bluuebunny/crossref_metadata_2025_split huggingface dataset:
Comparative metagenomic analysis of biosynthetic diversity across sponge microbiomes highlights metabolic novelty, conservation and diversification
Authors: Loureiro Catarina , Galani Anastasia , Gavriilidou Asimenia , de Mares Maryam Chaib , van der Oost John , Medema Marnix H. , Sipkema Detmer 
Published: April 2022

AbstractMarine sponges and their microbial symbiotic communities are rich sources of diverse natural products (NPs) that often display biological activity, yet little is known about their global distribution landscape and the symbionts that produce them. As the majority of sponge symbionts remain uncultured, it is a challenge to characterize their NP biosynthetic pathways, assess their prevalence within the holobiont and measure their diversity across sponge taxa and environments. Here, we explore the microbial biosynthetic landscapes of three sponge species from the Atlantic Ocean and the Mediterranean Sea. This dataset reveals striking novelty in its encoded biosynthetic potential, with less than 1% of the recovered gene cluster families (GCF) showing similarity to any characterized biosynthetic gene cluster (BGC). When zooming in on the microbial communities of each sponge, we observed higher variability of both secondary metabolic and taxonomic profiles between sponge species than within species. Nonetheless, we also identified conservation of GCFs, with 20% of sponge GCFs being shared between at least two sponge species, and a true GCF core comprised of 6% of GCFs shared across all species. Within this functional core, we identified a set of widespread and diverse GCFs encoding nonribosomal peptide synthetases (NRPS) that are potentially involved in the production of diversified ether lipids, as well as GCFs putatively encoding the production of highly modified proteusins. The present work contributes to the small, yet growing body of data characterizing NP landscapes of marine sponge symbionts, and to the cryptic biosynthetic potential contained in this environmental niche.
Surface microbiota of Mediterranean loggerhead sea turtles unravelled by 16S and 18S amplicon sequencing
Authors: Kanjer Lucija , Filek Klara , Mucko Maja , Majewska Roksana , Gračan Romana , Trotta Adriana , Panagopoulou Aliki , Corrente Marialaura , Bello Antonio Di , Bosak Sunčica 
Published: April 2022

AbstractLoggerhead sea turtle is considered a keystone species with major ecological role in Mediterranean marine environment. As with other wild reptiles, their outer microbiome is rarely studied. Although there are several studies on sea turtle’s macro-epibionts and endo-microbiota, there has been little research on epibiotic microbiota associated with sea turtles’ skin and carapace. Therefore, we provide the first identification of combined epibiotic eukaryotic and prokaryotic microbiota on Mediterranean loggerhead sea turtles. In this study, we sampled skin and carapace of 26 loggerheads from the Mediterranean Sea during 2018 and 2019. To investigate the overall microbial diversity and composition, amplicon sequencing of 16S and 18S rRNA genes was performed. We found that the Mediterranean loggerhead sea turtle epibiotic microbiota is a reservoir of a vast variety of microbial species. Microbial communities between samples mostly varied by different locations and seas, while within prokaryotic communities’ significant difference was observed between sampled body sites (carapace vs. skin). In terms of relative abundance, Proteobacteria and Bacteroidota were the most represented phyla within prokaryotes, while Alveolata and Stramenopiles thrived among eukaryotes. This study, besides providing a first survey of microbial eukaryotes on loggerheads via metabarcoding, identifies fine differences within both prokaryotic and eukaryotic microbial communities that seem to reflect the host anatomy and habitat. Multi-domain epi-microbiome surveys provide additional layers of information that are complementary with previous morphological studies and enable better understanding of the biology and ecology of these vulnerable marine reptiles.
Recall of pre-existing cross-reactive B cell memory following Omicron breakthrough infection
Authors: Kaku Chengzi I. , Bergeron Alan J. , Ahlm Clas , Normark Johan , Sakharkar Mrunal , Forsell Mattias N. E. , Walker Laura M. 
Published: April 2022

