Search Results (399)

Bacterial cellulose (BC) is an emerging biopolymer for skin tissue regeneration; however, its functionalization with natural antimicrobial agents remains limited. This study reports the preclinical evaluation of a BC-based dressing for diabetic wounds. BC membranes were obtained from mango pulp agro-waste by Komagataeibacter xylinus cultivation (6.32 g/L) and functionalized with grapefruit seed oil (GSO) at three v/v ratios (1:100, 1:200 and 1:500). FTIR spectroscopy confirmed GSO incorporation into the BC matrix through physical interactions, with a dose-dependent loading. Antimicrobial activity of the BC/GSO dressings was screened against Staphylococcus aureus, Escherichia coli and Candida albicans by agar diffusion, showing dose-dependent inhibition zones. Following the minimum effective dose principle, the BC/GSO 1:500 (v/v) formulation was selected for comprehensive biocompatibility evaluation (cytotoxicity, mutagenicity, pyrogenicity and sensitization) and for in vivo wound-healing testing in a streptozotocin-induced diabetic Wistar rat model. Cell viability above 70% was achieved from membrane-extract dilution 1:100,000, while mutagenicity, pyrogenicity and sensitization assays confirmed the absence of adverse biological responses. In vivo, BC/GSO 1:500 (v/v) dressings supported wound closure comparable to nitrofurazone, with no clinical signs of infection. Overall, these results position BC/GSO dressings as a sustainable, biocompatible and antimicrobial candidate for early-stage diabetic wound regeneration and demonstrate the technical feasibility of valorizing mango pulp agro-waste into a high-value biomedical biopolymer. Full article

Chicken feet, an abundant and low-cost poultry by-product rich in collagen, were used to extract gelatin, which was then formulated into active biodegradable films containing food-grade clove essential oil (CEO), glycerol, sorbitol, and Tween 20. Gelatin extraction involved 0.5 M NaOH pretreatment followed by 5% acetic acid extraction at 66 °C, yielding 11.22% gelatin. Eight gelatin–CEO films were prepared by varying the CEO concentration and plasticizer composition. The supplier-declared CEO composition was eugenol-dominant, and antibacterial activity against Escherichia coli, Kluyvera sp., and Enterobacter cloacae was assessed by agar disk diffusion, MIC, and MBC assays, each performed in triplicate. CEO inhibition zones of 22, 14, and 19 mm were recorded against E. coli, Kluyvera sp., and E. cloacae, respectively; the blank 6 mm control disks without oil produced no inhibition halo beyond the disk edge. MIC/MBC values were 5/6, 3/4, and 4/5 mg/mL for the same three strains. All films were continuous, smooth, and peelable; sorbitol-containing formulations were clearer and more flexible than sorbitol-free variants. Water solubility ranged from 37.67% to 48.78%, opacity from 5.26 × 10⁻³ to 9.20 × 10⁻³ A500 mm⁻¹, and thickness from 11.75 to 23.75 µm. Water vapor transfer was undetectable under the gravimetric screening protocol for all formulations. All films showed complete visual disappearance in soil within 6–10 days. In the cherry tomato trial, the best-performing coatings extended acceptable storage from about 5 days (uncoated control) to 10 days at 17–20 °C. Full article

The textile industry contributes significantly to environmental pollution through massive water usage and toxic synthetic dye effluents. Bioremediation offers a sustainable solution by using microorganisms, such as bacteria, to transform complex contaminants into simpler substances. This study evaluated the bioremediation potential of fifteen halotolerant endophytic bacteria isolated from black beans (Phaseolus vulgaris L.) against various textile dyes. The strains included Bacillus cereus, Bacillus amyloliquefaciens, Priestia megaterium, and Staphylococcus warneri. Initial screenings across different TSA (Tryptic Soy Agar) medium concentrations (10%, 50%, 100%) revealed that bacterial growth and discoloration—assessed via halo formation—were most pronounced in 50% medium. While several dyes showed no reaction, Malachite Green and Congo Red were successfully decolorized. In liquid medium assays TSB (Tryptic Soy Broth) (50%) quantitative analysis via spectrophotometry showed that strains PV57, PV107, and PV112 achieved approximately 45% discoloration for Congo Red. Most notably, PV18 and PV114 achieved discoloration efficiencies of 91.69% and 88.72%, respectively, for Malachite Green after 72 h. These findings indicate that salt-tolerant endophytic bacteria are promising candidates for the decolorization of textile dyes. However, further studies are required to determine whether the observed discoloration results from biodegradation, biotransformation, or biosorption. This study underscores the potential of agricultural endophytes in managing industrial waste effectively. Full article

