We configured a clinical linear accelerator to deliver ultra-high dose rate (6 kGy/min) irradiation to platelet and plasma blood components. Platelet aliquots spiked with 105CFU ofE.coliwere irradiated with 0.1-20 kGy, followed byE.coligrowth and platelet count assays. COVID Convalescent Plasma (CCP) aliquots were irradiated at a virus-sterilizing dose of 25 kGy and subsequently, RBD- specific antibody binding was assessed.

RESULTS

1 kGy irradiation of bacteria-spiked platelets reducedE.coligrowth by 2.7- log without significant change of platelet count, and 5 kGy or higher produced complete growth suppression. The estimated sterilization (6-log bacterial reduction) dose was 2.3 kGy, corresponding to 31% platelet count reduction. A 25 kGy virus sterilizing dose to CCP produced a 9.2% average drop of RBD-specific IgG binding.

CONCLUSION

This study shows proof-of-concept of a novel rapid blood sterilization technique using a clinical linear accelerator. Promising platelet counts and CCP antibody binding were maintained at bacteria and virus sterilizing doses, respectively. This represents a potential point-of-care blood product sterilization solution. If additional studies corroborate these findings, this may be a practical method for ensuring blood products safety.

HIGHLIGHTS

1 kGy irradiation of bacteria-spiked platelets reducedE.coligrowth by 2.7-log without significant change of platelet count, and 5 kGy or higher produced complete growth suppression.

The estimated sterilization (6-log bacterial reduction) dose was 2.3 kGy, corresponding to 31% platelet count reduction.

A 25 kGy virus sterilizing dose to CCP produced a 9.2% average drop of RBD- specific IgG binding.
Salmonellamultimutants enable efficient identification of SPI-2 effector protein function in gut inflammation and systemic colonization
Authors: Newson Joshua P. M. , Gürtler Flavia , Piffaretti Pietro , Meyer Annina , Sintsova Anna , Barthel Manja , Steiger Yves , McHugh Sarah C. , Enz Ursina , Alto Neal M. , Sunagawa Shinichi , Hardt Wolf-Dietrich
Published: December 2024

Salmonellaenterica spp. rely on translocation of effector proteins through the SPI-2 encoded type III secretion system (T3SS) to achieve pathogenesis. More than 30 effectors contribute to manipulation of host cells through diverse mechanisms, but interdependency or redundancy between effectors complicates the discovery of effector phenotypes using single mutant strains. Here, we engineer six mutant strains to be deficient in cohorts of SPI-2 effector proteins, as defined by their reported function. Using various animal models of infection, we show that three principle phenotypes define the functional contribution of the SPI-2 T3SS to infection. Multimutant strains deficient for intracellular replication, for manipulation of host cell defences, or for expression of virulence plasmid effectors all showed strong attenuationin vivo,while mutants representing approximately half of the known effector complement showed phenotypes similar to the wild-type parent strain. By additionally removing the SPI-1 T3SS, we find cohorts of effector proteins that contribute to SPI-2 T3SS-driven enhancement of gut inflammation. Further, we provide an example of how iterative mutation can be used to find a minimal number of effector deletions required for attenuation, and thus establish that the SPI-2 effectors SopD2 and GtgE are critical for the promotion of gut inflammation and mucosal pathology. This strategy provides a powerful toolset for simultaneous parallel screening of all known SPI-2 effectors in a single experimental context, and further facilitates the identification of the responsible effectors, and thereby provides an efficient approach to study how individual effectors contribute to disease.
Lonidamine, a Novel Modulator for the BvgAS System of Bordetella Species
Authors: Ota Natsuko , Nishida Takashi , Standley Daron M. , Sherif Aalaa Alrahman , Iwano Satoshi , Nugraha Dendi Krisna , Ueno Toshiya , Horiguchi Yasuhiko
Published: December 2024

The Gram‐negative bacteria Bordetella pertussis, B. parapertussis, and B. bronchiseptica cause respiratory diseases in various mammals. They share the BvgAS two‐component system, which regulates the phenotypic conversion between the virulent Bvg+ and avirulent Bvg– phases. In the BvgAS system, the sensor kinase BvgS senses environmental cues and transduces a phosphorelay signal to the response regulator BvgA, which leads to the expression of Bvg+ phase‐specific genes, including virulence factor genes. Bacteria grown at 37°C exhibit the Bvg+ phenotype. In contrast, at lower than 26°C or in the presence of modulators, such as MgSO4 and nicotinic acid, the BvgAS system is inactivated, leading bacteria to the avirulent Bvg– phase. Therefore, effective modulators are expected to provide a therapeutic measure for Bordetella infection; however, no such modulators are currently available, and the mechanism by which modulators inactivate the BvgAS system is poorly understood. In the present study, we identified lonidamine as a novel modulator after screening an FDA‐approved drug library using bacterial reporter systems with the Bvg+‐specific and Bvg–‐specific promoters. Lonidamine directly bound to the VFT2 domain of BvgS and inactivated the BvgAS system at concentrations as low as 50 nM, which was at least 2000‐ to 20,000‐fold lower than the effective concentrations of known modulators. Lonidamine significantly reduced the adherence of B. pertussis to cultured cells but unexpectedly exacerbated bacterial colonization of the mouse nasal septum. These results provide insights into the structural requirements for BvgAS modulators and the role of Bvg phenotypes in the establishment of infection.
Recent innovations for improving the techno‐functional properties of plant‐based egg analogs and egg‐mimicking products to promote their industrialization and commercialization
Authors: Khalifa Ibrahim , Li Zhihua , Nawaz Asad , Walayat Noman , Sobhy Remah , Jia Yangyang , Korin Ali , Zou Xiaobo , Maqsood Sajid
Published: December 2024

