Search Results (791)

The oral microbiome of patients with acute lymphoblastic leukemia (ALL) undergoes changes caused by the neoplasia as well as the antimicrobial activity of chemotherapy (CTX), which promotes the development of oral mucositis (OM). This study aimed to analyze the oral microbiome dynamics and salivary cytokine production in pediatric ALL patients before and during CTX, comparing children who did and did not develop OM. We conducted a longitudinal, observational, and analytical study including 32 newly diagnosed pediatric ALL patients (ages 2–16 years) undergoing CTX. Oral rinse and non-stimulated saliva samples were collected at baseline (day 0), day 14, and day 21 of induction of CTX, with an additional sample taken during OM episodes when possible. Microbiome analysis was performed using 16S rRNA sequencing on an Illumina MiSeq platform, and salivary cytokines were measured using a Luminex multiplex assay. The most pronounced microbiome changes occurred on day 14, particularly in patients who developed OM, characterized by higher α diversity, increased abundance of opportunistic taxa, and elevated IL-6 concentrations. In contrast, patients who did not develop OM exhibited a more stable microbial composition. Overall, these findings indicate that temporal oral dysbiosis and increased IL-6 may serve as early markers and potential predictors of OM development during chemotherapy in pediatric ALL patients. Full article

Tick-borne encephalitis virus (TBEV) causes tick-borne encephalitis (TBE), a severe disease of the human central nervous system. Currently, the data on the genetic variants of TBEV in Kyrgyzstan are practically absent. Therefore, the aim of this study was to analyze and describe the genetic diversity of TBEV in this region. The complete genome sequences of seven TBEV strains from the collection of the Scientific Centre for Family Health and Human Reproduction Problems (Irkutsk, Russia) were determined. These strains, isolated from Ixodes persulcatus ticks from Kyrgyzstan, were sequenced using the next generation sequencing approach on a MiSeq high-performance sequencer (Illumina, San Diego, CA, USA). A molecular genetic analysis of the obtained sequences, along with sequences of two previously isolated TBEV strains from Kyrgyzstan available in the GenBank database, demonstrated that the Siberian subtype of three genetic lineages (Zausaev, Vasilchenko and Bosnia) is predominantly distributed in Kyrgyzstan. The Far Eastern subtype of TBEV is also present. To date, this location probably represents the southernmost boundary of these TBEV subtypes’ ranges. Full article

Ammopiptanthus mongolicus is a relict species from the ancient Mediterranean of the Tertiary period and the only strong xerophytic evergreen broad-leaved shrub in the central Asian desert. Foliar endophytic and epiphytic bacteria jointly form phyllosphere microorganisms that influence plant health. This study investigated the dynamic changes in foliar endophytic bacterial communities across four leaf growth stages (Young, Mature, Old1, and Old2). Illumina 16S region (V5–V7) amplicon sequencing was used to analyze community composition, function, construction process, and environmental driving factors. The Old1 and Old2 stages were clearly separated from the Young and Mature stages, which demonstrated closer clustering. Community diversity and evenness first increased from the Young to Mature stages, declined at the Old1 stage, and finally reached maximum values at the Old2 stage; richness increased gradually. Total amplicon sequence variant (ASV) numbers, stage-specific ASVs, and their proportion increased with leaf development, whereas the proportion of shared ASVs between adjacent, interval, and all stages decreased. Dominant genera were Rhodococcus (Young), unclassified_f__Comamonadaceae (Mature), Rhodococcus (Old1), and Bacillus (Old2). Co-occurrence networks became progressively simpler, with reduced inter-node and positive connectivity. Functional predictions revealed that chemoheterotrophy and aerobic chemoheterotrophy decreased initially and then increased, with the lowest values at Old1. N, C/P, N/P, and SOD reached maximum at the Old2 stage. P was maximum at the Mature stage. P, C/P, and N/P were significantly positively correlated with the Young stage, N with the Mature stage, and SOD with the Old2 stage (p < 0.05). These findings enhance understanding of the diversity, composition, function, and plant–endophyte relationships in xerophytic relict species, particularly evergreen desert shrubs. Full article

