Search Results (621)

Fecal water syndrome (FWS) in horses is characterized by two-phase defecation, including both solid and liquid phases. While satisfactory explanations for FWS are unavailable, bacterial dysbiosis has been suggested as a contributing or causative factor. The objectives of this study were to determine whether fecal bacterial dysbiosis is associated with FWS in horses in the midwestern USA. Fecal samples were collected from horses with FWS and from unaffected horses at the same location. In total, 16S rRNA amplicon libraries produced from fecal bacterial DNA were sequenced using the Illumina sequencing platform. Significant differences in beta diversity were detected between affected and control horses (p = 7 × 10⁻⁴, F = 1.51), and differential abundance testing identified several features enriched in affected and control horses. These results agree with prior work regarding specific features in the bacterial microbiome associated with FWS, including Alloprevotella spp., and suggest fecal dysbiosis is associated with FWS. Full article

Hippophae rhamnoides subsp. sinensis is extensively found in China, where the annual precipitation ranges from 400 to 800 mm. It is the most dominant species in natural sea buckthorn forests and the primary cultivar for artificial ecological plantations. Additionally, it exhibits significant nutritional and medicinal value, making it a renowned eco-economic tree species. Despite extensive research into its ecological functions and health benefits, the mitochondrial genome of this widespread species has not yet been published, and knowledge of the mitochondrial genome is crucial for understanding plant environmental adaptation, evolution, and maternal inheritance. Therefore, the complete mitochondrial genome was successfully assembled by aligning third-generation sequencing data to the reference genome sequence using the Illumina NovaSeq 6000 platform and Nanopore Prometh ION technologies. Additionally, the gene structure, composition, repeat sequences, codon usage bias, homologous fragments, and phylogeny-related indicators were also analyzed. The results showed that the length of the mitochondrial genome is 454,489 bp, containing 30 tRNA genes, three rRNA genes, 40 PCGs, and two pseudogenes. A total of 411 C-to-U RNA editing sites were identified in 33 protein-coding genes (PCGs), with higher frequencies observed in ccmFn, ccmB, nad5, ccmC, nad2, and nad7 genes. Moreover, 31 chloroplast-derived fragments were detected, accounting for 11.86% of the mitochondrial genome length. The ccmB, nad4L, and nad7 genes related to energy metabolism exhibited positive selection pressure. The mitochondrial genome sequence similarity between H. rhamnoides subsp. sinensis and H. tibetana or H. salicifolia was 99.34% and 99.40%, respectively. Fifteen shared gene clusters were identified between H. rhamnoides subsp. sinensis and H. tibetana. Phylogenetically, the Rosales order showed close relationships with Fagales, Fabales, Malpighiales, and Celastrales. These findings provide fundamental data for exploring the widespread distribution of H. rhamnoides subsp. sinensis and offer theoretical support for understanding the evolutionary mechanisms within the Hippophae genus and the selection of molecular breeding targets. Full article

Bacterial symbionts play an important role in insect survival by contributing to key metabolic and defensive functions. While stingless bees are known to harbor diverse microbial communities, their core bacterial symbionts remain poorly characterized. In this study, we analyzed the gut microbiota of sixteen stingless bee species collected from different regions of Mexico using 16S rRNA gene sequencing on the Illumina^® MiSeq™ platform. Our results revealed that Proteobacteria, Firmicutes, and Actinobacteria are the most abundant bacterial phyla across species. Among the dominant genera, lactic acid bacteria, such as Lactobacillus spp., Bifidobacterium, and Fructobacillus spp., were the most prevalent. These bacteria are responsible for developing biochemical functions in metabolic processes like lactic fermentation and the biotransformation of complex organic compounds into molecules that are more easily assimilated by bees. This study offers a novel perspective on the diversity and predicted composition of gut microbiota in Mexican stingless bees. By highlighting differences in microbial communities among species with different feeding habits, our results emphasize the importance of preserving microbial biodiversity in these pollinators. Full article

