In the present study, an exopolysaccharide (EPS)-producing bacterial strain was isolated and identified as Sphingobium yanoikuyae SS02 using 16S rRNA gene sequencing. Optimization of the culture medium through a one-factor-at-a-time (OFAT) approach enhanced EPS yield to 6.5 ± 0.2g/L under mesophilic conditions. The purified EPS was structurally and functionally characterized using UV-Visible spectroscopy, Fourier Transform Infrared Spectroscopy (FTIR), X-ray Diffraction (XRD), Nuclear Magnetic Resonance (NMR), and Thermogravimetric Analysis (TGA). Spectroscopic analyses confirmed a glucose-rich heteropolysaccharide backbone with diverse functional groups, exhibiting moderate crystallinity and thermal stability, with major decomposition occurring between 150°C and 330°C. Biological evaluation revealed that the EPS possessed concentration-dependent antioxidant activity, with an IC50 of 0.635mg/mL in the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay and 92.5% and 78.3% inhibition for ascorbic acid and EPS, respectively, in the phosphomolybdenum assay. The EPS also exhibited notable anti-inflammatory activity with 72.57% inhibition at 800µg/ml, and broad-spectrum antibacterial activity against Staphylococcus aureus, Klebsiella pneumoniae, Bacillus subtilis, and Escherichia coli, with the highest sensitivity observed in S. aureus. These findings demonstrate the multifunctional bioactivity and biocompatibility of S. yanoikuyae SS02-derived EPS, suggesting its potential as a natural therapeutic agent for pharmaceutical and biomedical applications.

Standardization of oral sample collection methods is essential for accurate and reproducible microbiota quantification. This methodological study aimed to evaluate different oral collection methods to identify the most consistent approach for bacterial quantification by qPCR using samples from adolescent individuals. In addition, to assess the biological applicability of the best method, an exploratory analysis compared bacterial profiles between eutrophic and overweight/obese adolescents and explored associations between bacterial abundance and body composition parameters. Samples of unstimulated saliva, cheek swabs, and biofilm were collected from the same individuals, and qPCR was used to quantify total bacteria (16S rRNA gene), Bacillota, and Bacteroidota phyla. Unstimulated saliva produced the lowest variability in bacterial quantification compared with other methods (p < 0.05). Moderate correlations were observed between saliva and biofilm, whereas saliva and cheek swab showed weak associations. Although bacterial copy numbers tended to be higher in overweight/obese individuals, these differences were not statistically significant. Correlation matrices suggested group-specific associations between bacterial taxa and body composition parameters, demonstrating the potential of saliva for microbiome assessment in studies of nutritional and metabolic health. This study validated unstimulated saliva as a reproducible, non-invasive, and cost-effective biofluid for oral microbiota quantification by qPCR. The method provides consistent results suitable for large-scale, translational, or point-of-care applications.

This cross-sectional study aimed to compare the endodontic microbiome assessed from root canals of teeth associated with either symptomatic or asymptomatic apical periodontitis and analysed by 16S rDNA gene sequencing. 60 teeth presenting clinical and radiographic signs of symptomatic or asymptomatic apical periodontitis (n = 30) were included in this cross-sectional study after participants had given their written informed consent. After isolation with rubber dam, disinfection and access cavity preparation, glide paths were prepared using C-Pilot Files and K-Files under electronic root canal length control. Microbial samples were collected from a total of 120 root canals (symptomatic apical periodontitis, SAP: n = 62, asymptomatic apical periodontitis, AAP: n = 58) each with a sterile file (size 20/0.06) in a single length technique. Only one specimen per tooth was included in the analysis; in multi-rooted teeth, the specimen with highest sequencing depth. After DNA extraction, the hypervariable region V4 of the bacterial 16 S rRNA gene was amplified and sequenced (Illumina MiSeq). Taxonomy was assigned based on the expanded Human Oral Microbiome Database (eHOMD). Statistical analysis of diversity parameters comprised Mann-Whitney U tests and PERMANOVA. Compositional differences were evaluated by differential abundance analyses using DESeq2, LinDA, and ANCOM-BC2 methods. No differences were observed in richness and diversity (Shannon diversity index) on the genus or ASV level (p > 0.05). According to PERMANOVA, SAP and AAP microbiomes did not differ significantly both on genus and ASV levels (p > 0.05). Among highly abundant genera, Fusobacterium was indicated to be more abundant in SAP samples whereas Actinomyces was more abundant in AAP samples. The expression of clinical symptoms in apical periodontitis does not appear to be determined by specific microorganisms but may instead reflect shifts of the relative abundance of the microbial community.

