Ribosomal RNA Research Articles

Cold atmospheric plasma and skin wound healing: influence on microbial diversity and composition

BackgroundSkin wound healing presents a complex challenge, often compounded by the risk of infection. Cold atmospheric plasma (CAP) emerged as a novel therapeutic for reducing bacterial load and expediting wound healing. However, its effect on the wound microbiome remained unclear. This study aimed to characterize the microbiome of different types of wounds and determine whether CAP influenced microbial diversity.MethodsTwenty-five patients (ten with acute, fifteen with chronic skin wounds) and ten healthy controls were enrolled. CAP was tailored to individual clinical conditions. Skin samples were collected before and after CAP, and microbiota composition was determined by 16 S ribosomal RNA sequencing.ResultsMicrobial communities differed between acute and chronic groups. CAP could accelerate wound healing. However, it did not change microbial α and β-diversity in acute wounds. In chronic wounds, α-diversity indices, including the chao and ACE, were significantly increased, and a significant clustering was observed in post-CAP group. In addition, CAP led to higher abundance of Staphylococcus, lower levels of Proteobacteria and Pseudomonas in chronic wounds.ConclusionsThis study provided novel insights into the impact of CAP on skin wound microbiota. Further research was required to ascertain causality between microbiota and CAP and to develop personalized CAP treatment strategies.

Characterization of the Complete Mitochondrial Genome of Angulyagra polyzonata and Its Phylogenetic Status in Viviparidae.

Angulyagra polyzonata is an economically important mollusk in China, but detailed insights into its mitochondrial genome remain scarce. In this study, we sequenced and comprehensively analyzed the structural features and selection pressures of the A. polyzonata mitochondrial genome. The maximum likelihood method and Bayesian phylogenetic inference method were used to construct a phylogenetic tree of A. polyzonata with 21 other species, including gastropods and bivalves. The full-length mitochondrial genome of 17,379 bp was found to include 22 transfer RNA genes, 2 ribosomal RNA genes, and 13 protein-coding genes, exhibiting similarity to the composition and arrangement of mitochondrial genes in other gastropod species. Notably, the Ka/Ks ratios of mitochondrial protein-coding genes (nad5, cox3, nad3, nad2, cox1, cox2, atp8, atp6, nadl, nad6, cob, nad4l, and nad4) were <1, which indicates that the snail genes of the three genera of the family may have been subjected to strong natural selection pressure during the evolutionary process, so that the number of synonymous mutations (ks) in genes was much more than that of nonsynonymous mutations (ka). Comparative genomic analysis indicated that, apart from the absence of trnW and trnQ, the gene composition of A. polyzonata shares a high degree of homology with other members of the conical snail family. Phylogenetic analysis demonstrated that the selected species could be classified into two primary clades in which A. polyzonata clustered with the Viviparidae family. This study bridges the knowledge gap regarding the mitochondrial genome of A. polyzonata and offers valuable insights into the systematic relationships within the Viviparidae family.

Acupuncture attenuates experimental autoimmune thyroiditis by modulating intestinal microbiota and palmitic acid metabolism.

