bluuebunny/crossref_metadata_2025_split huggingface dataset:
Optimizing genome editing efficiency in
Streptomyces fradiae
via a CRISPR/Cas9n-mediated editing system
Authors: Wu Yuhan , Jin Hui , Yu Qiang , Wei Zihan , Zhu Jiang , Qiu Xiangqi , Luo Gan , Li Junhui , Zhan Yangyang , Cai Dongbo , Chen Shouwen 
Published: January 2025

ABSTRACT

Streptomyces fradiae
is an important bioresource to produce various antibacterial natural products, however, the time-consuming and labor-intensive genome editing toolkits hindered the construction and application of engineered strains, and this study aimed to establish an efficient CRISPR/Cas9n genome editing system in
S. fradiae
. Initially, the CRISPR/Cas9-mediated editing tool was employed to replace those awkward genome editing tools that relied on homologous recombination, while the off-target Cas9 exhibited high toxicity to
S. fradiae
Sf01. Therefore, the nickase mutation D10A, high-fidelity mutations including N497A, R661A, Q695A, and Q926A, and thiostrepton-induced promotor P

tipA

were incorporated into the Cas9 expression cassette, which reduced its toxicity. The deletion of single gene
neoI
and long fragment sequence (13.3 kb) were achieved with efficiencies of 77.8% and 44%, respectively. Additionally, the established tool was applied to facilitate the rapid deletion of
nagB
, replacement of P

frr

with P

ermE

*, and integration of exogenous
vgbS
, with respective efficiencies of 77.8%, 100%, and 67.8%, and all of the above modification strategies benefited neomycin synthesis in
S. fradiae
. Taken together, this research established an efficient CRISPR/Cas9n-mediated genome editing toolkit in
S. fradiae
, paving the way for developing high-performance neomycin-producing strains and facilitating the genetic modification of
Streptomyces
.

IMPORTANCE

This study describes the development and application of a genome editing system mediated by CRISPR/Cas9n in
Streptomyces fradiae
for the first time, which overcomes the challenges associated with genome editing caused by high GC content (74.5%) coupling with complex genome structure, and reduces the negative impact of “off-target effect.” Our work not only provides a facile editing tool for constructing
S. fradiae
strains of high-yield neomycin but also offers the technical guidance for the design of a CRISPR/Cas9n mediated genome editing tool in those creatures with high GC content genomes.
High-pressure continuous culturing: life at the extreme
Authors: Foustoukos Dionysis I. , Houghton Jennifer L. 
Published: January 2025

ABSTRACT

Microorganisms adapted to high hydrostatic pressures at depth in the oceans and within the subsurface of Earth’s crust represent a phylogenetically diverse community thriving under extreme pressure, temperature, and nutrient availability conditions. To better understand the microbial function, physiological responses, and metabolic strategies at
in-situ
conditions requires high-pressure (HP) continuous culturing techniques that, although commonly used in bioengineering and biotechnology applications, remain relatively rare in the study of the Earth’s microbiomes. Here, we focus on recent developments in the design of HP chemostats, with particular emphasis on adaptations for delivery and sampling of dissolved gases. We present protocols for sterilization, inoculation, agitation, and sampling strategies that minimize cell lysis, applicable to a wide range of chemostat designs.
Comparative metagenomics of tropical reef fishes show conserved core gut functions across hosts and diets with diet-related functional gene enrichments
Authors: Wu Derek G. , Harris Cassandra R. , Kalis Katie M. , Bowen Malique , Biddle Jennifer F. , Farag Ibrahim F. 
Published: January 2025