AbstractUnderstanding immune responses following SARS-CoV-2 breakthrough infection will facilitate the development of next-generation vaccines. Here, we profiled spike (S)-specific B cell responses following Omicron/BA.1 infection in mRNA-vaccinated donors. The acute antibody response was characterized by high levels of somatic hypermutation (SHM) and a bias toward recognition of ancestral SARS-CoV-2 strains, suggesting the early activation of vaccine-induced memory B cells (MBCs). BA.1 breakthrough infection induced a shift in B cell immunodominance hierarchy from the S2 subunit toward the receptor binding domain (RBD). A large proportion of RBD-directed neutralizing antibodies isolated from BA.1 breakthrough infection donors displayed convergent sequence features and broadly recognized SARS-CoV-2 variants of concern (VOCs). Together, these findings provide fundamental insights into the role of pre-existing immunity in shaping the B cell response to heterologous SARS-CoV-2 variant exposure.One sentence summaryBA.1 breakthrough infection activates pre-existing memory B cells with broad activity against SARS-CoV-2 variants.
Bacterial threat assessment of bacteriophage infection is mediated by intracellular polyamine accumulation and Gac/Rsm signaling
Authors: de Mattos Camilla D. , Nemudryi Artem A. , Faith Dominick , Bublitz DeAnna C. , Hammond Lauren , Kinnersley Margie A. , Schwartzkopf Caleb M. , Robinson Autumn J. , Joyce Alex , Michaels Lia A. , Brzozowski Robert S. , Coluccio Alison , Xing Denghui David , Uchiyama Jumpei , Jennings Laura K. , Eswara Prahathees , Wiedenheft Blake , Secor Patrick R. 
Published: April 2022

AbstractWhen eukaryotic cells are killed by pathogenic microorganisms, damage-associated and pathogen-associated signals are generated that alert other cells of nearby danger. Bacteria can detect the death of their kin; however, how bacteria make threat assessments of cellular injury is largely unexplored. Here we show that polyamines released by lysed bacteria serve as damage-associated molecules in Pseudomonas aeruginosa. In response to exogenous polyamines, Gac/Rsm and cyclic-di-GMP signaling is activated and intracellular polyamine levels increase. In the absence of a threat, polyamines are catabolized, and intracellular polyamines return to basal levels, but cells infected by bacteriophage increase and maintain intracellular polyamine levels, which inhibits phage replication. Phage species not inhibited by polyamines did not trigger polyamine accumulation by P. aeruginosa, suggesting polyamine accumulation and metabolism are targets in the phage-host arms-race. Our results suggest that like eukaryotic cells, bacteria can differentiate damage-associated and pathogen-associated signals to make threat assessments of cellular injury.Abstract Figure
Allopeptimicins: unique antibacterial metabolites generated by hybrid PKS-NRPS, with original self-defense mechanism in Actinoallomurus
Authors: Iorio Marianna , Gentile Andrea , Brunati Cristina , Tocchetti Arianna , Landini Paolo , Maffioli Sonia Ilaria , Donadio Stefano , Sosio Margherita 
Published: April 2022

AbstractIn the search for structurally novel metabolites with antibacterial activity, innovative approaches must be implemented to increase the probability of discovering novel chemistry from microbial sources. Here we report on the application of metabolomic tools to the genus Actinoallomurus, a poorly explored member of the Actinobacteria. From examining extracts derived from 88 isolates belonging to this genus, we identified a family of cyclodepsipeptides acylated with a C20 polyketide chain, which we named allopeptimicins. These molecules possess unusual structural features, including several double bonds in the amino-polyketide chain and four non-proteinogenic amino acids in the octapeptide. Remarkably, allopeptimicins are produced as a complex of active and inactive congeners, the latter carrying a sulfate group on the polyketide amine. This modification is also a mechanism of self-protection in the producer strain. The structural uniqueness of allopeptimicins is reflected in a biosynthetic gene cluster showing a mosaic structure, with dedicated gene cassettes devoted to formation of specialized precursors and modular assembly lines related to those from different pathways.
A single vesicle fluorescence-bleaching assay for multi-parameter analysis of proteoliposomes by total internal reflection fluorescence microscopy
Authors: Veit Sarina , Paweletz Laura Charlotte , Bohr Sören S.-R. , Menon Anant K. , Hatzakis Nikos S , Pomorski Thomas Günther 
Published: April 2022