Autoimmune thyroiditis (AIT) is a common and chronic autoimmune disease. Recent evidence indicates that serum metabolites anddysbiosis of the intestinal microbiota are associated with AIT, with theunderlying mechanism involving the apoptosis of thyroid follicular epithelial cells. Acupuncture, a traditional Chinese therapy, has demonstrated potential regulatory effects on various immune-related diseases. Clinical symptoms in AITpatients have shown improvement following acupuncture intervention. However, the mechanism underlying its therapeutic effects remain poorly understood. In this study, we investigated the mechanisms of acupuncture (Acu) treatment in rats with established experimental autoimmune thyroiditis (EAT) and evaluated the relationship between microbiota and serum metabolites after Acu treatment. After six weeks of acupuncture and Selenium yeast intervention (used as a positive control), enzyme-linked immunosorbent assay was used to employed to assess the expressions of serum thyroid function and inflammatory markers. Pathological changes in the thyroid gland were observed using hematoxylin-eosin staining and electron microscopy. Thyroid apoptosis was evaluated through TUNEL staining, immunohistochemistry and Western blot analysis. Additionally, changes in intestinal microbiota and serum metabolic profile were analyzed by 16S ribosomal RNA (16S rRNA) sequencing and LC-MS metabolomics, aiming to identify potential therapeutic targets for acupuncture intervention in AIT. The results revealed that Acu could effectively improve thyroid dysfunction and histopathological changes in EAT rats. Following Acu treatment, the content of B-cell lymphoma-2 (Bcl-2) increased, while the levels of Bax and the proportion of cleaved caspase-3 in thyroid tissue decreased. This may be associated with the amelioration of intestinal microbiota dysbiosis and metabolic disorders in EAT rats. Acu mitigated EAT-induced metabolic disorders by regulating the metabolism of palmitoleic acid, and adjusted intestinal microbiota dysbiosis by increasing the abundance of Prevotella. Furthermore, the microbiota (Prevotella) and metabolites (Cyclohexanecarboxylic acid, Tetradecanedioic acid) may serve as co-targets for both Acu and Selenium yeast treatment in EAT. Acu improves the apoptosis of thyroid follicular epithelial cells in rats in EAT model, and its mechanism may be related to intestinal microbiota and metabolism.

Taxonomic diversity of the intestinal microbiome landscape and its clinical significance in recurrent pregnancy loss

Aim: to study taxonomic diversity of the intestinal microbiome landscape in relation to neuro-immune-humoral biomarkers in patients with recurrent pregnancy loss (RPL).Materials and Methods. A cross-sectional comparative study was conducted by enrolling 55 pregnant women with history of RPL (main group) and 60 women with physiological pregnancy (control group). All women underwent serum tumor necrosis factor-alpha (TNF-α), interleukin (IL) IL-17, cortisol and melatonin levels assessment using enzyme-linked immunosorbent assay. The taxonomic composition of the intestinal microbiota at the birth level was examined using 16S ribosomal RNA gene sequencing. The Chao1, Sobs, and ACE (Abundance Coverage Estimator) indices were used to assess α-diversity of microbial community.Results. It was found that α-diversity of the bacterial community in patients with RPL was significantly decreased assessed by Chao1 index (p = 0.014). A significant decline in prevalence of the genera Bifidobacterium (p &lt; 0.001), Lаchnоsріra (p = 0.032), Roseburia (p = 0.003), Соррососcus (p = 0.012) was established along with rise in Ruminососсus (p &lt; 0.001) and Кlebsiеllа (p = 0.002) in women with RPL. Moreover, there were observed several significant relations: а positive correlation between abundance of Ruminococcus bacteria and TNF-α level (r = 0.49; p = 0.003), a negative correlation between abundance of Bifidobacterium and IL-17 (r = –0.54; p = 0.001), abundance of Lachnospira and cortisol level (r = –0.46; p = 0.002), as well as abundance of Coprococcus and melatonin level in blood serum (r = –0.58; p = 0.028).Conclusion. It was found out that patients with RPL are characterized by dysbiotic changes in the microbiome landscape. The statistically significant correlations between some microbiota representatives and neuro-immune-humoral biomarkers suggest that dysbiotic alterations in the intestine may be involved in developing immune disorders and dysregulation of the pineal-pituitary-adrenal axis underlying RPL pathogenesis.

The Nucleolus: A Central Hub for Ribosome Biogenesis and Cellular Regulatory Signals.

The nucleolus is the most prominent nuclear domain in eukaryotic cells, primarily responsible for ribosome biogenesis. It synthesizes and processes precursor ribosomal RNA (pre-rRNA) into mature rRNAs, assembling the 40S and 60S ribosomal subunits, which later form the 80S ribosome-the essential molecular machine for protein synthesis. Beyond ribosome production, the nucleolus lacks a delimiting membrane, allowing it to rapidly regulate cellular homeostasis by sequestering key stress response factors. This adaptability enables dynamic changes in size, number, and protein composition in response to cellular stress and signaling. Recent research highlights the nucleolus as a critical regulator of chemoresistance. Given its central role in cell survival and stress adaptation, the nucleolus has become an attractive therapeutic target, particularly in cancer treatment. A deeper understanding of nucleolar metabolism could pave the way for novel therapeutic strategies against various human diseases.