ABSTRACT

Fish gut microbial communities are important for the breakdown and energy harvesting of the host diet. Microbes within the fish gut are selected by environmental and evolutionary factors. To understand how fish gut microbial communities are shaped by diet, three tropical fish species (hawkfish,
Paracirrhites arcatus
; yellow tang,
Zebrasoma flavescens
; and triggerfish,
Rhinecanthus aculeatus
) were fed piscivorous (fish meal pellets), herbivorous (seaweed), and invertivorous (shrimp) diets, respectively. From fecal samples, a total of 43 metagenome assembled genomes (MAGs) were recovered from all fish diet treatments. Each host-diet treatment harbored distinct microbial communities based on taxonomy, with
Proteobacteria
,
Bacteroidota
, and
Firmicutes
being the most represented. Based on their metagenomes, MAGs from all three host-diet treatments demonstrated a baseline ability to degrade proteinaceous, fatty acid, and simple carbohydrate inputs and carry out central carbon metabolism, lactate and formate fermentation, acetogenesis, nitrate respiration, and B vitamin synthesis. The herbivorous yellow tang harbored more functionally diverse MAGs with some complex polysaccharide degradation specialists, while the piscivorous hawkfish’s MAGs were more specialized for the degradation of proteins. The invertivorous triggerfish’s gut MAGs lacked many carbohydrate-degrading capabilities, resulting in them being more specialized and functionally uniform. Across all treatments, several MAGs were able to participate in only individual steps of the degradation of complex polysaccharides, suggestive of microbial community networks that degrade complex inputs.

IMPORTANCE
The benefits of healthy microbiomes for vertebrate hosts include the breakdown of food into more readily usable forms and production of essential vitamins from their host's diet. Compositions of microbial communities in the guts of fish in response to diet have been studied, but there is a lack of a comprehensive understanding of the genome-based metabolic capabilities of specific microbes and how they support their hosts. Therefore, we assembled genomes of several gut microbes collected from the feces of three fish species that were being fed different diets to illustrate how individual microbes can carry out specific steps in the degradation and energy utilization of various food inputs and support their host. We found evidence that fish gut microbial communities share several core functions despite differences in microbial taxonomy. Herbivorous fish harbored a functionally diverse microbial community with plant matter degraders, while the piscivorous and invertivorous fish had microbiomes more specialized in protein degradation.
Exploring animal food microbiomes and resistomes via 16S rRNA gene amplicon sequencing and shotgun metagenomics
Authors: Ge Beilei , McDonald Ryan C. , Yang Qianru , Domesle Kelly J. , Sarria Saul , Li Xin , Hsu Chih-Hao , Jarvis Karen G. , Tadesse Daniel A. 
Published: January 2025

ABSTRACT

As a diverse and complex food matrix, the animal food microbiota and repertoire of antimicrobial resistance (AMR) genes remain to be better understood. In this study, 16S rRNA gene amplicon sequencing and shotgun metagenomics were applied to three types of animal food samples (cattle feed, dry dog food, and poultry feed). ZymoBIOMICS mock microbial community was used for workflow optimization including DNA extraction kits and bead-beating conditions. The four DNA extraction kits (AllPrep PowerViral DNA/RNA Kit, DNeasy Blood & Tissue Kit, DNeasy PowerSoil Kit, and ZymoBIOMICS DNA Miniprep Kit) were compared in animal food as well as the use of peptide nucleic acid blockers for 16S rRNA gene amplicon sequencing. Distinct microbial community profiles were generated, which varied by animal food type and DNA extraction kit. Employing peptide nucleic acid blockers prior to 16S rRNA gene amplicon sequencing was comparable with post-sequencing
in silico
filtering at removing chloroplast and mitochondrial sequences. There was a good agreement between 16S rRNA gene amplicon sequencing and shotgun metagenomics on community profiles in animal feed data sets; however, they differed in taxonomic resolution, with the latter superior at resolving at the species level. Although the overall prevalence of AMR genes was low, resistome analysis of animal feed data sets by shotgun metagenomics revealed 10 AMR gene/protein families, including beta-lactamases, erythromycin/lincomycin/pristinamycin/tylosin, fosfomycin, phenicol, and quinolone. Future expansion of microbiome and resistome profiling in animal food will help better understand the bacterial and AMR gene diversity in these commodities and help guide pathogen control and AMR prevention efforts.