Anaplasma species are globally distributed and pose significant public health threats as zoonotic pathogens. However, the epidemiological characteristics and genetic diversity of Anaplasma in rodents from Fujian Province, southeastern China, remain poorly understood. From 2015 to 2024, we systematically collected 966 rodents across 22 counties and cities in Fujian. Screening was performed using real-time PCR and nested PCR targeting the groEL and 16S rRNA genes. Positive samples were subjected to sequencing, followed by phylogenetic analysis (MEGA11) and haplotype network construction (PopART). The overall infection rate of Anaplasma was 4.35% (42/966), and all positive samples were identified as Anaplasma phagocytophilum. Infection rates showed significant differences among rodent species, cities, habitat types, and sampling years (P < 0.05). Haplotype network analysis indicated that H1 was the dominant haplotype. This study confirms the circulation of A. phagocytophilum in rodents in Fujian, reveals its association with rodent age, seasonal and habitat factors, and underscores the necessity for enhanced ongoing surveillance and control of rodent-borne Anaplasma in this region.

Metabolic dysfunction-associated steatotic liver disease (MASLD, formerly non-alcoholic fatty liver disease) is a prevalent and progressive condition closely linked to gut microbiota composition. Fecal microbiota transplantation (FMT) may help restore a health-associated microbiome, but its efficacy is often limited by inconsistent engraftment of beneficial taxa. Prebiotics may selectively support keystone microbes associated with reduced MASLD risk. This study evaluated two prebiotics, inulin and xylooligosaccharides (XOS), for their ability to modulate the microbiota of healthy FMT donors in an in vitro gut model, focusing on enriching beneficial taxa and functions associated with MASLD resilience. Stool from eight clinically qualified FMT donors was cultured anaerobically for 24 hours with or without prebiotics. Microbiota composition was assessed by 16S rRNA gene sequencing and short-chain fatty acid (SCFA) concentrations were measured using nuclear magnetic resonance. Functional potential was inferred using predictive metagenomic analysis. Prebiotic responses were highly donor-specific, yet both inulin and XOS consistently enriched Bifidobacterium and Bacteroides-genera associated with SCFA production and metabolic health. XOS preferentially enriched Lactobacillus and Parabacteroides, while inulin enhanced Holdemanella and Mediterraneibacter. Functional pathways relevant to MASLD pathophysiology were enriched, including carbohydrate metabolism, vitamin biosynthesis, fatty acid metabolism, and tryptophan degradation. Both prebiotics significantly increased acetate levels, while butyrate showed a donor-dependent increasing trend. These findings suggest that prebiotic supplementation can selectively enrich MASLD-relevant microbial taxa and functions in donor-derived FMT material, supporting their potential as adjuvants to enhance the efficacy and disease-specificity of FMT interventions for MASLD.

Density fluctuations of Burkholderia and Achromobacter in cystic fibrosis sputum.

Sweet potatoes were cultivated in hydroponics with low nitrogen fertilization. Through 16S rRNA gene amplicon sequencing, the most abundant genera in storage roots of them were identified as Azospirillum, Sphingomonas, Burkholderia, Bradyrhizobium, and Arthrobacter, all of which have been reported as nitrogen-fixing bacteria. These results suggest that nitrogen-fixing bacteria may selectively increase to compensate for the insufficient nitrogen fertilization.