Morphological, Pathological and Phylogenetic Analyses of Phytophthora heterospora Causing Durian Stem Canker Disease in the Eastern Coast of Peninsular Malaysia

ABSTRACTDurian (Durio zibethinus), a highly popular tropical fruit in Southeast Asia, faces a major challenge from stem canker disease, primarily caused by oomycetes. This study aimed to identify and characterise the causal agent of durian stem canker on the eastern coast of Peninsular Malaysia. A combination of morphological, pathological and phylogenetic analyses was employed to comprehensively understand the pathogen's characteristics. A series of samplings was conducted on several durian orchards. Isolates were identified using morphological characteristics, DNA sequences and phylogenetic analysis of multiple markers, namely the internal transcribed spacer (ITS) region of the ribosomal RNA gene cluster, β‐tubulin (BTUB) and cytochrome c oxidase subunit I (COXI). Three representative isolates were selected for pathogenicity tests on durian seedlings. The sampling sites had disease incidence ranging from 10% to 54%. A total of 25 isolates were recovered from symptomatic tissues and showed morphological features that resembled those of Phytophthora species. Phylogenetic analyses using concatenated markers put all of the isolates in the same group as Phytophthora heterospora, supported with 98% maximum‐likelihood bootstrap values. The result of the pathogenicity tests revealed P. heterospora as the causal agent of stem canker of durian.

Reference Whole Genome Sequence Analyses and Characterization of a Novel Carnobacterium maltaromaticum Distinct Sequence Type Isolated from a North American Gray Wolf (Canis lupus) Gastrointestinal Tract.

We hypothesize that bacteria isolated from free-ranging animals could potentially be useful for practical applications. To meet this objective a Gram-positive bacterium was isolated from the gastrointestinal (GI) tract of a Gray Wolf (Canis lupus) using Brucella broth with hemin and vitamin K (BBHK). By small ribosomal RNA (16S) gene sequencing the bacterium was initially identified as a novel Carnobacterium maltaromaticum strain. The bacterium could be propagated both anaerobically and aerobically and was both catalase/oxidase negative and negative by the starch hydrolysis as well as negative using lipase assays. The reference whole genome sequence (WGS) was obtained using both Illumina and Nanopore sequencing. The genome assembly was 3,512,202 bp in length, encoding core bacterial genes with a GC% content of 34.48. No lysogenic bacteriophage genes were detected, although the genome harbors genes for the expression of bacteriocin and other secondary metabolites with potential antimicrobial properties. Multilocus sequence typing (MLST), WGS phylogenetics, average nucleotide identity (ANI), and single nucleotide polymorphism (SNP) analyses of the isolate's genome indicate this bacterium is a newly identified Carnobacterium maltaromaticum sequence type (ST). Members of the Carnobacteria have anti-listeria activities, highlighting their potential functional properties. Consequently, the isolate could be a potential probiotic for canids and this is the first report on an axenic C. maltaromaticum culture from the genus Canis.

Mitochondrial Genomes of Six Snakes (Lycodon) and Implications for Their Phylogeny.