IMPORTANCE
With the growing interest and application of metagenomics in understanding the structure/composition and function of diverse microbial communities along the One Health continuum, this study represents one of the first attempts to employ these advanced sequencing technologies to characterize the microbiota and AMR genes in animal food. We unraveled the effects of DNA extraction kits on sample analysis by 16S rRNA gene amplicon sequencing and showed similar efficacies of two strategies at removing chloroplast and mitochondrial reads. The in-depth analysis using shotgun metagenomics shed light on the community compositions and the presence of an array of AMR genes in animal food. This exploration of microbiomes and resistomes in representative animal food samples by both sequencing approaches laid important groundwork for future metagenomic investigations to gain a better understanding of the baseline/core microbiomes and associated AMR functions in these diverse commodities and help guide pathogen control and AMR prevention efforts.
MASP1 modulation as a novel therapeutic target in severe pediatric pertussis: insights from a multi-omics approach
Authors: Xu Lin , Wang Caiying , Liu Yuhuan , Zhang Yanlan , Li Zhen , Pang Lin 
Published: January 2025

ABSTRACT

Pertussis, a severe infectious disease in children, has become increasingly prominent in recent years. This study aims to investigate the role of the MASP1 protein in severe pertussis in children through multi-omics analysis, providing a theoretical basis for the development of novel therapeutic strategies. The study retrieved macro-genome and 16S rRNA data of pediatric pertussis from public databases to analyze microbial diversity and specific flora abundance, conducting pathway functional enrichment analysis. Differential expression analysis of transcriptome data and Gene Ontology (GO)/Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analysis, combined with machine learning, identified the key gene MASP1. A
Bordetella pertussis
infection model was established using human bronchial epithelial cell line HBE135-E6E7 to validate MASP1 expression changes and investigate its relationship with airway epithelial cell damage by constructing cell lines overexpressing and knocking down MASP1. Finally, the impact of inhibiting MASP1 expression on infection symptoms was evaluated using a mouse pertussis infection model. The results revealed significant differences in microbial diversity and specific flora abundance between healthy children and those with pertussis, with MASP1 significantly upregulated in severe pertussis and its inhibition alleviating infection symptoms. The study highlights the critical role of MASP1 in pertussis, providing a crucial foundation for developing therapeutic strategies targeting MASP1.
TSST-1 promotes colonization of
Staphylococcus aureus
within the vaginal tract by activation of CD8
+
T cells
Authors: Dufresne Karine , Al Kait F. , Craig Heather C. , Coleman Charlotte E. M. , Kasper Katherine J. , Burton Jeremy P. , McCormick John K. 
Published: January 2025

ABSTRACT

Toxic shock syndrome toxin-1 (TSST-1) is a superantigen produced by
Staphylococcus aureus
and is the determinant of menstrual toxic shock syndrome (mTSS); however, the impact of TSST-1 on the vaginal environment beyond mTSS is not understood. Herein, we assessed how TSST-1 affects vaginal colonization by
S. aureus
, host inflammatory responses, and changes in microbial communities within the murine vagina. We demonstrated that TSST-1 induced a CD8
+
T-cell-dependent inflammatory response in 24 h that correlated with
S. aureus
persistence within the vaginal tract. This increase was due to superantigen-dependent T-cell activation that triggered a change in microbial composition within the vaginal tract. Altogether, this study demonstrates that within the vaginal tract, TSST-1 modulates the vaginal microbiota to favor the survival of
S. aureus
in the absence of mTSS.

IMPORTANCE

Toxic shock syndrome toxin-1 (TSST-1) is a superantigen toxin produced from
Staphylococcus aureus
that causes the menstrual form of toxic shock syndrome. This research demonstrates that TSST-1 also has a wider function within the vaginal tract than previously expected. We show that TSST-1, by activating CD8
+
T cells, induces an inflammatory environment that modifies the vaginal microbiota to favor colonization by
S. aureus
. These are important findings as
S. aureus
can colonize the human vaginal tract efficiently and subsequently trigger dysbiosis within the microbial communities leading to several adverse outcomes such as decreased fertility, increased risks for sexually transmitted diseases, and issues related to pregnancy and birth.
Anidulafungin is a useful surrogate marker for predicting
in vitro
susceptibility to rezafungin among five
Candida
species using CLSI methods and interpretive criteria
Authors: Winkler Marisa L. , Deshpande Lalitagauri , Kimbrough John H. , Karr Maura , Rhomberg Paul , Klauer Abby L. , Castanheira Mariana 
Published: January 2025