AbstractReconstitution of membrane proteins into model membranes is an essential approach for their functional analysis under chemically defined conditions. Established model-membrane systems used in ensemble average measurements are limited by sample heterogeneity and insufficient knowledge of lipid and protein content at the single vesicle level, which limits quantitative analysis of vesicle properties and prevents their correlation with protein activity. Here, we describe a versatile total internal reflection fluorescence microscopy-based bleaching protocol that permits parallel analyses of multiple parameters (physical size, tightness, unilamellarity, membrane protein content and orientation) of individual proteoliposomes prepared with fluorescently tagged membrane proteins and lipid markers. The approach makes use of commercially available fluorophores including the commonly used nitrobenzoxadiazole (NBD) dye and may be applied to deduce functional molecular characteristics of many types of reconstituted fluorescently tagged membrane proteins.
Head-to-head comparison of CCN4, DNMT3A, PTPN11, and SPARC as suppressors of anti-tumor immunity
Authors: Pirkey Anika C. , Deng Wentao , Norman Danielle , Razazan Atefeh , Klinke David J. 
Published: April 2022

BackgroundEffective communication between innate and adaptive immunity is essential for mounting an effective antitumor immune response. Emergent cancer cells likely secrete factors that inhibit this communication. To identify such factors, we applied an in vitro workflow that coupled a functional assay with proteomics to a syngeneic mouse model of melanoma known to be resistant to immunotherapies. Collectively, these in vitro results suggested CCN4, DNMT3A, PTPN11, and SPARC as secreted factors that potentially mediate immunosuppression and could potentially become easily accessible targets for novel immunotherapies. The objective of this study was to test for consistent clinical correlates in existing human data and to verify in vivo whether knocking out tumor cell production of these secreted factors improved immune-mediated control of tumor growth.ResultsAnalyses of available human data indicate that high CCN4 expression is associated with reduced survival in primary melanoma patients. High DNMT3A, PTPN11, and SPARC expression showed no association with overall survival. In addition, scRNAseq data from patients with melanoma confirmed CCN4 is expressed by the tumor cells, as opposed to other cells in the tumor microenvironment. An experimental system was created using a CRISPR/Cas9 approach to generate knock out cell lines for each of the four genes of interest using the B16F0 mouse melanoma cell line. In a immunocompetent C57BL/6, DNMT3A, PTPN11, and SPARC knockouts had no effect on overall survival compared to mice challenged with wildtype B16F0 cells while the CCN4 knockout significantly increased survival. This increase in overall survival was lost when the CCN4 knockout cells were injected into severely immunocompromised NSG hosts, indicating that CCN4’s effect on the tumor microenvironment is immune mediated. A kinetic analysis leveraging a Markov Chain Monte Carlo approach quantified the various knockouts’ effect on cells’ intrinsic growth rate. The analysis shows CCN4 is the only knockout tested that decreased the net proliferation rate in immunocompetent mice compared to wildtype cells.ConclusionsThe results suggest that CCN4 is a mediator of immunosuppression in the melanoma tumor microenvironment and a potential collateral immunotherapy target.
CryoGrid-PIXUL-RNA: High throughput RNA isolation platform for tissue transcript analysis
Authors: Schactler Scott A. , Scheuerman Stephen J. , Lius Andrea , Altemeier William A. , An Dowon , Matula Thomas J. , Mikula Michal , Kulecka Maria , Denisenko Oleg , Mar Daniel , Bomsztyk Karol 
Published: April 2022