Colubridae, known to be one of the most species-rich snake families, remains relatively understudied in termshe context of complete mitochondrial genome research. This study provide the first systematic characterization of the mitochondrial genomes of six colubrid species: Lycodon subcinctus, Lycodon rosozonatus, Lycodon fasciatus, Lycodon gongshan, Lycodon futsingensis, and Lycodon aulicus. In this study, mitochondrial genomes were sequenced using Sanger sequencing. The raw data were subjected to quality- filtered withing using Fastp and subsequently assembled into complete mitochondrial genomes via SPAdes. Gene annotation was performed by Tblastn, Genewise (for CDS coding sequences), MiTFi (for transfer RNAs), and Rfam (for ribosomal RNAs). Sequence analyses were conducted with various tools, including MEGA, tRNAscan-SE, DnaSP, MISA, and REPuter. Finally, phylogenetic trees were reconstructed based on 13 protein-coding genes from 14 species. The mitogenomes of these six species ranged from 17,143 to 17,298 bp in length and con-sisted of 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs), 2 ribosomal RNA genes (rRNAs), and 2 control regions. The nucleotide composition of the Colu-bridae mitogenomes was comparable with an A + T composition ranging from 52.1% to 58.8% except for the trnS1 and trnC. All the tRNAs could fold into a stable secondary structure. The Pi and Ka/Ks values suggested that atp8 was the fastest-evolving gene, while cox1 was the most conserved gene. Bayesian inference and maximum likelihood phylogenetic analyses yielded consistent results, with the six sequenced species clus-tering together with their congeneric species. These findings will provide valuable references for further research on the phylogeny of Colubridae.

Optimization of Mapping Tools and Investigation of Ribosomal RNA Influence for Data-Driven Gene Expression Analysis in Complex Microbiomes.

For gene expression analysis in complex microbiomes, utilizing both metagenomic and metatranscriptomic reads from the same sample enables advanced functional analysis. Due to their diversity, metagenomic contigs are often used as reference sequences instead of complete genomes. However, studies optimizing mapping strategies for both read types remain limited. In addition, although transcripts per million (TPM) is commonly used for normalization, few studies have evaluated the influence of ribosomal RNA (rRNA) in metatranscriptomic reads. This study compared Burrows-Wheeler Aligner-Maximal Exact Match (BWA-MEM) and Bowtie2 as mapping tools for metagenomic contigs. Even after optimizing Bowtie2 parameters, BWA-MEM showed higher efficiency in mapping both metagenomic and metatranscriptomic reads. Further analysis revealed that rRNA sequences contaminate predicted protein-coding regions in metagenomic contigs. When comparing TPM values across samples, contamination by rRNA led to an overestimation of TPM changes. This effect was more pronounced when the difference in rRNA content between samples was larger. These findings suggest that metatranscriptomic reads mapped to rRNA should be excluded before TPM calculations. This study highlights key factors influencing read mapping and quantification in gene expression analysis of complex microbiomes. The findings provide insights for improving analytical accuracy and advancing functional studies using both metagenomic and metatranscriptomic data.

Spegazzinia menglaensis, A Novel Species of Didymosphaeriaceae from Air in China.

A novel fungal species, viz. Spegazzinia menglaensis belonging to Didymosphaeriaceae (Pleosporales, Dothideomycetes), was isolated from air samples in China. Spegazzinia menglaensis is characterized by basauxic conidiophores, with a light brown conidiophore mother cell, and 4-celled, stellate α conidia and disc-shaped β conidia. To further confirm its taxonomic placement, in-depth molecular analyses were conducted, utilizing multiple loci including internal transcribed spacers (ITS), large subunit of nuclear ribosomal RNA gene (LSU), translation elongation factor 1-α (tef1-α). The molecular analyses revealed that S. menglaensis formed distinct clade within Spegazzinia, which is close to S. jinghaensis. To provide a comprehensive understanding of the new fungal species, detailed descriptions, accompanied by illustrative images, as well as a phylogenetic tree showcasing the position of S. menglaensis within the taxonomic framework, have been included.

MYCN and KAT2A form a feedforward loop to drive an oncogenic transcriptional program in neuroblastoma

The oncoprotein MYCN drives malignancy in various cancer types, including neuroblastoma (NB). However, our understanding of the mechanisms underlying its transcriptional activity and oncogenic function, as well as effective strategies to target it, remains limited. We discovered that MYCN interacts with the transcriptional coactivator KAT2A, and this interaction significantly contributes to MYCN’s activity in NB. Our genome-wide analyses indicate MYCN recruits KAT2A to bind to DNA, thereby transcriptionally regulating genes associated with ribosome biogenesis and RNA processing. Moreover, we identified that MYCN directly activates KAT2A transcription, while KAT2A acetylates MYCN, increasing MYCN protein stability. Consequently, MYCN and KAT2A establish a feedforward loop that effectively regulates global gene expression, governing the malignant NB phenotype. Treatment of NB cells with a KAT2A Proteolysis Targeting Chimera (PROTAC) degrader reduces MYCN protein levels, antagonizes MYCN-mediated gene transcription regulation and suppresses cell proliferation. This study highlights the potential of transcriptional cofactors as viable targets for developing anti-MYCN therapies.