Background/Objectives: Given the ongoing threat of antimicrobial resistance, the identification and characterization of multidrug-resistant isolates are essential. An increase in antimicrobial-resistant bacteria has been reported in Romania, but national data are still scarce. This study aimed to evaluate beta-lactamase-producing Gram-negative bacteria (GNB) isolated over two years at a Romanian nephrology hospital, while comparing carbapenemase detection phenotypic methods. Methods: Gram-negative bacterial isolates collected between January 2022 and May 2024 that met antimicrobial resistance screening criteria were evaluated. After identification, extensive disk diffusion antibiograms were performed, read, and interpreted, complemented by testing on cloxacillin/oxacillin-supplemented Mueller–Hinton agar. The colistin minimum inhibitory concentration (MIC) was not assessed, and aztreonam–avibactam was not tested for Enterobacterales. For non-fermenter GNB, the colistin MIC was determined. Phenotypic carbapenemase production tests were performed for all strains (BlueCarba Test, CIM, mCIM, zCIM, and rCIM). Carbapenemase detection immunochromatographic tests were performed for a set of strains. Results: Among the 397 evaluated strains, 335 (84.38%) were Enterobacterales and 62 (15.62%) non-fermenter GNB, showing high antimicrobial resistance levels. Of these, 188 (47.35%) were Klebsiella pneumoniae; 139/188 (73.93%) showed carbapenem resistance and carbapenemase production; 49/188 (26.06%) produced two carbapenemases; and 45/188 (23.93%) presented resistance to all tested antimicrobials. MALDI-TOF identified 28 KPC-producing K. pneumoniae strains. Lateral flow assays revealed NDM, VIM, KPC, and OXA-48-like enzymes in 48 of 56 tested Enterobacterales; 12/48 strains produced two carbapenemases. Of the 62 non-fermenter GNB, 33 were Pseudomonas spp. and 20 Acinetobacter baumannii; one Pseudomonas spp. was susceptible only to colistin and seven only to cefiderocol; four A. baumannii were susceptible only to colistin and three only to cefiderocol. Lateral flow assays detected VIM or IMP enzymes in 13/33 Pseudomonas spp. and OXA-23 and/or OXA-40/-58 enzymes in all 20 A. baumannii. Conclusions: Among the evaluated strains, many showed resistance to multiple antimicrobial classes. Furthermore, strains co-producing two carbapenemases were identified. Full article

Prevalence of soil-transmitted helminth (STH) infections in Sri Lanka has declined from 73.3% in 1954 to <1% in 2017, leading to revised deworming policies and discontinuation of routine deworming in three districts, including Anuradhapura, in 2019. As countries approach elimination, this study assessed human intestinal nematode infection (HINI) prevalence in Anuradhapura, a low-endemic setting, using microscopic and molecular methods. From January to November 2023, 967 primary school children were recruited and screened for HINI using direct smears, Kato–Katz, formalin ether concentration, agar plate culture, scotch tape, and qPCR. Combined microscopy and molecular prevalence was 41.2% for STH and 54.1% for HINI. The most prevalent HINI was Enterobius vermicularis (191, 24.5%), followed by Strongyloides stercoralis (120, 19.2%), Ascaris lumbricoides (113, 18.1%), Trichuris trichiura (40, 6.4%), and hookworm (43, 6.9%). Against the composite reference standard, qPCR demonstrated superior diagnostic performance across all species, achieving sensitivities of 86.7% for Ascaris, 85.0% for Trichuris, and 97.7% for hookworm, consistently exceeding those of microscopy-based methods. Correspondingly, qPCR also identified the highest infection prevalences, detecting Ascaris, Trichuris, and hookworm infections in 15.7%, 5.4%, and 6.7% of participants, respectively. All infections were low-intensity as determined by Kato–Katz. Microscopy significantly underestimates STH prevalence. In post-deworming settings, integrated surveillance including molecular diagnostics is essential. The high prevalence of S. stercoralis and E. vermicularis highlights the need for expanded surveillance and targeted interventions to support sustainable control. Full article