Treatment of Mycobacterium tuberculosis requires multi-drug regimens, and resistance to any individual antibiotic can compromise outcomes. For slow-growing organisms like M. tuberculosis, rapid detection of resistance-conferring mutations enables timely initiation of effective therapy. Conversely, confirming wild-type status in resistance-associated genes supports confidence in standard regimens. We developed an amplicon-based next generation sequencing (amplicon tNGS) assay on the Illumina platform targeting eight genes linked to resistance to isoniazid, rifampin, ethambutol, pyrazinamide, and fluoroquinolones. Sequencing results were analyzed using a custom bioinformatics pipeline. Forty-seven samples were used for assay development, and 37 additional samples underwent post-implementation clinical validation. Compared to whole genome sequencing (WGS), amplicon tNGS demonstrated 97.7% sensitivity, 98.9% specificity, and 98.7% overall accuracy for variant detection in targeted regions. Resistance prediction showed 79.3% concordance with WGS; discrepancies were primarily due to mutations outside of target regions. Among post-implementation samples, 27/37 passed quality metrics for all targets, with 95.7% concordance between amplicon tNGS results and final susceptibility results. This assay is now in use in our laboratory and offers significantly faster turnaround than both WGS and phenotypic methods on cultured isolates, enabling more rapid, informed treatment decisions for tuberculosis patients. Full article

Gray mold disease, caused by the fungus Botrytis cinerea, affects a wide variety of plants. In this study, we conducted several in vitro tests and genomic analyses on three strains of this fungus (BcPgIs-1, BcPgIs-3, MIC) previously isolated from diseased pomegranate fruits, collected at two geographic locations in Mexico. Our goal was to identify possible differences among these strains. The development of the three strains in distinct culture media, the production of extracellular enzymes, and their effect on the progression of infection in pomegranate fruits were evaluated. The genomes were sequenced using the Illumina platform and analyzed with various bioinformatics tools. All strains possess genetic determinants for virulence and cell wall polymer degradation, but MIC exhibited the highest pectinolytic activity in vitro. This strain also produced sclerotia in a shorter time (7 days) in PDA medium. BcPgls-3 demonstrated the highest conidia production across all the culture media used. Both BcPgls-3 and MIC damaged all the pomegranate fruits 8 days after inoculation, while the BcPgls-1 required up to 9 days. Sequencing of the three strains yielded high-quality sequences, resulting in a total of 17 scaffolds and genomes that exceed 41 million bp, with a GC content of approximately 42%. Phylogenomic analysis indicated that the MIC strain is situated in a group separate from BcPgIs-1 and BcPgIs-3. BcPgIs-3 possesses more coding sequences, but MIC has more genes for CAZymes and peptidases. The three strains share 10,174 genes, while BcPgIs-3 and MIC share 851. These findings highlight the differences among the strains studied, which may reflect their adaptive capacities to their environment. Results contribute to our understanding of the biology of gray mold in pomegranates and could assist in developing more effective control strategies. Full article

Introduction: Genetic plasticity and adaptive camouflage in critical pathogens have contributed to the global surge in multidrug-resistant (MDR) infections, posing a serious threat to public health and therapeutic efficacy. Antimicrobial resistance, now a leading cause of global mortality, demands urgent action through diagnostics, vaccines, and therapeutics. In India, the Indian Council of Medical Research’s surveillance network identifies Escherichia coli as a major cause of urinary tract infections, with increasing prevalence in human gut microbiomes, highlighting its significance across One Health domains. Methods: Whole-genome sequencing of E. coli strain ECG015, isolated from a human gut sample, was performed using the Illumina NextSeq platform. Results: Genomic analysis revealed multiple antibiotic resistance genes, virulence factors, and efflux pump components. Phylogenomic comparisons showed close relatedness to pathovars from both human and animal origins. Notably the genome encoded protein tyrosine kinases (Etk/Ptk and Wzc) and displayed variations in the envelope stress-responsive CpxAR two-component system. Promoter analysis identified putative CpxR-binding sites upstream of genes involved in resistance, efflux, protein kinases, and the MazEF toxin–antitoxin module, suggesting a potential regulatory role of CpxAR in stress response and persistence. Conclusions: This study presents a comprehensive genomic profile of E. coli ECG015, a gut-derived isolate exhibiting clinically significant resistance traits. For the first time, it implicates the CpxAR two-component system as a potential central regulator coordinating antimicrobial resistance, stress kinase signaling, and programmed cell death. These findings lay the groundwork for future functional studies aimed at targeting stress-response pathways as novel intervention strategies against antimicrobial resistance. Full article