Multi-omics analysis reveals CMTR1 upregulation in cancer and roles in ribosomal protein gene expression and tumor growth

BackgroundCMTR1 (cap methyltransferase 1), a key nuclear mRNA cap methyltransferase, catalyzes 2’-O-methylation of the first transcribed nucleotide, a critical step in mRNA cap formation. Previous studies have implicated CMTR1 in embryonic stem cell differentiation and immune responses during viral infection; however, its role in cancer biology remains largely unexplored. This study aims to elucidate CMTR1’s function in cancer progression and evaluate its potential as a novel therapeutic target in certain cancer types.MethodsWe conducted a comprehensive multi-omics analysis of CMTR1 across various human cancers using TCGA and CPTAC datasets. Functional studies were performed using CRISPR-mediated knockout and siRNA knockdown in human and mouse basal-like breast cancer models. Transcriptomic and pathway enrichment analyses were carried out in CMTR1 knockout/knockdown models to identify CMTR1-regulated genes. In silico screening and biochemical assays were employed to identify novel CMTR1 inhibitors.ResultsMulti-omics analysis revealed that CMTR1 is significantly upregulated at the mRNA, protein, and phosphoprotein levels across multiple cancer types in the TCGA and CPTAC datasets. Functional studies demonstrated that CMTR1 depletion significantly inhibits tumor growth both in vitro and in vivo. Transcriptomic analysis of CMTR1 knockout cells revealed that CMTR1 primarily regulates ribosomal protein genes and other transcripts containing 5’ Terminal Oligopyrimidine (TOP) motifs. Additionally, CMTR1 affects the expression of snoRNA host genes and snoRNAs, suggesting a broader role in RNA metabolism. Mechanistic studies indicated that CMTR1’s target specificity is partly determined by mRNA structure, particularly the presence of 5’TOP motifs. Finally, through in silico screening and biochemical assays, we identified several novel CMTR1 inhibitors, including N97911, which demonstrated in vitro growth inhibition activity in breast cancer cells.ConclusionsOur findings establish CMTR1 as an important player in cancer biology, regulating critical aspects of RNA metabolism and ribosome biogenesis. The study highlights CMTR1’s potential as a therapeutic target in certain cancer types and provides a foundation for developing novel cancer treatments targeting mRNA cap methylation.

First report of grapevine red blotch virus causing grapevine red blotch disease in New England vineyards.