Analyzing the composition and structure of the gut bacterial community in Antheraea pernyi is essential for improving its economic traits, as well as for understanding gut bacteria–host interactions in lepidopteran insects. This study utilized the Illumina MiSeq PE 300 platform to conduct 16S rRNA gene sequencing for a comparative analysis of gut bacterial community in laboratory-reared and field-released (spring and autumn) Antheraea pernyi larvae of the same strain. The study revealed the specific effects of rearing environment and seasonal variation on the structural and functional dynamics of the larval gut bacterial communities. The composition of the dominant gut bacteria varied significantly with rearing environment and season. Laboratory-reared and spring field-released groups exhibited similar bacterial community structures, whereas the autumn field-released group showed a significant trend toward specialization, characterized by enrichment of specific bacterial taxa. Linear discriminant analysis effect size identified statistically significant biomarkers across samples. Taxonomic analysis revealed that Actinomycetota, Actinobacteria, Mycobacteriales, Dietziaceae, and Dietzia were characteristic of the gut bacteria profile in spring field-released, Lactobacillales, Enterococcaceae, and Enterococcus were enriched in the autumn field-released group, and the laboratory-reared group exhibited a relative dominance of Alphaproteobacteria. Functional prediction indicated that gut bacterial community structure likely influences its metabolic potential, which may suggest an adaptive response of the Antheraea pernyi to distinct ecological environments. This study provides important insights into the highly complex nature of insect-microbe interactions. Full article

In Saudi Arabia, cucurbit crops such as zucchini (Cucurbita pepo) and snake gourd (Trichosanthes cucumerina) are major vegetables and key dietary components, yet their associated viral threats remain poorly understood. We surveyed symptomatic cucurbit samples from greenhouses and open fields in the Al-Ahsa and Qatif regions. The detection methods employed included PCR, RCA, and Illumina NGS. Based on nucleotide sequence comparisons and maximum-likelihood phylogenetic analysis, we identified three viruses, i.e., TYLCV, WmCSV, and ToLCPalV, present as both single and mixed infections. Sequence analyses revealed a novel strain, TYLCV-Hasa, representing a distinct lineage of TYLCV. Analysis revealed that recombination occurred solely in the DNA-A components of the identified viruses, while DNA-B segments showed no evidence of recombination. Notably, no DNA satellites were detected, suggesting cucurbits may act as independent reservoirs of begomovirus diversity. These results provide a comprehensive genomic insight into cucurbit-infecting begomoviruses in Eastern Saudi Arabia. The discovery of TYLCV-Hasa and evidence of recombination raise concerns about the emergence of novel viral variants that could pose risks to cucurbit cultivation. The results establish a foundation for advanced molecular surveillance and breeding strategies, contributing to improved food security and supporting Saudi Arabia’s Vision 2030 goals for sustainable agriculture. Full article