Recent advancements in plant‐based egg analogs (PBEAs) and plant‐based egg‐mimicking products (PEMP) are driven by growing consumer demand and the need for alternatives to animal‐derived products. This movement is being encouraged by several factors, including a drive toward specifically sustainability, increased allergenicity, and consumer preferences for flexitarian diets. We comprehensively overviewed the current research on the formulation and characterization of PBEAs and PEMP that are vital for diverse food applications. We also highlighted the techno‐functional features of these ingredients and their impact in PBEA and PEMP‐based formulas and evaluated up‐to‐date outcomes that display the availability of economically viable substitutes. However, to efficiently mimic the sensorial and textural features of eggs, further innovation and intensive work are still needed. For instance, challenges persist in achieving desired quality attributes, controlling costs, and scaling‐up the production, which limit broader market adoption of PBEAs and PEMP. Addressing these obstacles through persistent research and development can improve the functionality and acceptance of PBEAs in the food industry, aligning with evolving consumer preferences for plant‐based protein alternative options.
Ricemilks made from pigmented and aromatic rice varieties: Sensory analysis and physiochemical properties
Authors: Ardoin Ryan , Chaffee Olivia , Lea Jeanne , Sookraj Alysza , Boue Stephen , Smith Brennan , Dupre Rebecca , Olson Douglas W. , Broussard William , Priddy Daniel
Published: December 2024

Along with dairy allergic/intolerant and vegetarian/vegan individuals, flexitarian consumers contribute to the market for plant‐based milk alternatives. Niche pigmented (purple and red) and/or aromatic (aromatic brown and white) rice varieties were used to formulate ricemilks, which were compared to a product made from commercial nonaromatic white rice. Descriptive sensory analysis (9 trained panelists) and affective sensory testing (161 consumers) were used to explore perceptions of ricemilks. Additional sample differences were characterized by instrumental color, viscosity, 2‐acetyl‐1‐pyrroline (2‐AP), starch quality, total phenolic content (TPC), oxygen radical absorbance capacity (ORAC), and proximate analyses. Compared to nonaromatic white ricemilk, increased bioactive and antioxidant potential was exhibited for pigmented varieties (TPC ranging 56.6–64.5 µg/mL; ORAC ranging 628–753 µM Trolox equivalents). However, the pigments also resulted in decreased acceptability of product color. For red ricemilk, this was overcome after tasting, resulting in the highest grouping of overall liking scores along with aromatic brown and aromatic white ricemilks (5.6–5.7 on a 9‐point hedonic scale)—all associated with a desirable “popcorn” aromatic from 2‐AP. Highest purchase intent was found for aromatic brown ricemilk (47% overall and 69% among regular dairy and nondairy milk consumers) which was associated with the “dairy” aromatic. Consumers also favored colors and mouthfeel more reminiscent of dairy milk. Ricemilks made from niche pigmented and/or aromatic rices provided unique flavors and outperformed nonaromatic white ricemilk in overall acceptability. Ricemilks made from aromatic whole grain varieties can offer a new health‐promoting functional beverage option to consumers, and a new use of niche rice varieties.
Cold atmospheric plasma inactivation of polyphenol oxidase: Focus on the protective and boosting effect of mono‐ and disaccharides
Authors: Laika Jessica , Sabatucci Annalaura , Sacchetti Giampiero , Di Michele Alessandro , Molina Hernandez Junior Bernardo , Ricci Antonella , Dalla Rosa Marco , Chaves Lopez Clemencia , Neri Lilia
Published: December 2024

Polyphenol oxidase (PPO) is among the most detrimental enzymes in processed plant foods, being responsible for enzymatic browning. To propose a “mild” alternative to traditional enzymatic inactivation methods, this study investigated the effect of cold atmospheric plasma (CAP) on PPO inactivation and highlighted the role of different sugars on both inactivation and structural modification of this enzyme. Different model systems were prepared in phosphate buffer using a purified PPO either alone or added with glucose, fructose, sucrose, and trehalose at different concentrations. CAP treatments (6 KV; 23 KHz; duty cycle 10%) were applied at times ranging from 5 to 30 min. Different spectroscopic analyses were conducted before and after treatments to evaluate the PPO activity and changes in tertiary and secondary structures. CAP induced a significant reduction (p < 0.05) in PPO activity across all systems, ranging from 70% to 94% after 30 min of treatment. Among sugars, fructose enhanced (p < 0.05) the PPO inactivation (+23% on average with respect to the phosphate buffer system), possibly by promoting the loss of secondary structures containing the copper‐binding site of the catalytic pocket. The effect of other sugars on PPO inactivation was strictly dependent on their type and concentration; specifically, disaccharides at the highest concentrations and treatment times showed a protective effect on the structure and functionality of the protein. Thus, the results of this study highlight that sugars can modulate the effectiveness of CAP, offering promising perspectives for optimizing this food processing technology.

Practical Application

Cold atmospheric plasma (CAP) is a promising nonthermal technology for food preservation. In particular, surface dielectric barrier discharge (SDBD) CAP could be applied as an alternative to chemical and thermal treatments to inactivate polyphenol oxidase (PPO), an enzyme responsible for browning reactions and quality loss in most processed fruit and vegetable products. However, as shown by this study, PPO inactivation induced by CAP is affected by sugars. Specifically, fructose can positively influence the inactivation of this enzyme. Therefore, CAP potentially could find main applications for the PPO stabilization of high fructose‐content plants (e.g., pears, apples, bananas, grapes, peppers, and squashes).