Rapid and accurate detection of microorganisms in sterile body fluids, particularly cerebrospinal fluid (CSF), is crucial for effective diagnosis and treatment. Conventional methods, such as centrifugation, may result in low microbial recovery and false negatives, limiting diagnostic accuracy. An alternative, efficient, and accessible microbial concentration method is needed. This study evaluates a nanoparticle-based microbial concentration method to enhance pathogen recovery from CSF. The method was optimized for interaction time (1 min) and nanoparticle dosage (0.01 g/mL) using standard microbial strains, including Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa, and Candida albicans. Its clinical performance was assessed using 800 CSF samples, and microbial identification was confirmed via MALDI-TOF MS. The method significantly improved microbial recovery across 10-2–10-9 CFU/mL concentrations, achieving a detection limit as low as 2 CFU/mL. Clinical validation demonstrated 100% sensitivity and specificity, detecting 15 additional true-positive cases missed by centrifugation. While centrifugation fails to detect bacteria below 10-7 CFU/mL, our method reliably detects even at 10-9 CFU/mL, demonstrating superior microbial enrichment, especially in low-biomass samples. This method enhances diagnostic accuracy by reducing false negatives and expediting pathogen detection. Its resource-conscious, low-cost and equipment-free nature makes it particularly beneficial for resource-limited laboratories, offering a scalable alternative for microbial concentration in CSF diagnostics.Graphical

Bdellovibrio bacteriovorus is an agent that stands out with its predatory properties and has recently been used against pathogens that are frequently resistant to antibiotics. The study was conducted experimentally to determine the effect of dressing application containing Bdellovibrio bacteriovorus on superficial incisional surgical site infection caused by Staphylococcus aureus in mice. In the study, mice were divided into 6 different groups, BB: B. bacteriovorus; NC: Negative Control; PC: Positive Control Methicillin Resistant S. aureus; MRSA + BB: Methicillin Resistant S. aureus + B. bacteriovorus dressing; MRSA + V: Methicillin Resistant S. aureus + Vancomycin; MRSA + BB + V: Methicillin Resistant S. aureus + B. bacteriovorus dressing + Vancomycin group. The treatment procedures were applied over a period of 3 days. Infection symptoms were monitored and recorded at the 24th, 48th, and 72nd hours. In the Staphylococcus aureus + Vancomycin group, all mice developed edema, redness, and fever at 24 h. At 48 h, all mice exhibited edema and redness, with 50% showing fever. At 72 h, 70% of the mice showed edema and redness, and 10% showed fever. In the Staphylococcus aureus + Bdellovibrio bacteriovorus + Vancomycin combined treatment group, all mice exhibited edema, redness, and fever at 24 h. At 48 h, only 20% of the mice showed redness. At 72 h, no edema, redness, fever, purulent discharge, or suture dehiscence was observed. Sepsis developed in 2 of 10 mice in the Staphylococcus aureus + Bdellovibrio bacteriovorus + Vancomycin group. The most effective treatment was in the Staphylococcus aureus + Bdellovibrio bacteriovorus + Vancomycin group. It was determined that sepsis findings were the least in the Staphylococcus aureus + Bdellovibrio bacteriovorus + Vancomycin group. B. bacteriovorus holds the potential to be an effective control agent in preventing or slowing resistance development.

Methicillin-resistant Staphylococcus aureus (MRSA) biofilms pose a severe risk to public health, showing resistance to standard antibiotics, which drives the need for novel antibacterial strategies. Bacteriophages have emerged as potential agents against biofilms, especially through their phage-encoded enzymes that disrupt the biofilm matrix, enhancing bacterial susceptibility. In this study, two bacteriophages, UPMK_1 and UPMK_2, were propagated on MRSA strains t127/4 and t223/20, respectively. Biofilms formed by these strains were treated with phages at specified concentrations, followed by protein extraction and analysis. Comparative proteomic profiling was performed using one-dimensional and two-dimensional SDS-PAGE, with protein identification facilitated by MALDI-TOF/TOF MS spectrometry, to observe biofilm degradation effects. Proteomic analysis revealed that phage treatment induced significant changes in biofilm protein expression, particularly with upregulated ribosome-recycling factors and elongation factors linked to enhanced protein synthesis, reflecting a reactivation of amino acid metabolism in the treated biofilms. This was marked by upregulated intracellular proteases like CIpL, which play a role in protein refolding and degradation, critical for phage progeny production and biofilm disruption. Phage treatment demonstrated notable effects on the metabolic and protein synthesis pathways within MRSA biofilms, suggesting that phages can redirect bacterial cellular processes to favour biofilm breakdown. This indicates the potential of bacteriophages as a viable adjunct to traditional antimicrobial approaches, particularly in combating antibiotic-resistant infections like MRSA. The study underscores the efficacy of bacteriophages as anti-biofilm agents, offering a promising strategy to weaken biofilms and combat antibiotic resistance through targeted disruption of bacterial metabolic pathways and biofilm integrity.

Bacteria form consortia as integral components of diverse ecosystems, where they interact with various organisms. Within these communities, bacterial–bacterial communication plays a pivotal role by driving numerous specific interactions. A key aspect of this chemical communication is the production of secondary metabolites. Recent research demonstrates that interspecies interactions between microorganisms can serve as physiological triggers, activating silent biosynthetic gene clusters and leading to the synthesis of novel secondary metabolites by the interacting species. This review focuses on mixed cultivation strategies involving actinobacteria, with an emphasis on utilizing mycolic acid-containing bacteria such as Tsukamurella pulmonis as inducer organisms. It comprehensively examines recent advances striving to understand these bacterial interactions, specifically involving the ability of actinomycetes to recognize and respond to mycolic acid-containing bacteria to activate secondary metabolism. Furthermore, the genetic basis of secondary metabolism activation was explored and newly discovered secondary metabolites induced by actinobacteria–mycolic acid-containing bacteria co-culture were highlighted. Finally, the integration of combined-culture strategies with genetic engineering methods and the ecological relevance of actinobacteria–mycolic acid-containing bacteria interactions were discussed. These bacterial interactions provide an excellent model system for understanding the molecular mechanisms regulating secondary metabolism and could open new avenues for drug discovery.