ABSTRACT

This study addresses the use of other echinocandins as surrogate markers to predict the susceptibility of rezafungin against the six most common
Candida
spp. The Clinical Laboratory Standards Institute (CLSI) reference broth microdilution method was performed to test 5,720 clinical isolates of six different
Candida
species. Species-specific interpretative criteria by CLSI breakpoints or epidemiological cutoff values were applied. Essential agreement was 100% within two doubling dilutions for all species and comparisons. The categorical agreement of rezafungin using anidulafungin against all
Candida
spp. was 97.6% (2.9% very major errors [VMEs], 0.2% major errors [MEs], and 2.2% minor errors [miEs]); for caspofungin, it was 99.6% (11.4% VME, 0.09% ME, and 0.19% miE); and for micafungin, it was 99.6% (14.3% VME, 0.15% ME, and 0.17% miE). There were species-specific differences that led to unacceptably high VME for
Candida dubliniensis
with all agents and for
Candida parapsilosis
when caspofungin or micafungin but not anidulafungin was used as the comparator. Genetic analysis showed rezafungin nonsusceptibility correlated well with FKS hotspot mutations. The best-performing surrogate was anidulafungin, which can be used to predict rezafungin susceptible or nonsusceptible in
Candida albicans, Candida glabrata, Candida parapsilosis, Candida tropicalis,
and
Candida krusei
with low error rates and ≥90% essential and categorical agreement. Micafungin or caspofungin can also be used as a surrogate marker for predicting rezafungin susceptible or nonsusceptible in
C. albicans
,
C. glabrata
,
C. tropicalis
, and
C. krusei
. No surrogate performs appropriately to determine rezafungin susceptibility for
C. dubliniensis
.
High astrovirus diversity in an endemic bat species suggests multiple spillovers from synanthropic rodents and birds
Authors: Leong Rachel , Hoarau Axel O. G. , Carcauzon Victoria , Köster Marie , Dietrich Muriel , Tortosa Pablo , Lebarbenchon Camille 
Published: January 2025

ABSTRACT

Beyond the role of bats as natural host reservoirs of infectious agents, the impact of viral spillover from other animal species to bats has been neglected. Given the limited virus-host specificity of astroviruses (AstVs) and their propensity for cross-species transmission, we hypothesized that AstVs could be transmitted within animal communities (rodents, birds, and bats) and that native endemic bats may be exposed to viruses hosted by other species. We investigated the presence of AstV RNA in 3,796 biological samples collected in Reunion Island from
Mormopterus francoismoutoui
(
N
= 3421), an endemic free-tailed bat species, and also from small terrestrial mammals and birds:
Rattus rattus
(
N
= 146),
Rattus norvegicus
(
N
= 74),
Mus musculus domesticus
(
N
= 36),
Suncus murinus
(
N
= 99), and
Columba livia
(
N
= 20). We found significant differences in AstV prevalence between species (ranging from 1% ± 1.97% in
S. murinus
to 75% ± 18.9% in
C. livia
) as well as variation of pairwise identity of AstV sequences among host species, with a high diversity in
M. francoismoutoui
(mean pairwise identity: 53.2% ± 11.2%). Phylogenetic analyses further revealed that AstVs detected in bats did not cluster in a single clade and were genetically related to AstVs found in birds, rodents, pigs, dogs, cats, and reptiles. Our study suggests that bats may be exposed to viruses associated to synanthropic and alien species. Cascade effects of virus spillover toward endemic and endangered bat species need to be fully assessed.

IMPORTANCE
Epidemiological consequences of cross-species transmission of zoonotic viruses are mostly considered from a health and economic perspective. Virus spillovers resulting from human-introduced species are much less considered, although they may have major consequences on the conservation of endemic and endangered bat species, in particular in an island context. Based on astrovirus detection and sequencing in an endemic bat species and five non-native species on a tropical island, we identified multiple and repeated viral introductions from synanthropic rodents and birds to bats, rather than the opposite. Such findings call for more investigations in these isolated and vulnerable ecosystems to better understand and mitigate the risks associated with pathogen spillovers.
Retreatment of patients with chronic hepatitis C, subtype 3a, and cirrhosis, who previously failed a regimen containing second-generation NS5A inhibitors with sofosbuvir + glecaprevir/pibrentasvir and ribavirin for 16–24 weeks
Authors: Fedorchenko Sergii V. , Klimenko Zhanna , Martynovich Tatiana , Solianyk Iryna , Suprunenko Tatiana 
Published: January 2025