Background/Objectives: Dogs are important carriers and transmitters of staphylococci from surface microbiota. Carriage screening allows for the identification of animals colonised with pathogens such as methicillin-resistant S. pseudintermedius (MRSP) and methicillin-resistant S. aureus (MRSA), which are spread between animals and from dogs to humans. This cross-sectional study determined the diversity of staphylococci from the surface microbiota of clinically healthy dogs in Bulgaria and their susceptibility to antimicrobial agents. Methods: The study was performed with 30 healthy dogs reared in the region of Stara Zagora, Bulgaria in 2024 and 2025. Swabs were obtained from eight body sites from each dog and incubated on blood and mannitol salt agar. Random isolates were identified by MALDI-TOF MS and tested for resistance to oxacillin/cefoxitin and to 14 classes of antimicrobial drugs (AMD). Results: Ninety out of 100 tested isolates were confirmed as Staphylococcus spp. from 15 different species. The total share of coagulase-positive (CoPS) staphylococci significantly exceeded that of coagulase-negative (CoNS) ones. Fifteen phenotypically methicillin-resistant staphylococci were identified—eight CoNS and seven CoPS—and confirmed by MIC test. The highest resistance was against penicillin (64.4%), ampicillin and minocycline (52.2%), whereas the highest sensitivity was to rifampin, amikacin, cefquinome and amoxicillin + clavulanic acid. Conclusions: Data about the carriage of MRSP, MRSA and multidrug-resistant coagulase-negative staphylococci in healthy dogs are important in view of the increased risk of colonisation/infection for people in contact with these dogs in households and veterinary facilities (clinics, hospitals). This supports the “One Health” approach integrating animal, human and environmental health. Full article

Background: Acinetobacter baumannii (A. baumannii) is a critical-priority pathogen of major concern in healthcare settings. Colistin remains a last-resort antibiotic for multidrug-resistant (MDR) A. baumannii infections; however, resistance is increasingly reported worldwide yet remains understudied in Bahrain. Reliable detection methods and understanding clonal dissemination are essential for infection control. Objectives: This study aimed to (1) determine the rate of colistin resistance in 102 clinical A. baumannii isolates from Bahrain, (2) evaluate the diagnostic performance of the colistin agar test (CAT) and E-test against broth microdilution (BMD method), and (3) assess clonal relationships using BOX-PCR fingerprinting. Methods: 102 clinical isolates from multiple hospitals in Bahrain underwent susceptibility testing via the BMD method, CAT, and E-test; screening for mcr-1 to mcr-5 genes; and BOX-PCR DNA fingerprinting. Results: Colistin resistance was detected in 14.7% of isolates by BMD method, higher than regional and global averages. All resistant isolates were mcr-negative, suggesting chromosomally mediated resistance. CAT showed 86.7% sensitivity, 98.8% specificity, and a 13.3% very major error rate. The E-test failed to detect resistant isolates (very major error 100%). BOX-PCR revealed predominant clonal relatedness with intra- and inter-hospital spread. Conclusions: Colistin resistance in A. baumannii from Bahrain exceeds regional and global levels, likely driven by chromosomal mechanisms under selective pressure. The BMD method remains the gold standard for colistin testing, while CAT may serve as a screening tool requiring confirmation. Strengthened stewardship and infection control measures are vital to contain dissemination. Full article