Typhoid fever is a potentially fatal systemic infection caused by Salmonella typhi (S. typhi), a Gram-negative, rod-shaped, facultative anaerobe. This study aimed to characterize the efflux pump genes, molecular mutations, and antibiotic susceptibility patterns in clinical isolates. A total of 2950 blood samples were collected from suspected typhoid patients. Of these, 380 (12.88%) bacterial isolates were recovered, of which 236 (62.10%) were Gram-negative. S. typhi was detected in 95 isolates (25% of all bacterial isolates), corresponding to an overall prevalence of 3.22%. Identification was performed using API 20-E strips and partial 16S rRNA gene sequencing. Resistance profiling classified the isolates into multidrug-resistant (MDR, 13.68%) and extensively drug-resistant (XDR, 86.31%) categories. High resistance was observed to ampicillin (98.94%) and chloramphenicol (93.68%), whereas no resistance was detected to azithromycin or meropenem. Minimum inhibitory concentration (MIC) testing using E-Strips showed the highest MIC values for trimethoprimsulfamethoxazole (MIC50: 0.125 μg/ml, MIC90: 0.50 μg/ml) and the lowest for ciprofloxacin (MIC₅₀: 0.008 μg/ml, MIC₉₀: 0.023 μg/ml). Among the efflux pump genes, mdtB (32.63%) was the most prevalent, followed by mdtC (30.52%) and mdtA (29.47%). Multiple mutations were identified and evaluated using the I-Mutant 2.0 server. Protein structures were modeled using trRosetta and Discovery Studio. Phylogenetic analysis using MEGA-11 revealed evolutionary relationships with other Salmonella enterica serovars. A serious public health issue that increases mortality and morbidity rates worldwide is antibiotic resistance. Among the mechanisms underlying antibiotic resistance in bacteria are degradation of antibiotics and/or modification of enzymes, changing drug targets to alter the affinity of antibiotics, altering bacterial surfaces by changing the expression of external membrane proteins, and active bacterial efflux of drugs. The drug's binding affinity can be changed by mutations in efflux pump genes that change the shape of the substrate- binding pocket. These structural alterations may improve the pump's capacity to identify and eliminate antibiotics, reducing intracellular drug levels and increasing resistance to several drugs. This study highlights the alarming rise of multidrug resistance in S. typhi isolates and identifies key genetic mutations that may influence efflux pump function, potentially contributing to enhanced antibiotic resistance.

Calodium hepaticum is a neglected zoonotic parasite of rodents, primarily affecting rats. Highly sensitive molecular assays such as nested PCR are not available to screen the parasite in rat, man and dog. In the present study, parasite specific nested PCR primers were designed to amplify 171bp partial 18S rRNA gene of C. hepaticum and compared with an existing semi nested PCR. Both nested PCR and semi nested PCR assays were sensitive enough to detect at least 10 C. hepaticum eggs in rat liver samples. The limit of detection of nested PCR assay was 420 zM, and it was 35-fold more sensitive than that of semi nested PCR assay. The nested and semi nested PCR assays specifically amplified C. hepaticum egg DNA from human and dog DNA samples spiked with parasite DNA. The molecular prevalence of C. hepaticum in household rats in Chennai was 41.81% based on nested PCR assay. This study suggests that the 18S rRNA gene based nested PCR assay could be a more sensitive detection system for molecular screening of C. hepaticum in rat liver samples and highly suitable for epidemiological studies. Further, both the 18S rRNA gene based nested and semi nested PCR assays can potentially used for detection of C. hepaticum infection in man and dog.

Iron deficiency anemia (IDA) is a common nutritional deficiency disease caused by iron deficiency. Oral iron supplementation, the simplest and most commonly used iron repletion strategy in the clinic, primarily induces gastrointestinal inflammation, which subsequently elicits complications, including nausea, gastrointestinal bleeding, and constipation. Oral iron supplementation, the most simple and widespread method of iron replenishment in the clinic, primarily causes gastrointestinal inflammation, which, in turn, leads to complications such as nausea, gastrointestinal bleeding, and constipation. Furthermore, gastrointestinal inflammation impedes the absorption of iron, thereby exacerbating anemia. Gold nanoparticles with an inherent anti-inflammatory effect make them a promising weapon for alleviating iron-induced gut inflammation. Herein, we studied the effects of 4,6-diamino-2-pyrimidinethiol (DAPT)-functionalized gold nanoparticles (DAu NPs) on the iron supplementation efficiency and gut inflammation in the IDA model. We discussed the mechanisms of gut microbiota and immune responses on gut inflammation using the 16S rRNA (rRNA) gene sequencing and the polarization of RAW 264.7 cells in vitro. DAu NPs with oral iron supplementation could effectively treat IDA. DAu NPs played a significant role in reshaping gut microbiota, promoting short-chain fatty acid production, and regulating immune responses to reduce inflammation caused by excess iron. In vitro, DAu NPs could inhibit iron-dependent bacteria (Escherichia coli) proliferation while promoting probiotic (Lactobacillus) growth. Oral administration of DAu NPs could regulate M2 polarization of gut macrophages, reduce neutrophil and Th17 cell infiltration, and increase Treg cells recruitment. DAu NPs accumulated primarily in the colon and were excreted via feces, demonstrating excellent biosafety. Our study provides a potential method for the treatment of IDA and other metal element deficiencies.