Background: Cholera, caused by Vibrio cholerae, remains endemic in many developing countries, including Pakistan. The extensive use of antibiotics has led to the emergence of antimicrobial resistance in V. cholerae, limiting available treatment options. In this study, we performed molecular characterisation of antibiotic-resistant V. cholerae serotype Ogawa isolates from a recent cholera outbreak in Khyber Pakhtunkhwa, Pakistan. Methodology: Suspected cholera stool samples were collected from hospitalised patients at various district hospitals of Khyber Pakhtunkhwa Province (KPK), Pakistan. The samples were transported to the Public Health Reference Microbiology Laboratory at Khyber Medical University, Peshawar. V. cholerae were identified based on colonial morphology, Gram staining, and biochemical tests using EPI 10E. For serotype identification, monovalent antisera were used. Antibiotic susceptibility testing (AST) was performed using CLSI M45 and EUCAST guidelines. DNA was extracted from pure colonies of multidrug-resistant (MDR) V. cholerae and subjected to whole-genome sequencing (WGS) for genomic characterisation using an Illumina MiSeq platform. Results: Of the 350 active diarrheal cases investigated, 70 were confirmed as V. cholerae. The outbreak was initially reported in Dir and was subsequently followed by a high incidence of cholera in the Peshawar district of KPK. All strains belong to the Ogawa serotype, which shows high antibiotic resistance, particularly to ampicillin (n = 62, 88.57%), Sulfamethoxazole/Trimethoprim (n = 60, 85.71%), Erythromycin (n = 59, 84.29%), and Tetracycline (n = 53, 75.71%). The lowest resistance was against Meropenem (n = 1, 1.4%), followed by amikacin (n = 7, 10.0%) and levofloxacin (n = 13, 18.57%). Furthermore, 34 (48.57%) of the isolates were MDR, while 13 (18.57%) were extensively drug-resistant. Six samples were selected for whole-genome sequencing. The selection of six V. cholerae samples for WGS was based on their drug resistance pattern and origin of isolation. At the genomic level, all sequenced V. cholerae strains harboured multiple antimicrobial resistance determinants. Quinolone resistance was associated with mutations and genes in gyrA, gyrB, parC, and parE; resistance to sulfamethoxazole–trimethoprim with folA, folP, and dfr; tetracycline resistance with tetA and tet35; chloramphenicol resistance with catB and S10p; and aminoglycoside resistance with hns, S12p, and gigB. In addition, β-lactam resistance was linked to the presence of efflux and β-lactamase genes, including blaSHV and mox-3. Mutations were identified in gyrA at positions S83I, S177A, and S202A, and in parC at positions S85L and I231V. Collectively, the presence of these resistance determinants likely enables V. cholerae to survive exposure to high concentrations of multiple antibiotics. Conclusions: Our V. cholerae isolates showed close genetic relatedness to previously sequenced strains from Pakistan (2010 and 2022), as well as to recently reported international strains from the USA, Australia, and China. These findings highlight both the long-term persistence of these lineages within Pakistan and their international dissemination, likely facilitated by globalisation. Full article

Background: Bicuspid aortic valve (BAV) is the most common congenital heart defect, and its complications (namely, dilatation of the thoracic ascending aorta) raise concerns regarding the proper timing of aortic surgery. The study aim is to unravel the genetic basis of BAV and its complications through a high-throughput sequencing (HTS) approach and segregation analysis if family members were available. Methods: Fifty-two Italian BAV patients were analyzed by HTS using the Illumina MiSeq platform. Targeted sequencing of 97 genes known to be or plausibly associated with connective tissue disorders or aorthopathy was performed. Thirty-five first-degree relatives of N = 10 probands underwent mutational screening for variants identified in the index cases. Results: HTS identified 194 rare (MAF < 0.01) variants in 63 genes. Regarding previously reported genes, five NOTCH1 variants in four BAV patients, four FBN1 variants in two patients and one GATA5 variant in one patient were identified. Interestingly, among further loci, the possible contribution of PDIA2, LRP1 and CAPN2 was suggested by (a) the increased prevalence of rare genetic variants, independently from their ACMG classification in the whole BAV cohort, and (b) segregation analyses of variants identified in family members. Moreover, the present data also suggest the possible contribution of rare variants to BAV complications, specifically MYLK in aortic dilatation, CAPN2 in BAV calcification and VHL and AGGF1 in valve stenosis. Conclusions: Our results underline clinical and genetic diagnosis complexity in traits considered monogenic, such as BAV, but characterized by variability in disease phenotypic expression (incomplete penetrance), as well as the contribution of different major and modifier genes to the development of complications. Full article

Background/Objectives: The pharyngeal microbiota plays a critical role in respiratory health by supporting immune modulation, colonization resistance, and metabolic functions. Disruptions in this microbial ecosystem are associated with respiratory diseases; however, standard diagnostics often target individual pathogens, overlooking overall microbial dynamics. This study investigates the composition and diversity of the pharyngeal microbiota in three populations: individuals with pre-COPD (with and without concurrent acute respiratory infection [ARI]) and those with stable COPD. Methods: Pharyngeal swabs were analyzed using 16S rDNA sequencing on the Illumina MiSeq platform. Taxonomic and functional profiles were generated with MicrobAT^®, while microbial diversity was evaluated using the Shannon index and PERMANOVA. Group differences in microbiota composition were assessed via Kruskal–Wallis tests and robust PCA. The sample size was estimated at 8 subjects per group to detect significant differences (α = 0.05, 80% power, SD ≈ 20). Results: Twenty-nine swabs were collected: 11 from pre-COPD subjects (PC), 9 from ARI patients receiving antibiotics, and 9 from individuals with stable severe COPD. Microbial diversity was preserved in the PC group (100%) but markedly reduced in ARI (25%) and COPD (15%). Microbiota composition differed significantly across groups (R² = 0.371, p = 0.001), particularly at the phylum level. Functional analysis revealed minimal deficits in PC (<10%) but major impairments in ARI (81%) and COPD (56%), indicating reduced microbial functional capacity. Conclusions: Broad-spectrum microbial analysis highlights the importance of assessing pharyngeal microbiota beyond traditional pathogen detection, offering potential for innovative diagnostic and therapeutic approaches. Full article