Marine endophytic fungi inhabit the internal tissues of seaweed, seagrass, and mangroves without causing harm. These fungi are known to produce extracellular enzymes, including proteases and cellulases, which play crucial roles in various biological processes and have potential applications in diverse industrial sectors. This study aimed to screen the enzymatic potential of marine endophytic fungi, identify selected isolates, and characterize their enzyme activities. A total of 20 fungal isolates were obtained, comprising 16 isolates from seaweed, three from seagrass, and one from mangrove leaves, collected from the coastal areas of the Seribu Islands (Jakarta), Sukabumi (West Java), Nusa Dua (Bali), and the Buton Islands (Southeast Sulawesi). Screening results showed that 50% of the isolates exhibited proteolytic activity on skim milk agar, while 40% demonstrated cellulolytic activity on carboxymethylcellulose (CMC) agar. Two isolates with the highest clear zone indices for protease and cellulase activity were identified as Penicillium citrinum and Fomitopsis sp., with distinct morphological characteristics including velvety colonies and filamentous hyphal structures. The specific activities of the protease and cellulase were 5475.42 ± 2724.25 U/mg protein and 620.77 ± 607.71 U/mg protein, respectively, indicating high catalytic potential. Full article

Background/Objectives: Antimicrobial agents play an important role in the pathogenesis and treatment of Clostridioides (C.) difficile infections. C. difficile isolates have shown different genotypic and phenotypic resistance patterns and could serve as antimicrobial resistance reservoirs. Methods: To gain insight into accordance and potential disagreements between genotypic and phenotypic antimicrobial resistances in C. difficile, we compared the genotypic and phenotypic resistance patterns of 108 bovine C. difficile isolates collected in Germany between 2010 and 2012. These isolates represent a collection of different ribotypes (RT) and originated from different husbandries in Germany. Whole genome sequencing of all isolates was performed with Illumina^® Miseq™, and sequences were screened for antimicrobial resistance determinants. For phenotypic antimicrobial susceptibility testing, the agar dilution procedure according to the CLSI document M11 was used. Minimal inhibitory concentration values were determined for penicillin, meropenem, tetracycline, moxifloxacin, vancomycin, metronidazole, erythromycin and clindamycin. Results: Various phenotypic and genotypic antimicrobial resistances were found in the isolates examined that belonged to different ribotype/sequence type (ST) lineages, even if these originated from the same source and geographical region (bovine isolates from Germany). Agreement between phenotypic and genotypic resistance was seen for most antimicrobial agents tested. A total of 92% (83/90) of the investigated ST11 isolates showed phenotypic resistance or were classified as non-wild type to at least one of the antimicrobials tetracycline, moxifloxacin, erythromycin and clindamycin. Conclusions: The results of this comparison contribute to a better understanding of antimicrobial resistance in C. difficile by relating phenotypic susceptibility patterns to genomic resistance determinants. Full article

The accumulation of polyethylene (PE) in aquatic ecosystems represents a significant environmental challenge due to the polymer’s high molecular weight and chemical stability. This study investigates the biodegradation potential of Hafnia paralvei UUNT_MP29, a bacterial strain isolated from the gut of common carp (Cyprinus carpio), for low-density polyethylene (LDPE). Initial screening on LDPE-emulsified agar confirmed extracellular enzymatic activity through the formation of distinct clear zones. Quantitative analysis showed a cumulative mass loss of 24.10% by Day 16, with the most intensive degradation occurring between Days 4 and 8, which closely correlated with maximum bacterial count (CFU/mL). Kinetic modeling indicated that the degradation followed a first-order rate law (R² = 0.9269), with a rate constant (k) of 0.2991 days⁻¹ and a remarkably short half-life (t_1/2) of 2.32 days. Structural characterization via FTIR spectroscopy demonstrated oxidative transformation, evidenced by a reduction in sp³ C-H stretching and the emergence of C-O/C-O-C functional groups. SEM micrographs further confirmed extensive bio-deterioration, including surface pitting and macroscale erosion. Thermal analysis (TGA/DTG) supported these findings, showing a significant 10.95 °C decrease in the maximum degradation temperature (T_max), indicating a reduction in polymer chain length. These results suggest that H. paralvei UUNT_MP29 is a highly efficient agent for the rapid breakdown of polyethylene and highlight the potential of aquatic gut microbiota as reservoirs for plastic-degrading biotechnologies. Full article