Grapevine red blotch virus (GRBV), a member of the Grabovirus genus in the Geminiviridae family, is the causative agent of grapevine red blotch disease (Al Rwahnih et al. 2013). This disease affects the production and quality of grapevines (Vitis vinifera L.) (Krenz et al. 2023) and has been reported in several grape-producing states across the US (EPPO 2025), but not in the Southeastern New England American Viticultural Area (SNE-AVA). During an extensive survey conducted from 2018 to 2020 on various grapevine RNA viruses in SNE-AVA vineyards (Dumas et al. 2023), we noticed that 28 leaf samples from five vineyards (two from Connecticut, one from Massachusetts, and two from Rhode Island) from plants displaying symptoms similar to leafroll disease (redness, yellowing, leaf distortion, and chlorotic mottling) were negative for all RNA viruses tested. Total RNA was extracted from the five vineyard samples and pooled into a composite sample (0.5µg of each RNA sample). The composite sample was used for library preparation following the ribosomal RNA depletion protocol (NEB Waltham, MA) and sequencing on an Illumina NextSeq2000 platform (producing 150 bp paired-end reads). A total of 66,300,500 clean reads were obtained and assembled into contigs using Megahit version 1.1.2 (Li et al. 2015). Nine contigs demonstrated a nucleotide identity ranging from 84% to 97% with the GRBV isolate CF214-1 (NC_022002), while none were mapped to other viral or viroidal sequences in GenBank. Total DNA was extracted from all 28 samples and amplified using rolling circle amplification (RCA) with random primers. The RCA product was used as a template for the PCR primers 1402c/1404v and 2287v/604c (Ouro-Djobo et al. 2023) targeting the GRBV genome. PCR products were Sanger sequenced and assembled on SnapGene software (Dotmatics; version 7.2) using the GRBV isolate CF214-1 (Al Rwahnih et al. 2013) as the reference genome. Multiple sequence alignment was conducted using ClustalW, and phylogenetic trees were constructed using the Maximum likelihood method based on the Tamura-Nei model on MEGA-11 Software (Version 11.0.3). Eighteen of the 28 samples analyzed tested positive for GRBV by PCR and were confirmed by blasting analysis results using the sequences acquired. The whole genome sequence (3204 bp) was acquired for seven isolates (PQ601719-PQ601725) and partial genome sequences (~1409 bp) for eleven isolates (PQ601707-PQ601718). Phylogenetic analyses of the sequences acquired in this study separated the SNE-AVA isolates into two distinct phylogenetic clades: sixteen isolates grouped in clade 1. In contrast, only two isolates, one from CT and another from RI, were grouped in clade 2. To the best of our knowledge, this is the first report of GRBV in the SNE-AVA vineyards. This report highlights the need to instate a grapevine testing program in NE to improve the sanitary status of planting material.

The winner takes it all: a single genotype of Kalanchoe ×houghtonii is a global invader.

The winner takes it all: a single genotype of Kalanchoe ×houghtonii is a global invader.

Mass spectrometry analysis of PM2.5 in poultry farms and the cytotoxicity and metabolism perturbation of BEAS-2B cells.

To evaluate the potential risks posed by farm-derived fine particulate matter (PM2.5), we conducted a comprehensive analysis of PM2.5 samples collected from chicken farms. Specifically, water-soluble ions, metal and metalloids, and volatile organic compounds (VOCs) were quantitatively determined via ion chromatography, inductively coupled plasma mass spectrometry (ICP-MS), and gas chromatography‒mass spectrometry (GC‒MS), respectively. Furthermore, the microbial composition was elucidated through 16S ribosomal RNA (rRNA) high-throughput sequencing and ribosomal DNA (rDNA)-internal transcribed spacer (ITS) analysis. The study revealed that the water-soluble ion profile of PM2.5 was dominated by NO3-, NH4+, and SO42-, among others. Notably, aluminum, zinc, and manganese emerged as metals with relatively high concentrations. The primary VOCs identified were formic acid, acetic acid, and propionic acid. Microbiologically, Aspergillus and Faecalibacterium were the predominant genera detected. Upon exposure to PM2.5, BEAS-2B cells exhibited marked morphological alterations and a decrease in cell viability. Additionally, a dose-dependent increase in intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) levels was observed, accompanied by a decrease in superoxide dismutase (SOD) activity. This oxidative stress was further corroborated by elevated levels of inflammatory cytokines, including IL-6, IL-8, and TNF-α. Our findings suggest that livestock-generated PM2.5 significantly impacts cellular metabolism, particularly amino acid and nucleotide metabolism. Notably, PM2.5 from these environments can elicit cellular oxidative stress and inflammatory responses, which, with prolonged exposure, may lead to adverse health outcomes in both animals and humans. Therefore, the physical, chemical, and microbial characteristics of PM2.5 in poultry farms cannot be overlooked, emphasizing the critical need to improve the air quality within these facilities.