This study evaluated how different additives—Lactiplantibacillus plantarum (LP), Lentilactobacillus buchneri (LB), and a composite enzyme (CE)—affect the fermentation quality, nutritional value, and microbial community of Leymus chinensis silage. Fresh forage was wilted to 65% moisture, treated with additives (dissolved in distilled water), and vacuum-sealed in polyethylene bags for 60 days of ensiling. Fermentation parameters and nutritional composition were analyzed using standard methods (e.g., HPLC for organic acids, Kjeldahl for crude protein), and the microbial community was profiled via Illumina MiSeq sequencing of the 16S rRNA gene V3-V4 region. Data were subjected to one-way ANOVA and Duncan’s test in SAS. All additives significantly improved key fermentation parameters (p < 0.05). The LP treatment yielded the most favorable profile, with the lowest pH (4.26) and the highest lactic acid (6.52 g/kg DM) and acetic acid (2.58 g/kg DM) contents. LP also best preserved nutrients, showing the highest dry matter (581.62 g/kg FW), water-soluble carbohydrates (24.76% g/kg DM), and crude protein (7.09% DM) (p < 0.05). The CE treatment most effectively degraded fiber, resulting in the lowest acid detergent fiber (428.87% g/kg DM) and neutral detergent fiber (628.43% g/kg DM) (p < 0.05). Additives significantly reduced bacterial alpha-diversity but enriched beneficial phylum such as Bacillota and genus such as Lentilactobacillus spp. LB), while suppressing harmful genera. Correlation analysis confirmed LP was positively correlated with lactic acid and water-soluble carbohydrates (p < 0.0001). In conclusion, additives, particularly LP, enhance silage quality by modulating the microbial community. Full article

The oral cavity contains several microbial niches, including saliva, dental plaque and tongue coating, each shaped by distinct local environments and host factors. This study compared the ecological and functional characteristics of the microbiomes of these three oral sites within the same individuals and examined host conditions associated with their variation. Saliva, supragingival plaque and tongue coating samples were collected simultaneously from 31 adults without clinical oral lesions. The bacterial 16S rRNA gene (V3–V4 region) was sequenced using the Illumina MiSeq platform, and analyses included α and β diversity, Mantel correlations, differential abundance tests, network analysis and functional prediction. The three sites displayed a clear ecological gradient. Saliva and tongue coating were taxonomically similar but were influenced by different host factors, whereas plaque maintained a distinct, biofilm-like structure with limited systemic influence. Functional divergence was most pronounced on the tongue coating despite its taxonomic similarity to saliva, whereas functional differences between saliva and plaque were modest despite larger taxonomic separation. These findings indicate that microbial composition and function vary independently across oral niches and support the need for multi-site sampling to more accurately characterize oral microbial ecology. Full article