Background: Saxifraga umbellulata var. pectinata (Saxifragaceae) is recognized as a genuine medicinal material from the Qinghai–Tibet Plateau in China. This paper presents the chloroplast (cp) genome of S. umbellulata var. pectinata, marking the first report for this genus. The Tibetan medicinal plants documented in ‘Chinese Medicinal Plant Resources’ are associated with their chloroplast genomes and medicinal mechanisms. Objective: In order to resolve any potential ambiguity in conventional classifications, this study reconstructs the evolutionary position of S. umbellulata var. pectinata within the genus by comparing its chloroplast genetic information with that of other groupings. Methods: The chloroplast genome of S. umbellulata var. pectinata was sequenced using the Illumina NovaSeq 6000 platform. Subsequent sequence assembly, annotation, and characterization were performed using bioinformatics analysis. The NJ phylogenetic tree was constructed using MEGA 7.0 software. Results: The complete chloroplast genome of S. umbellulata var. pectinata is 146,549 bp in length, comprising four subregions: a large single-copy (LSC) region of 79,318 bp and a small single-copy (SSC) region of 16,390 bp, separated by a pair of inverted repeat (IR) regions each 25,421 bp long. This cp genome contains 131 genes, including 86 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. The overall GC content is 38.1%. Phylogenetic analysis based on 20 cp genomes indicates that S. umbellulata var. pectinata is closely related to Saxifraga sinomontana and Saxifraga stolonifera. Full article

The leading causes of pediatric arterial ischemic stroke (PAIS) are arteriopathies, which refer to pathologies of the arterial walls in the brain. Since traditional risk factors for cardiovascular diseases in children play a smaller role than in adults, it can be supposed that genetic factors may be of particular importance in this age group. Therefore, this study aimed to identify mutations affecting the formation of vascular wall pathologies, which can subsequently lead to ischemic stroke. The study used a database of 92 Caucasian children diagnosed with ischemic stroke. From this group, 25 children with arteriopathies were selected. The study had an exploratory and descriptive design, with the aim of characterizing rare genetic variants in a selected cohort, without attempting formal statistical association testing. The sequencing was performed using the Illumina NextSeq 550 platform. A panel of 161 genes known to be associated with stroke or arteriopathies was selected for further analysis. We identified 10 pathogenic or likely pathogenic mutations in 15 patients. Among these, three are likely monogenic causes of stroke (ELN, SCN5A, and VHL genes), two are considered risk factors (FV and ADAMTS13), two have conflicting interpretations (ACAD9 and ENG), and three are most likely benign (CBS, PMM2, and PKD1). The frequency of genetic variants underlying ischemic stroke or acting as risk factors for the disease in the studied group is significantly higher than the estimated frequency of monogenic forms of stroke in young adults and higher than in the general population. NGS testing is worth considering, especially in patients who exhibit certain symptoms that may suggest the presence of mutations. Full article

Momordica balsamina, a plant traditionally used in African medicine, contains a 30 kDa protein, MoMo30, previously identified by our group as an anti-HIV agent that binds glycan residues on the gp120 envelope protein, thereby acting as an entry inhibitor. In this study, we investigated whether prolonged exposure to MoMo30 exerts selective pressure on HIV-1 and induces mutations in the viral envelope (env) gene. T-lymphocyte cells were infected with HIV-1_NL4-3 and continuously treated with MoMo30 over a 24-day period. Viral RNA was isolated at regular intervals, and env genes were sequenced using the Illumina platform. RNA sequence variant calling was performed using iVar, which uses a frequency-based binomial test with a default allele frequency threshold of 3% and a minimum base quality of 20 and applies Bonferroni correction for multiple testing. The infectivity of the MoMo30-exposed virus was assessed using MAGI-CXCR4 cells, visualized by β-galactosidase staining, and compared to untreated controls. Statistical significance was determined via two-way ANOVA. MoMo30-treated HIV-1 exhibited multiple detrimental mutations in gp120 and gp41, including missense, nonsense, and frameshift changes. Notably, 32% of N-linked glycosylation sites were deleted in the treated virus, while no such changes were observed in controls. Functionally, the MoMo30-treated virus demonstrated a sixfold reduction in infectivity compared to untreated HIV-1_NL4-3. These findings suggest that MoMo30 imposes genetic pressure on HIV-1_NL4-3, selecting for mutations that reduce viral fitness. Full article

The aim of this study was to examine the genetic characteristics that could be associated with the virulence characteristics of Escherichia coli collected from clinical samples. A collection of 100 non-repetitive E. coli isolates was analyzed. All isolates were typed by MLST. String production, biofilm formation and serum resistance were examined for all isolates. Twenty E. coli isolates were completely sequenced Illumina platform. The results showed that the majority of E. coli isolates (87%) produced significant levels of biofilm, while none of the isolates were positive for string test and resistance to serum. Additionally, the presence of CRISPR/Cas systems (type I-E or I-F) was found in 18% of the isolates. Analysis of WGS data found that all sequenced isolates harbored a variety of virulence genes that could be implicated in adherence, invasion, iron uptake. Also, WGS data confirmed the presence of a wide variety of resistance genes, including ESBL- and carbapenemase-encoding genes. In conclusion, an important percentage (87%) of the E. coli isolates had a significant ability to form biofilm. Biofilms, due to their heterogeneous nature and ability to make microorganisms tolerant to multiple antimicrobials, complicate treatment strategies. Thus, in combination with the presence of multidrug resistance, expression of virulence factors could challenge antimicrobial therapy of infections caused by such bacteria. Full article