Oligo-cyanobacterial microalgae-bacteria granular sludge for mitigating cyanotoxin risk: Cultivation, characteristics, and formation mechanism.

Very few starter cultures were specifically developed for African cereal foods. This study aimed to develop starter cultures for cereal fermentations that can be stored at ambient temperature. Model mahewu fermentations were conducted with two starter culture combinations. The starter cultures competed at 15 °C and 25 °C with strain cocktails of finger millet malt isolates that were formulated based on the bacterial abundance in finger millet malt. Fermentations were analysed by plating and sequencing of 16S rRNA gene amplicons. Without starter cultures, Enterobacteriaceae were dominant members of the microbial community for 24 h (25 °C) and 72 h (15 °C), respectively. Because 16S rRNA gene amplicon sequencing identified Salmonella spp. with more than 0.1% abundance in all six finger millet malt samples, the persistence of Enterobacteriaceae at low temperatures represents a risk for food safety. At 25 °C, the starter cultures Limosilactobacillus fermentum and Lactiplantibacillus plantarum dominated the microbial community. At 15 °C, Lp. plantarum and Weissella confusa were dominant. Starter cultures were produced by air-drying and by freeze-drying and stored at 4 °C and 20 °C. Results provide an alternative approach to the production of starter cultures.

To develop and validate an ecology-driven strategy that leverages natural manure-soil depth gradients as a screening system for the targeted isolation of nutrient-solubilizing bacteria (NSB) with high biofertilizer potential. A full-factorial sampling design was implemented across gradients of distance-from-manure (5 points, 8-m intervals) and soil depth (0-20, 20-40, 40-60 cm) in a coconut plantation. Culturable bacteria were isolated using a culture-dependent approach on a nutrient-rich medium, identified via 16S rRNA gene sequencing, and functionally screened in vitro for nitrogen fixation, phosphate solubilization (PS), and potassium solubilization (KS) capabilities. Manure input and soil depth interacted to form a heterogeneous soil nutrient landscape, with available phosphorus (AP) identified as the most influential environmental factor shaping the bacterial community. Phosphate-solubilizing bacteria (PSB) were significantly enriched in low-P habitats, verifying the niche-based selection of functional bacteria. This gradient-based screening strategy enabled the targeted recovery of multifunctional NSB strains (e.g., Klebsiella and Enterobacter) with concurrent nitrogen fixation, phosphate and potassium solubilization capacities, which were isolated from specific microhabitats including deep, nutrient-depleted soil layers. This study demonstrates that intersecting manure and soil depth gradients form a powerful, predictable natural screening system for the targeted isolation of beneficial bacteria. This ecology-driven strategy effectively links microbial ecology to bioprospecting. It provides a curated library of isolates with defined ecological origins and a predictive framework for developing customized biofertilizers, thereby enhancing microbial resource mining efficiency and contributing to sustainable agriculture.

Human gut microbiota is associated with obesity. Gut microbiota-derived extracellular vesicles (EVs), lipid coated nanoparticles secreted by bacteria, have been suggested as a communication mechanism between gut microbiota and the host. This study characterized the effect of Roux-en-Y gastric bypass (RYGB) on gut microbiota and gut microbiota-derived EVs in patients with obesity. Fecal samples were collected from 30 recruited patients at baseline and 6 months after surgery. EVs were isolated from fecal samples, and their origin and protein content were analyzed. The number of unique proteins was increased in gut microbiota-derived EVs after the surgery as compared to baseline. A significant difference in both microbiota composition (p = 0.001; PERMANOVA) and microbiota-derived EVs (p = 0.001; PERMANOVA) was observed in response to surgery. Based on 16S rRNA gene sequencing data, a random forest classifier accurately classified both gut microbiota (AUC = 0.93) and EVs (AUC = 0.80) to baseline and after surgery groups. This study found that gastric bypass surgery altered both the composition and characteristics of gut microbiota and gut microbiota-derived EVs in patients with obesity. Thus, gut microbiota-derived EVs may play a role in obesity and influence the health effects of bariatric surgery beyond the gut. Trial Registration: ClinicalTrials.gov identifier: NCT00950003.

Gut microbiota modulates the effects of host-derived fecal microRNAs on cultured gut microbiota in mice.

Identification of gut microbiota features of diarrheic calves using the full-length 16S rRNA gene amplicon sequencing and machine learning.