The endophytic bacterium Bacillus subtilis 10-4 is a potent bioinoculant, previously shown to enhance growth and resilience to abiotic/biotic stresses across various crops. However, the genetic basis underlying these beneficial traits remains unexplored. In this study, a whole-genome analysis of B. subtilis 10-4 was performed to gain the molecular determinants of its plant-beneficial effects. The Illumina MiSeq-based assembly revealed a genome of 4,278,582 bp (43.5% GC content) distributed across 19 contigs, encoding 4314 predicted protein-coding sequences, 42 tRNAs, and 6 rRNAs. This genomic architecture is comparable to other sequenced B. subtilis strains. The genomic annotation identified 331 metabolic subsystems with a total number of 1668 functions, predominantly associated with amino acid (281) (16.9%) and carbohydrate (247) (14.9%) metabolism. In silico genomic analysis uncovered a diverse repertoire of genes significant for plant growth and stress resilience. These included genes for colonization (i.e., exopolysaccharide production, biofilm formation, adhesion, motility, and chemotaxis), nutrient acquisition (i.e., nitrogen, phosphorus, iron, potassium, and sulfur metabolisms), and synthesis of bioactive compounds (auxins, salicylic acid, siderophores, gamma-aminobutyric acid, vitamins, and volatiles) and antimicrobials. The latter was supported by identified biosynthetic gene clusters (BGCs) for known antimicrobials (100% similarity) bacilysin, bacillaene, subtilosin A, and bacillibactin, as well as clusters for surfactin (82%), fengycin (80%), and plipastatin (46%), alongside a unique terpene cluster with no known similarity. Additionally, genes conferring abiotic stress tolerance via glutathione metabolism, osmoprotectants (e.g., proline, glycine betaine), detoxification, and general stress response were identified. The genomic evidence was consistent with observed plant growth improvements in laboratory assays (radish, oat) and a field trial (wheat) upon 10-4 inoculation. Thus, the findings elucidate the genomic background of B. subtilis 10-4’s beneficial effects, solidifying its potential for utilization as a bioinoculant in sustainable crop production under changing climate accompanied by multiple environmental stresses. Full article

Kentucky bluegrass powdery mildew, caused by the fungus Blumeria graminis f. sp. poae, is a destructive disease affecting Poa pratensis L. In this study, endophytic bacteria were isolated from the resistant Kentucky bluegrass cultivar ‘Taihang’. Employing a combination of conidia germination inhibition assays and control efficacy tests, the biocontrol endophytic bacterial strains were screened. The impact of inoculation with the powdery mildew pathogen and biocontrol endophytic bacteria on the difference in endophytic bacterial community in the leaves of Kentucky bluegrass were studied via Illumina Miseq high-throughput 16S ribosomal RNA gene sequencing technology. A total of 18 endophytic bacterial isolates were obtained from ‘Taihang’, belonging to 3 phyla: Proteobacteria (3 isolates), Actinobacteria (6 isolates), and Firmicutes (9 isolates). The conidia germination assay revealed that isolates 6213 (Bacillus sp.) and 718 (Neobacillus sp.) exhibited the strongest inhibitory against Blumeria graminis f. sp. poae, with inhibition rate exceeding 80%. Isolate 718 exhibited superior control efficacy over strain 6213. A concentration of 10⁹ colony-forming units per milliliter (CFU/mL) was the most effective in suppressing powdery mildew on Kentucky bluegrass. The abundance of Proteobacteria on Kentucky bluegrass after the application of isolate 718 may enhance the resistance of Kentucky bluegrass to powdery mildew, and the dominant endophytic bacterial communities were Burkholderiales, Burkholderiaceae and Cupriavidus, indicating that the application of isolate 718 modulated the plant’s response to powdery mildew infection. These results demonstrate that isolate 718 enhanced the resistance of Kentucky bluegrass against powdery mildew by reshaping the endophytic bacterial community within the leaves. These findings provide molecular insights into plant−pathogen−endophytic bacteria interactions and support the development of sustainable strategies, eco-friendly strategies for plant diseases management. Full article

Emerging evidence indicates a high rate (>10%) of drug resistance (DR) associated with integrase strand transfer inhibitors (INSTIs) in developed countries, although there is limited information on DR during INSTI treatment in Uganda. With the increased use of INSTIs as standard first-line treatment, monitoring for DR using next-generation sequencing (NGS) has become essential. NGS can detect the lower-frequency variants that may be missed by traditional Sanger sequencing (SS). This study evaluates the diagnostic accuracy of next-generation sequencing (NGS) compared to Sanger sequencing for detecting HIV-1 INSTI resistance mutations and estimates the prevalence and factors associated with drug resistance among adults with virologic failure on INSTI-based regimens in Uganda. Utilizing the Illumina MiSeq platform for NGS, data was analyzed using STATA V.18 and a logistic regression model at 5% level of significance. This study demonstrates that NGS achieved 100% sensitivity, specificity, positive predictive value, negative predictive value, and overall accuracy in detecting major mutations. NGS identified INSTI DRMs in 4% of adults at a ≥20% threshold and was able to detect both high- and low-abundance variants, which could have important implications for clinical outcomes. This study emphasizes the need for HIVDR testing before antiretroviral therapy (ART) initiation, given the increasing use of INSTIs. We recommend that healthcare providers adopt more sensitive diagnostics such as NGS and use detailed resistance profiles to tailor antiretroviral therapies. This approach is critical for effectively managing and preventing drug-resistant HIV strains. Full article