ABSTRACT

The outcomes of retreatment patients infected with hepatitis C virus genotype 3, cirrhosis, with velpatasvir may be affected by treatment failure with velpatasvir. The efficacy of SOF+GLE/PIB+RIB 16–24 weeks of treatment has been shown. The presence of NS5A resistance-associated substitution mutations, including Y93H, and the number and regimens of the past failed therapy do not influence the likelihood of achieving sustained virological response. When velpatasvir treatment fails, pibrentasvir should be used as the first choice for retreatment.
On the analytic generalization of particle deflection in the weak field regime and shadow size in light of EHT constraints for Schwarzschild-like black hole solutions
Authors: Pantig Reggie C. 
Published: January 2025

Abstract
In this paper, an analytic generalization of the weak field deflection angle (WDA) is derived by utilizing the current non-asymptotically flat generalization of the Gauss–Bonnet theorem. The derived formula is valid for any Schwarzschild-like spacetime, which deviates from the classical Schwarzschild case through some constant parameters. This work provided four examples, including Schwarzschild-like solutions in the context of Bumblebee gravity theory and the Kalb–Ramond framework, as well as one example from a black hole surrounded by soliton dark matter. These examples explore distinct mechanisms of Lorentz symmetry breaking, with results that are either new or in agreement with existing literature. The WDA formula provided a simple calculation, where approximations based on some conditions can be done directly on it, skipping the preliminary steps. For the shadow size analysis, it is shown how it depends solely on the parameter associated with the metric coefficient in the time coordinate. A general formula for the constrained parameter is also derived based on the Event Horizon Collaboration (EHT) observational results. Finally, the work realized further possible generalizations on other black hole models, such as RN-like, dS/AdS-like black hole solutions, and even black hole solutions in higher dimensions.
A Prognostic Riskscore Model Related to Helicobacter pylori Infection in Stomach Adenocarcinoma
Authors: Peng Jing , Yan Qi , Pei Wennan , Jiang Yi , Zhou Li , Li Ruoqing 
Published: January 2025

Background: Helicobacter pylori (HP) is associated with the development of various stomach diseases, one of the major risk factors for stomach adenocarcinoma (STAD).Methods: The HP infection score between tumor and normal groups was compared by single‐sample gene set enrichment analysis (ssGSEA). The key modules related to HP infection were identified by weighted gene coexpression network analysis (WGCNA), and functional enrichment analysis was conducted on these module genes. Further, the limma package was used to screen the differentially expressed genes (DEGs) between HP‐positive and HP‐negative STAD. The prognostic genes were obtained to construct the riskscore model, and the performance of the model was validated. The correlation between riskscore and tumor immune microenvironment (TIME) was analyzed by Spearman’s method. The single‐cell atlas of HP‐positive STAD was delineated. The mRNA expression levels of the prognostic genes were verified using STAD cells, and the migration and invasion capacities of STAD cells were evaluated by using the wound healing assay and transwell assay.Results: The HP infection score in the tumor group was significantly higher than that in the normal group. The purple and royal blue modules showed higher correlation with HP infection in STAD, and these module genes were enriched in the immune‐related pathway. Further, five prognostic genes (CTLA4, CPVL, EMB, CXCR4, and FAM241A) were screened from the HP infection–related DEGs, which were utilized for establishing the riskscore model, with good robustness. Riskscore exhibited strong correlation with TIME in STAD. Single‐cell atlas of HP‐positive STAD revealed that CXCR4 is highly expressed in Epithelial Cell 1, Epithelial Cell 2, and parietal cells of the tumor group. CPVL, EMB, CTLA4, FAM241A, and CXCR4 showed high expression in STAD cells, and the silencing of CPVL could suppress the migration and invasion of STAD cells.Conclusion: This study established a riskscore model based on HP infection–related genes, which could provide reference for prognostic prediction and treatment targets of STAD.