Off-grid field-deployable molecular diagnostic platform for malaria surveillance

BackgroundMalaria, a major global health concern, continues to cause substantial morbidity and mortality, particularly in tropical regions. Traditional malaria diagnostic methods such as microscopy and quantitative polymerase chain reaction (qPCR) are effective but face challenges in field settings because of their requirement for laboratories with specialized equipment and trained personnel. This study presents the development and validation of a portable, cost-effective, field-deployable real-time qPCR platform for detecting Plasmodium species.MethodsField-compatible DNA isolation was performed using DNAzol, and TaqMan probes targeting 18S ribosomal RNA (rRNA) were employed to detect five Plasmodium species—P. falciparum, P. vivax, P. malariae, P. ovale, and P. knowlesi—using the bCUBE qPCR platform. In vitro-cultured P. falciparum and experimentally infected Anopheles gambiae were used to quantify P. falciparum infections, with infection prevalence compared to microscopy. The bCUBE qPCR system was also evaluated under field conditions to detect P. falciparum infections in field-collected An. gambiae mosquitoes.ResultsThe bCUBE qPCR demonstrated a strong linear correlation (R2 = 0.993) with a standard laboratory qPCR machine for detecting P. falciparum infections. It successfully detected as few as 0.5 parasites/µl of blood, one oocyst in mosquito guts, and 5–10 sporozoites in salivary glands. It was also capable of discriminating between P. falciparum, P. vivax, P. malariae, P. ovale, and P. knowlesi. Field evaluations in Cameroon confirmed its accuracy in identifying P. falciparum in mosquito samples, with same-day results. The capability of the bCUBE qPCR system to detect infections in both individual and pooled mosquito surveillance further highlights its potential for in-field large-scale malaria monitoring surveillance.ConclusionsThe bCUBE qPCR system offers a portable, sensitive, and scalable solution for malaria diagnostics, enabling real-time surveillance in resource-limited settings. Its ability to provide rapid, on-site results reduces the need for centralized laboratory testing, facilitating timely decision-making in malaria control programs.Graphical

Assessment of rhizosphere microbial activity using optimized RNA extraction coupled with universal ribosomal RNA depletion techniques

BackgroundMetatranscriptomics, which analyzes gene expression patterns in a heterogeneous community, is a powerful tool to evaluate microbial functional activity. A key challenge in this process is obtaining high-quality RNA, which is complicated by the composition and ecosystem complexity of the soil matrix. Another crucial step involves the removal of highly abundant ribosomal RNA (rRNA), as its presence can dominate sequencing results and obscure the detection of messenger RNA (mRNA) expression. Conventional library preparation methods often struggle to efficiently remove rRNA from a complex mix of prokaryotic and eukaryotic organisms, further complicating mRNA isolation.MethodsTo overcome these limitations, we have developed an optimized method for extracting RNA from soybean rhizosphere microbes, followed by universal rRNA depletion to create rRNA-free samples for sequencing. These samples were sequenced using an Illumina high-throughput sequencer.ResultsOur optimized cetyltrimethylammonium bromide (CTAB) phenol–chloroform extraction protocol significantly improved RNA yield and quality from clay-rich soils, outperforming previously published methods and commercial kits. Illumina sequencing data revealed minimal rRNA contamination, and the resulting short reads could be successfully assembled into transcripts. These findings also demonstrate that using the Zymo-Seq RiboFree Total RNA Library Kit effectively enabled library preparation from complex environmental samples for Illumina sequencing.DiscussionThis RNA sample preparation method, combined with our optimized extraction technique, provides a valuable approach for studying rhizosphere microbes that hold exciting potential for advancing soil health assessments and understanding plant–microbe-pathogen interactions.

TlyA is a 23S and 16S 2'-O-methylcytidine methyltransferase important for ribosome assembly in Bacillus subtilis.

Ribosomal RNA (rRNA) is methylated in organisms ranging from bacteria to metazoans. Despite the pervasiveness of rRNA methylation in biology, the function of rRNA methylation on ribosome function is poorly understood. In this work, we identify a biological function for the rRNA 2'-O-methylcytidine methyltransferase TlyA, conserved between Bacillus subtilis and Mycobacterium tuberculosis (Mtb). The tlyA deletion in B. subtilis confers a cold sensitive phenotype and resistance to aminoglycoside antibiotics that target the 16S rRNA. We show that ΔtlyA cells have ribosome assembly defects characterized by accumulation of the 50S subunit. Using a genetic approach and based on sequence alignments with other rRNA methyltransferases we tested the importance of potential catalytic residues and S-adenosyl-L-methionine (SAM) cofactor binding sites. We show that TlyA shares the common rRNA methyltransferase catalytic triad KDK and a SAM binding motif GxSxG which differs from Mtb TlyA. Together our work demonstrates that B. subtilis tlyA is critical for ribosome assembly and we identify key residues for TlyA function in vivo. Since E. coli lacks TlyA or a functional equivalent, our work highlights key differences in ribosome maturation between B. subtilis, Mtb and more divergent Gram-negative bacteria providing new insight into translation and antibiotic resistance mechanisms.