Background/Objectives: Endplate lesions of the lumbar spine are often asymptomatic and frequently observed incidentally by radiological assessment. Variants in the vitamin D receptor gene (VDR) and an increase in some biochemical markers related to the osteo-cartilaginous metabolism were found in patients with endplate lesions. The aim of this study was to identify biochemical and genetic markers putatively associated with the presence of endplate lesions of the lumbar spine. Methods: Quantification of circulating bone remodeling proteins was obtained from 10 patients with endplate lesions and compared with age- and sex-matched controls. Whole exome sequencing (WES) was performed on patient genomic DNA using the Novaseq 6000 platform (Illumina, San Diego, CA, USA), obtaining a median read depth of 117×–200×, with ≥98% of regions covering at least 20×. The sequencing product was aligned to the reference genome (GRCh38.p13-hg38) and analyzed with Geneyx software. Results: We observed modifications in the levels of circulating proteins involved in bone remodeling and angiogenesis. We identified variants of interest in aggrecan (ACAN), bone morphogenetic protein 4 (BMP4), cytochrome P450 family 3 subfamily A member 4 (CYP3A4), GLI family zinc finger 2 (GLI2), heparan sulfate proteoglycan 2 (HSPG2), and mesoderm posterior bHLH transcription factor 2 (MESP2). VDR polymorphism (rs2228570) was present in nine patients, with the homozygotic ones having more severe endplate lesions and higher levels of the analyzed circulating markers in comparison with heterozygotic patients. Conclusions: These data represent interesting evidence of genetic variants, particularly in VDR, and altered levels of circulating markers of bone remodeling associated with endplate lesions, which should be confirmed in a larger population. The hypothesis suggested by our results is that the endplate lesions could be the consequence of an altered ossification mechanism at the vertebral level. Full article

Background: Antimicrobial resistance (AMR) is a critical global health threat, with AMR Salmonella enterica serovar Typhimurium strains being a major foodborne pathogen. Integrons, a type of mobile genetic element, capture and transfer resistance genes, thereby playing a role in the spread of AMR. Objectives: This study aimed to characterize the locations of integrons carrying AMR genes within the whole genomes of 32 Salmonella Typhimurium isolates collected from dairy cattle by two U.S. Veterinary Diagnostic Laboratories between 2009 and 2012. Methods: Class I integrons were sequenced from PCR-amplified products. DNA was extracted, quantified, barcoded, and sequenced on the Illumina MiSeq platform. Whole genome sequences were trimmed and assembled using the SPAdes assembler in Geneious Prime^®, and plasmids were identified with the PlasmidFinder pipeline in Linux. Integron locations were determined by aligning their sequences with whole genome contigs and plasmids, while AMR genes were identified through BLAST with the MEGARes 3.0 database and confirmed by alignment with isolate, plasmid, and integron sequences. Statistical analysis was applied to compare the proportions of isolates harboring integrons on their chromosome versus plasmids and also to examine the associations between integron presence and AMR gene presence. Results: Seven plasmid types were identified from all isolates: IncFII(S) (n = 14), IncFIB(S) (n = 13), IncC (n = 7), Inc1-I(Alpha) (n = 3), and ColpVC, Col(pAHAD28), and Col8282 (1 isolate each). Of the 32 isolates, 16 (50%) carried at least one size of integron. Twelve of them carried both 1000 and 1200 bp; 3 carried only 1000 bp and 1 carried 1800 bp integrons. Of the 15 isolates that carried 1000 bp integron, 12 harbored it on IncFIB(S) plasmids, 2 on IncC plasmids, and 1 on the chromosome. The 1200 bp integrons from all 12 isolates were located on chromosomes. There were significant positive associations between the presence of integrons and the presence of several AMR genes including sul1, aadA2, blaCARB-2, qacEdelta1, tet(G), and floR (p < 0.05). AMR genes were located as follows: aadA2 on IncFIB(S) and IncC plasmids; bla_CMY-2 on IncC plasmid; qacEdelta1 on IncFIB(S), IncC, and chromosome; bla_CARB-2, floR, tet(A) and tet(G) on the chromosome. Conclusions: The findings highlight the genomic and plasmid complexity of Salmonella Typhimurium which is impacted by the presence and location of integrons, and this study provides genomic insights that can inform efforts to enhance food safety and protect both animal and public health. Full article