ABSTRACTDisease molecular complexity requires high throughput workflows to map disease pathways through analysis of vast tissue repositories. Great progress has been made in life sciences analytical technologies. To match the high throughput of these advanced analytical platforms, we have previously developed a multipurpose microplate sonicator, PIXUL, that can be used in multiple workflows to extract analytes from cultured cells and tissue fragments for various downstream molecular assays. And yet, the sample preparation devices, such as PIXUL, along with the downstream analytical capabilities have not been fully exploited to interrogate tissues because storing and sampling of such specimens remain, in comparison, inefficient. To mitigate this bottleneck, we have developed a low-cost user-friendly system, the CryoGrid, that consists of CryoBlock, thermometer/thermocouple, and QR coded CryoTrays to freeze and store frozen tissue fragments, and hand-held CryoCore tool for tissue sampling supported by iPad and Google apps to display tissues, direct coring and share metadata.RNA is one of the most studied analytes. There is a decades-long history of developing methods to isolate and analyze RNA. Still, the throughput of sampling and RNA extraction from tissues has not matched that of the high throughput transcriptome analytical platforms. To address this need, we have integrated the CryoGrid system with PIXUL-based methods to isolate RNA for gene-specific qPCR and genome-wide transcript analyses. TRIzol is commonly used to isolate RNA but it is labor-intensive, hazardous, requires fume-hoods, and is an expensive reagent. We developed a PIXUL-based TRIzol-free RNA isolation fast protocol that uses a buffer containing proteinase K (PK). Virtually every disease (and often therapeutic agents’ toxicity) is a systemic syndrome but often only one organ is examined. CryoGrid-PIXUL, integrated with either TRIzol or PK buffer RNA isolation protocols, yielded similar RNA profiles in a multiorgan (brain, heart, kidney and liver) mouse model of sepsis. Thus, RNA isolation using the CryoGrid-PIXUL system combined with the PK buffer offers an inexpensive user-friendly workflow to study transcriptional responses in tissues in health and disease as well as in therapeutic interventions.
Efflux-linked Accelerated Evolution of Antibiotic Resistance at a Population Edge
Authors: Bhattacharyya Souvik , Bhattacharyya Madhumita , Pfannenstiel Dylan M. , Nandi Anjan K. , Hwang YuneSahng , Ho Khang , Harshey Rasika M. 
Published: April 2022

SUMMARYEfflux is a common mechanism of resistance to antibiotics. We show that efflux itself promotes accumulation of antibiotic-resistance mutations (ARM). This phenomenon was initially discovered in a bacterial swarm where the linked phenotypes of high efflux and high mutation frequencies spatially segregated to the swarm edge, driven there by motility. We uncovered and validated a global regulatory network that connects high efflux to downregulation of specific DNA repair pathways even in non-swarming states. The efflux-DNA repair link was corroborated in a clinical ‘resistome’ database: genomes with mutations that increase efflux exhibit a significant increase in ARMs. Accordingly, efflux inhibitors decreased evolvability to antibiotic resistance. Swarms also revealed how bacterial populations serve as reservoir of ARMs even in absence of antibiotic selection pressure. High efflux at the edge births mutants that, despite compromised fitness, survive there because of reduced competition. This finding is relevant to biofilms where efflux activity is high.
The temporal transcriptomic signature of cartilage formation
Authors: Takács Roland Á. , Vágó Judit , Póliska Szilárd , Pushparaj Peter N. , Ducza László , Kovács Patrik , Jin Eun-Jung , Barrett-Jolley Richard , Zákány Róza , Matta Csaba 
Published: April 2022