The growing threat of antimicrobial resistance necessitates alternative strategies to conventional antibiotics. Insects represent a promising source of antimicrobial peptides (AMPs) due to their potent innate immune responses. In this study, we investigated the hemolymph peptide extracts from Hermetia illucens larvae as a bioresource of infection-induced AMPs. Larvae were challenged with Gram-negative (Escherichia coli) and Gram-positive (Micrococcus flavus) bacteria, and hemolymph-derived peptides were extracted and fractionated by RP-HPLC. Peptide fractions were screened in vitro by agar diffusion assay against a panel of pathogenic strains. While extracts from uninfected larvae were essentially inactive, fractions 9–13 from infected larvae showed broad-spectrum antibacterial activity, including against multidrug-resistant clinical isolates such as carbapenem-resistant Klebsiella pneumoniae, suggesting an inducible immune response. Fractions were further analyzed by SDS-PAGE and LC-MS/MS to identify candidate AMPs, while shotgun proteomics on unfractionated extracts enabled quantitative comparison among conditions. Proteomics raw data are available via ProteomeXchange Consortium, with the dataset identifier PXD071144. In silico prediction using ProP 1.0, APD3, and CAMP_R4 supported the antimicrobial potential of candidate peptides. Overall, our integrative approach demonstrates that bacterial infection induces a diverse and bioactive immunopeptidome in H. illucens, supporting its potential as a sustainable platform for the discovery of novel antimicrobial agents. Full article

Iron (Fe) and zinc (Zn) are essential micronutrients for plant metabolism; however, their bioavailability in tropical soils is often limited by low solubility and complex mineral interactions. Root-associated bacteria may enhance micronutrient availability through siderophore production and the solubilisation of insoluble mineral forms. This study aimed to functionally characterise three bacterial isolates from rice roots—Bacillus siamensis TUR07-02b, Priestia aryabhattai SMNCH17-07, and Priestia megaterium SMBH14-02—under controlled in vitro conditions. Siderophore activity was evaluated qualitatively using Chrome Azurol S (CAS) agar, where percentages represent halo-based indices relative to colony diameter, and quantitatively using the CAS–shuttle assay, expressed as percent siderophore units relative to an uninoculated reference. Zinc solubilisation was assessed in solid media as halo-based indices and in liquid media as Zn-equivalent signals (mg L⁻¹) obtained by spectrophotometry. All strains produced siderophores, with P. aryabhattai showing the highest qualitative index (167%), while P. aryabhattai and B. siamensis showed statistically similar activity in liquid medium (~23%). Zinc solubilisation was substrate-dependent: B. siamensis showed the broadest solubilisation spectrum in solid media, whereas P. aryabhattai achieved the highest Zn-equivalent signals for ZnCO₃ and Zn₃(PO₄)₂ after 20 days. These results demonstrate strain-specific functional differences and represent a preliminary screening method based on relative in vitro estimations. Full article