Influence of Maternal Weight Dynamics Prior to and Throughout Gestation on Early Infant Gut Microbiome Colonization

This study is aimed at exploring the relationship between maternal weight categories, including pre-pregnancy body mass index (P-BMI) and gestational weight gain (GWG), and the composition of the infant gut microbiome in the early days of life. We recruited 71 mother-infant pairs from Kangwon National University Hospital and Bundang CHA Hospital, collecting meconium samples from the infants within the first 5 days postpartum. Using 16S ribosomal RNA gene sequencing (V3–V4 region), this study assessed microbial diversity and the relative abundance of specific bacterial taxa in these initial stool samples. Participants were categorized into groups based on maternal P-BMI and GWG, enabling a comprehensive comparison of the microbiota composition in the infants’ meconium across different maternal weight metrics. Our analysis identified significant variations in the infant gut microbiome correlated with maternal weight categories. Key findings include a differential abundance of genera such as Sphingobacteriaceae, Bacillaceae, Cytophagaceae, and Alteromonadaceae across maternal P-BMI groups, whereas Moraxellaceae and Rhodospirillaceae varied across GWG groups. In the P-BMI category, infants born to overweight mothers demonstrated a higher abundance of Pseudopedobacter, and a lower abundance of Citrobacter and Lachnospira, while infants in the underweight group showed a higher abundance of Lachnospira and Weissella. In the normal weight group, Citrobacter and Pseudopedobacter were more abundant. Within the GWG category, infants in the inadequate group showed a higher abundance of Klebsiella, whereas the normal group showed a higher abundance of Holdemania. The composition of the infant gut microbiome in the early postnatal period is significantly influenced by maternal weight categories. Understanding the role of maternal weight in shaping early microbial colonization may provide insights into developing strategies to optimize infant health outcomes through targeted interventions before and during pregnancy.

The gut microbial community structure of the oriental armyworm Mythimna separata (Walker) (Lepidoptera: Noctuidae) affects the the virulence of the entomopathogenic fungus Metarhizium rileyi

Mythimna separata, the oriental armyworm, is a lepidopteran pest that threatens cereal crops. In the current study, two strains (XSBN200920 and JHML200710) of entomopathogenic fungus Metarhizium rileyi were tested for virulence against oriental armyworms. When treated with spore suspensions of both strains at a concentration of 1.0 × 108 spores/mL, the 3rd instar larvae's survival rate was considerably different (P < 0.01). The median lethal time of the insects exposed to XSBN200920 was about 3 d longer than that of JHML200710. The results of 16S ribosomal RNA sequencing showed that Chao1 richness in the JHML200710 treatment group was significantly decreased compared with the CK ( 0.02% Tween 80). The dominant gut bacteria species at the phylum level were Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidota in the three groups. The CK group had a much higher associated abundance of cyanobacteria than the other two fungal treatment groups. Sixteen genera revealed significant variations in the gut bacteria of the insects at the genus level. The Kyoto Encyclopedia of Genes and Genomes (KEGG) functional gene and enzyme analysis showed that when compared with the CK group, the XSBN200920 treatment group showed a significant reduction in six aspects, including betalain biosynthesis, spliceosome, and neuroactive ligand-receptor interaction. These findings suggested that healthy and fungus-infected insects' intestinal microbial community structure differed significantly. And the virulence of M. rileyi is closely linked to its ability to alter the structure of the intestinal microbiome of insects. The results offer a starting point for examining the relationship between the gut microbial diversity of oriental armyworms and variations in the virulence of pathogenic fungi.