ABSTRACTChondrogenesis is a multistep process, in which cartilage progenitor cells generate a tissue with distinct structural and functional properties. Although several approaches to cartilage regeneration rely on the differentiation of implanted progenitor cells, the temporal transcriptomic landscape ofin vitrochondrogenesis in different models has not been reported. Using RNA sequencing, we examined differences in gene expression patterns during cartilage formation in micromass cultures of embryonic limb bud-derived progenitors. Principal component and trajectory analyses revealed a progressively different and distinct transcriptome during chondrogenesis. Differentially expressed genes (DEGs), based on pairwise comparisons of samples from consecutive days were classified into clusters and analysed. We confirmed the involvement of the top DEGs in chondrogenic differentiation using pathway analysis and identified several chondrogenesis-associated transcription factors and collagen subtypes that were not previously linked to cartilage formation. Transient gene silencing ofATOH8orEBF1on day 0 attenuated chondrogenesis by deregulating the expression of key osteochondrogenic marker genes in micromass cultures. These results provide detailed insight into the molecular mechanism of chondrogenesis in primary micromass cultures and present a comprehensive dataset of the temporal transcriptomic landscape of chondrogenesis, which may serve as a platform for new molecular approaches in cartilage tissue engineering.GRAPHICAL ABSTRACTThis study provides insights into the mechanisms of cartilage formation. Gene expression patterns during cartilage formation in micromass cultures were analysed using RNA sequencing. SilencingATOH8orEBF1attenuates chondrogenesis.Key points•We examined the global gene expression patterns during in vitro chondrogenesis.•Using WGCNA, we created a module of genes with patterns similar to those of SOX9, ACAN, and COL2A1.•We identified ATOH8 and EBF1 transcription factors with a yet unexplored role in chondrogenesis.
Social media communication with intensified pandemic fears: evaluating the relative impact of user- and firm-generated content on brand loyalty
Authors: Heng Wei Lee , Chuan Huat Ong , Arumugam Prakash V. 
Published: April 2022

PurposeThe purpose of this study is to analyse user-generated content and firm-generated content on perceived quality and brand trust, and eventually how it impacted brand loyalty with pandemic fear as the moderator.Design/methodology/approachThis study employed an online survey questionnaire method in the Facebook online shopping groups to collect the data. The filter question technique was adopted to verify the respondent's validity. A total of 434 samples was collected using purposive sampling. The data were then analysed using SmartPLS 3.0.FindingsThe analysis showed that firm-generated content appeared to have a stronger positive relationship on perceived quality and brand trust than on user-generated content. Brand trust and perceived quality are found to influence brand loyalty positively. However, pandemic fears only moderate the relationship between firm-generated content and brand trust and perceived quality. This study revealed that the main components in social media communication do not influence perceived quality and brand trust to the same extent.Originality/valueThis study contributes to the knowledge of social media communication during the pandemic period that has not been studied empirically in the Malaysian context. The main components in social media communication were delineated and the impact of pandemic fears on how they would possibly affect the established relationships in the literature were examined. This study enables the researchers and practitioners to take a closer look at how the pandemic crisis could provide a shift on what has been understood so far.
Understanding the role of image, quality and price for developing prestigious mass brands
Authors: Singh Balgopal 
Published: April 2022

PurposeThis research article aims to understand the role of brand image, service quality and price (charge) in revitalising functional mass brands into prestigious mass brands.Design/methodology/approachThe empirical research framework was developed by synthesising the past literature on masstige marketing and brand extension. Data was collected using a survey questionnaire from 396 respondents availing M-Wallet. Structural equation modelling was used to validate the brand revitalization attributes; further, the binary logistic regression model examined the effect of revitalization attributes on the chance of increasing customer's perception of masstige.FindingsThe exploratory study suggested brand image, service quality and value for money pricing as essential attributes to revitalize mass brands into masstige brands; furthermore, path analysis validated the positive effects of these attributes on the perception of masstige. The proposed binary logistic regression model suggested brand image as sensitive attributes, increasing the odds ratio by 9.39 times in favour of perceiving brand as masstige followed by the perceived service quality that is 5.87 times. The prediction capability of the proposed binary logistic regression model is found to be 96%.Practical implicationsThe methodology of this study provides the basis for future researchers to advance research on masstige. This study will assist the marketers of mass brands to make better marketing decisions related to how masstige image can be sustained or a new or less known brand can be revitalized into a prestigious brand.Originality/valueThis study is the first to provide empirical evidence of how the mass brand can be revitalised as masstige brands by considering image, quality and price attributes.