Outbreaks of infectious diseases contribute significantly to morbidity and mortality in resource-limited settings, yet the capacity to identify their etiology remains limited. We aimed to characterize microbes and antimicrobial resistance (AMR) genes in Tanzanian children affected by an acute febrile illness (AFI) outbreak using metagenomic next-generation sequencing (mNGS). A cross-sectional study was conducted on archived blood samples from children who presented with AFI between 2018 and 2019. Total nucleic acids were extracted from 200 µL of blood, and complementary DNA (cDNA), along with enriched pathogenic DNA, was sequenced using the Illumina MiSeq platform. mNGS data were analyzed using CZ-ID Illumina mNGS bioinformatics pipeline v7.0. Results were obtained from 25 participants (mean age: 11.6 years; SD ± 5), of whom 36% had a moderate to high-grade fever. The following five potential microbial causes of AFI were identified: Escherichia coli (n = 19), Paraclostridium bifermentans (n = 2), Pegivirus C (n = 2), Shigella flexneri (n = 1) and Pseudomonas fluorescens (n = 1), with E. coli being the most prevalent. Twelve AMR genes were detected, including mdtC, acrF, mdtF, and emrB. E. coli harbored most of the AMR genes previously associated with resistance to commonly used antibiotics. mNGS offers a promising complementary approach to conventional diagnostics for identifying pathogens and AMR profiles in vulnerable populations. Full article

Background/Objectives: Brain arteriovenous malformations (bAVMs) are rare vascular anomalies characterized by direct connections between arteries and veins, bypassing the capillary network. This study aimed to identify potential genetic factors contributing to the development of sporadic bAVMs. Methods: Three patients (AVM1–3) from Kazakhstan who underwent microsurgical resection at the National Centre for Neurosurgery (NCN) in Astana, Kazakhstan, were analyzed. Brain AVMs were diagnosed using magnetic resonance imaging (MRI). Genomic DNA was isolated from whole venous blood samples, and whole-exome sequencing was performed on the NovaSeq 6000 platform (Illumina). Variants were filtered according to standard bioinformatics protocols, and candidate gene prioritization was conducted using the ToppGene tool. Results: In silico analysis further revealed candidate genes likely associated with lesion development, including COL3A1, CTNNB1, LAMA1, NPHP3, SLIT2, SLIT3, SMO, MAPK3, LRRK2, TTN, ERBB2, PARD3, and OBSL1. It is essential to focus on the genetic variants affecting the following prioritized genes: ERBB2, SLIT3, SMO, MAPK3, and TTN. Mutations in these genes were predicted to be “damaging”. Most of these genes are involved in signaling pathways that control vasculogenesis and angiogenesis. Conclusions: Defects in genes associated with ciliary structure and function may be critical to the pathogenesis of brain AVMs. These findings provide valuable insights into the molecular underpinnings of bAVM development, emphasizing key biological pathways and potential candidate genes. Further research is needed to establish robust correlations between specific genetic mutations and clinical phenotypes, which could ultimately inform the development of improved diagnostic, therapeutic, and prognostic approaches. Full article

Conventional diagnostic methods (CDMs) for lower respiratory infections (LRIs) have limitations in detecting causative pathogens. This study evaluates the utility of shotgun metagenomic sequencing (SMS) as a complementary diagnostic tool using bronchoalveolar lavage (BAL) fluid. Sixteen BAL fluid samples from pneumonia patients with positive CDM results—including bacterial/fungal cultures; PCR for Mycobacterium tuberculosis or cytomegalovirus; and the BioFire^® FilmArray^® Pneumonia Panel (BioFire Diagnostics LLC, Salt Lake City, UT, USA)—underwent 10 Gb SMS on the Illumina NovaSeq 6000 platform (Illumina, San Diego, CA, USA). Reads were aligned to the NCBI RefSeq database; with fungal identification further supported by internal transcribed spacer (ITS) analysis. Antibiotic resistance genes (ARGs) were annotated using the Comprehensive Antibiotic Resistance Database. Microbial reads accounted for 0.00002–0.04971% per sample. SMS detected corresponding bacteria in 63% of cases, increasing to 69% when subdominant taxa were included. Fungal reads were low; however, Candida species were identified in four samples via ITS. No viral reads were detected. ARGs meeting perfect match criteria were found in two cases. This is the first real-world study comparing SMS with CDMs, including semiquantitative PCR, in BAL fluid for LRI. SMS shows promise as a supplementary diagnostic method, with further research needed to optimize its performance and cost-effectiveness. Full article