This study aimed to isolate lactic acid bacteria (LAB) from the intestines of whiteleg shrimp ( Litopenaeus vannamei ) for potential probiotic use and to evaluate the effects of dietary synbiotic supplementation on shrimp growth performance and digestive enzyme activity. A total of 249 LAB isolates were obtained from shrimp intestines collected in extensive ponds in Ca Mau province. Among these, 15 isolates exhibited strong antagonistic activity against Vibrio parahaemolyticus , with inhibition zones ranging from 6 to 11 mm. Most isolates were Gram-positive, non-motile, non-spore-forming, and exhibited cell shapes ranging from short rods to spherical. Four isolates demonstrated strong protease and leu-aminopeptidase activities, although α-amylase production was relatively low. Strain LAB8.1CM was identified as Lactiplantibacillus plantarum based on 16S rRNA gene sequencing (99.7% homology) and exhibited promising probiotic characteristics, indicating its potential as a safe feed additive. In a 60-day feeding trial, shrimp were assigned to five dietary treatments: control (basal diet), prebiotic (0.2% banana extract), probiotic ( L. plantarum 10 8 CFU/kg), synbiotic-0.2% (banana extract 0.2% + L. plantarum 10 8 CFU/kg), and synbiotic-0.4% (banana extract 0.4% + L. plantarum 10 8 CFU/kg), each with three replicates. Synbiotic supplementation significantly improved digestive enzyme activities, including α-amylase, β-galactosidase, protease, and leu-aminopeptidase. Growth performance parameters, such as weight gain, daily weight gain, specific growth rate, and feed conversion ratio, were also significantly enhanced, particularly in shrimp fed the synbiotic-0.2% diet. Survival rates did not differ significantly among treatments. These findings indicate that dietary supplementation with selected LAB strains, in combination with prebiotics, can effectively enhance growth, digestion, and health in whiteleg shrimp, highlighting their potential as synbiotic feed additives for sustainable aquaculture practices.

Microbial pollution disables the chemical defenses of sea fans.

Two novel actinomycete strains, AG03ᵀ and ODS28ᵀ, were isolated from worker ants Anochetus graeffei and Odontomachus simillimus, respectively, collected in Thailand. Strain AG03ᵀ exhibited the highest 16S rRNA gene sequence similarity (98.28%) to Nocardia grenadensis NBRC 108939ᵀ, whereas strain ODS28ᵀ showed the highest similarity (97.73%) to Actinacidiphila bryophytorum NEAU-HZ10ᵀ. Chemotaxonomic analysis of AG03ᵀ revealed the presence of meso-diaminopimelic acid in the cell wall, glucose and mannose as whole-cell sugars, and MK-8(H₄ω-cycl) as the predominant menaquinone. In contrast, ODS28ᵀ contained LL-diaminopimelic acid, glucose, ribose, and xylose as whole-cell sugars, with MK-9(H₈) and MK-9(H₆) as the predominant menaquinones. Whole genome sequence-based relatedness values, including average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH), supported the classification of both strains as representing novel species. Based on polyphasic taxonomic evidence, strains AG03ᵀ and ODS28ᵀ are proposed as the type strains of Nocardia anocheti sp. nov. (AG03ᵀ = TBRC 16206ᵀ = NBRC 115864ᵀ) and Streptomyces odontomachicola sp. nov. (ODS28ᵀ = TBRC 18345ᵀ = NBRC 116641ᵀ), respectively. The genomes of both strains contained biosynthetic gene clusters (BGCs) potentially responsible for the production of bioactive secondary metabolites. These findings suggest that social insects, such as ants, represent a promising source of novel actinomycetes.

The development of beekeeping in Ecuador has generated the need to strengthen the bee health program. Research on the main pathogens responsible for diseases like nosemosis, which can severely impact bee health, is of special interest. This study aims to identify the Nosema apis and/or Nosema ceranae species infecting honey bee colonies located in the northern Andean region of Ecuador using multiplex PCR targeting the RNA polymerase II gene (RPB1), and the phylogenetic analysis of N. ceranae based on the 16 S rRNA gene sequences. Among the 164 honey bee samples collected from colonies in the provinces of Carchi, Imbabura, and Pichincha, the prevalence of Nosema apis and Nosema ceranae was 14.63% and 21.34%, respectively. Phylogenetic analysis showed that N. ceranae from Ecuador is closely related to the sequences from Argentina and Brazil. These findings provide the first molecular confirmation of N. ceranae in Ecuador and support the need for molecular monitoring of honey bee pathogens in the region.