In this study, multiple samples were collected from different urea-fertilized agricultural lands, and their fungal strains were isolated using the tenfold serial dilution method on potato dextrose agar plates. In total, 21 strains were identified as urease-positive through primary screening on Christensen medium. Secondary screening of selected fungal isolates conducted through submerged fermentation could then identify the fungal strain 10⁻⁵ S₁₁ brown as the most effective urease producer that exhibited maximum urease activity (682 U/mL/min). It was identified by scotch tape microscopy for morphological characterization and subsequently confirmed through 18S rRNA sequencing as Aspergillus terreus. Further, optimization of fermentation conditions showed that M9 medium containing 1.5% urea as a nitrogen source at pH 5.5, in addition to 3% sucrose as a carbon source, 4% inoculum size, and 7 days of incubation at 30 °C, produced the best fermentation and enhanced the urease activity from 682 U/mL/min to 1050 U/mL/min. Subsequently, the optimized urease enzyme was mixed with clean sand to induce carbonate precipitation to enhance its unconfined compressive strength from 22.5 kPa for untreated samples to 154.2 kPa for treated samples after 28 days, with subtle permeability reduction from 4.26 × 10⁻³ cm/s to 1.7 × 10⁻³ cm/s. Full article

Background/Objectives: Multidrug-resistant (MDR) Escherichia coli resistant to third-generation cephalosporins are a growing One Health concern, but data on extraintestinal pathogenic E. coli (ExPEC) from wildlife in North Africa remain scarce. We aimed to characterize ESBL/AmpC-producing ExPEC from captive wild mammals in Tunisia and to situate these isolates in a global genomic context. Methods: In 2018, 30 fecal samples from 14 captive wild mammals in a private farm were screened on cefotaxime agar. Four cefotaxime-resistant E. coli isolates were recovered from a llama, lion, hyena, and tiger. Antimicrobial susceptibility testing and Illumina whole-genome sequencing were combined with in silico typing, resistome and virulome profiling, plasmid and mobile element analysis, human pathogenicity prediction and core-genome MLST-based minimum-spanning trees. Results: All isolates were MDR but remained susceptible to carbapenems, colistin and tigecycline. Two ST162/B1 isolates from the llama and tiger carried bla_CMY-2, whereas two ST69/D isolates from the lion and hyena harbored bla_CTX-M-15 and qnrS1. Genomes encoded 61–68 antimicrobial resistance genes and 114–131 virulence-associated genes, together with IncF-, IncI1- and IncY-type plasmids and IS26-rich insertion sequence profiles. Mating-out assays yielded cefotaxime-resistant transconjugants, supporting plasmid transferability of bla_CMY-2 or bla_CTX-M-15. PathogenFinder predicted a ≥0.93 probability of human pathogenicity for all isolates. cgMLST-based trees showed that Tunisian ST69 and ST162 clustered within internationally disseminated lineages containing human, animal and food isolates, rather than forming wildlife-restricted branches. Conclusions: Captive wild mammals in Tunisia can harbor high-risk ExPEC lineages combining ESBL/AmpC production, multidrug resistance and extensive virulence and mobility gene repertoires. These findings highlight captive wildlife as potential reservoirs and sentinels of clinically relevant E. coli and underscore the need for integrated WGS-based One Health surveillance at the human–animal–environment interface in North Africa. Full article

Microorganisms represent the Earth’s most abundant biomass and a vast reservoir of genetic diversity. However, traditional agar plate methods fail to recover the vast majority of these species, leaving a “microbial dark matter” that holds immense potential for the discovery of novel antibiotics and bioactive compounds. While conventional techniques such as selective media and enrichment culture remain foundational, they are inherently limited by community biases and the inability to support low-abundance, oligotrophic species. To address these bottlenecks, a diverse array of innovative isolation strategies has emerged. This review systematically categorizes and evaluates these methodologies, ranging from in situ cultivation to high-resolution single-cell manipulation. We first examine membrane diffusion-based cultivation (e.g., iChip), which mimics natural microenvironments to resuscitate recalcitrant microbes. Subsequently, we explore high-throughput single-cell technologies, including microfluidics for physicochemical separation, optical tweezers for precise manipulation, and fluorescence-activated cell sorting (FACS). Special attention is given to Raman-activated cell sorting (RACS) as a label-free functional screening tool and reverse genomics for targeted capture. By synthesizing the strengths and limitations of these approaches, we propose integrated workflows designed to accelerate the mining of untapped microbial resources. Full article