In order to clarify the changes and correlations among microbial community structure and soil environmental factors in the rhizosphere soil of peppers under healthy and diseased conditions, Illumina MiSeq technology was used to perform high-throughput sequencing of the V3-V4 region of the bacterial 16S rRNA gene and the ITS hypervariable region of fungi in the rhizosphere soil of peppers. The dominant species and key environmental factors affecting the occurrence of pepper Phytophthora blight were analyzed and screened, and the functions of bacteria and fungi in the samples were predicted by PICRUSt2 and FUNGuild. The results showed that except for soil pH, the contents of microbial biomass carbon, magnesium, zinc, and iron in the rhizosphere soil of healthy peppers were significantly higher than those in the diseased soil. Alpha diversity analysis showed that the diversity index of the bacterial community in healthy soil was higher than that in diseased soil, while the diversity index of the fungal community was significantly lower than that in diseased soil. The relative abundance of beneficial bacteria such as Proteobacteria, Actinobacteriota, Burkholderiales, and Rhodanobacteraceae in the rhizosphere soil of healthy peppers was higher. Pathogens such as Penicillium and Fusarium were significantly enriched in the rhizosphere soil of diseased pepper plants. The functional prediction results showed that soil bacteria were mainly metabolized, including the biosynthesis of ansamycin, the biosynthesis of vancomycin antibiotics, the biosynthesis of valine, leucine, and isoleucine, the metabolism of C5-branched dicarboxylic acid, and the biosynthesis of fatty acids. The main nutritional strategies of the fungal community are disease prototype and saprophytic. Combined with the key environmental factors, microbial composition, and correlation analysis of pepper rhizosphere soil, it is speculated that the occurrence of pepper Phytophthora blight may be related to the synergistic effect of soil nutrients and microbial flora, which provides a theoretical basis for the biological control of pepper Phytophthora blight in the future. Full article

The growing demand for precision diagnostics in cystic fibrosis and other genetic disorders, such as cancers, is driving the need for sequencing platforms that combine analytical robustness, scalability, and cost-efficiency. In this study, we performed a direct comparison between two leading Next-Generation Sequencing (NGS) platforms, MiSeq (Illumina, CA, USA) and DNBSEQ-G99RS (MGI Tech Co., Shenzhen, China), using a CE-IVD-certified CFTR panel (Devyser AB), selected for its complexity and variant spectrum, including SNVs, CNVs, and intronic polymorphisms. A total of 47 genomic DNA samples from routine clinical activity were analyzed on both platforms. Illumina sequencing covered all CFTR variants using standard workflows, while MGI data were generated from residual diagnostic DNA, with informed consent. Sequencing data were processed using Amplicon Suite v3.7.0 for variant calling, annotation, and ACMG classification. Quality control metrics and platform-specific parameters were also evaluated. Both platforms demonstrated complete concordance in variant detection, including SNVs, CNVs, and complex alleles (e.g., Poly-T/TG). Illumina exhibited slightly superior basecalling quality and allelic frequency uniformity, while MGI achieved higher sequencing depth (mean ~2793×) and demultiplexing efficiency. No false positives, false negatives, or discordant HGVS annotations were observed. The use of full-gene CFTR sequencing enabled granular and technically rigorous cross-platform validation. These findings confirm the analytical equivalence of Illumina and MGI for diagnostic genotyping. Moreover, MGI’s greater data output and flow cell capacity may offer tangible advantages in high-throughput settings, including somatic applications such as liquid biopsy and molecular oncology workflows. Full article