A culture-based strategy was used to isolate and screen representative actinomycetes from six sampling sites in the Karakum Desert, Turkmenistan. A total of 459 actinomycete isolates were obtained using 16 selective media, and 270 representative strains were subjected to 16 S rRNA gene sequencing. Comparative 16 S rRNA gene sequence analyses on colour-group representatives led to their assignment to 17 genera with validly published names which included many isolates assigned to novel or putatively novel species including ones belonging to rare genera, such as Actinocorallia, Actinomadura, Jiangella and Nonomuraea. Mining of whole-genome sequences of 32 isolates which formed distinct lineages in phylogenomic trees revealed biosynthetic gene clusters predicted to encode for many novel specialized metabolites, notably antibiotics. The genomes of most of these isolates included genes associated with the promotion of plant growth while bioinformatic analyses of stress-related genes provided on insight into how filamentous actinomycetes have adapted to harsh environmental conditions in the Karakum Desert. This extensive bioprospecting campaign shows that the Karakum Desert is a unique source of novel, rare and gifted filamentous actinomycetes with the ability to synthesise new specialized metabolites needed to address key existential issues facing humankind, especially, the urgent need to find a new generation of therapeutic antibiotics to control multidrug-resistant microbial pathogens and compounds that protect and promote plant growth.

Characterization of lipid-degrading microorganisms and understanding their metabolite production mechanisms are essential for enhancing biodegradation efficiency of lipid-rich wastewater. In this study, we isolated Burkholderia arboris strain JYK2, which demonstrated lipid degradation at 46.29 mg/(l • h) with lipase activity reaching 59.92 U/ml when lipid was provided as the sole carbon source. Experimental results revealed that strain JYK2 secretes lipase in media containing lipids and fatty acids but not in glycerol-containing media, a phenomenon likely attributable to quorum sensing mechanisms. Furthermore, extracellular polymeric substances (EPS) were produced in the presence of lipids and fatty acids. Compositional analysis showed that EPS primarily consisted of proteins (approximately 50%) and polysaccharides (approximately 25%). This protein-rich characteristic conferred high hydrophobicity to the EPS, contributing to its lipase adsorption capacity as verified in this study. Additionally, biosurfactant activity was detected in the EPS produced by JYK2. These functions collectively enhance substrate utilization by microorganisms and promote lipid biodegradation. However, minimal EPS production was observed at low fatty acid concentrations, suggesting that the EPS production mechanism cannot be solely attributed to quorum sensing. This study provides insights into the interactions between lipid-degrading bacteria, lipase production, and EPS functionality, which are crucial for optimizing biological treatment of lipid-rich wastewater.Graphical

Limited research has investigated the ability of psychrophilic and psychro-tolerant microorganisms to produce cold-active keratinases, despite their potential as an efficient alternative for substrate conversion at reduced energy expenditure. A screening of 32 Penicillium and Talaromyces isolates for keratinolytic activity at temperatures of 5, 10, and 15ºC identified a promising P. oxalicum strain as the most potent at 10ºC, yielding 242.39 U/mL. Following six days of incubation at pH 8.0 and 15 °C with 0.2% yeast extract as the nitrogen source, the P. oxalicum strain exhibited keratinase activity of 359.42 U/mL. The keratinase underwent purification with a 4.13-fold increase, utilizing an MP 800 anion exchanger and Sephacryl S 200 , resulting in a specific activity of 684.46 U/mg and a yield of 5.34%. The SDS-PAGE analysis identified a keratinase with a molecular weight of 37.51 kD, exhibiting peak activity at pH 9.0 and 20ºC, with a specific activity of 721.8 U/mg. Mg2+, Zn2+, and Mn2+ enhanced keratinase activity by 156.0%, 140.60%, and 156.0%, respectively. The keratinase activity was significantly enhanced (p < 0.05) by the addition of 5 mM SDS (139.15%), 5 and 10% mercaptoethanol (1125.70 and 1327.0%, respectively), and 5 and 10% DMSO (128.30 and 227.40%, respectively). The dehairing potential of P. oxalicum AUMC 15084, utilizing crude keratinase on goat skin, demonstrated complete dehairing after 20 h at 20ºC with the crude preparation. This study provides a promising Penicillium oxalicum strain that could be used for production of cold-active keratinase. The effectiveness of the produced keratinase in the dehairing process was demonstrated as an environmentally friendly alternative to the traditional chemical procedure.

Pseudomonas aeruginosa displays major biofilm formation, food spoilage, and significant losses for the food industry. It extremely exhibits inherent resistance to various antibiotics, including quinolones, which can spread via plasmids. The present study investigates the prevalence of biofilm-forming P. aeruginosa in meat products and assesses its antibiotic resistance patterns. Evaluating the effectiveness of electrolyzed-oxidized water (EOW) and per-acetic acid (PAA) alone or in a combination against P. aeruginosa biofilm formed on stainless steel (SS) surfaces. Besides, the effect on quinolone-resistance (qnr) gene expression level using real-time PCR. P. aeruginosa was isolated from 24.6% of the total tested samples (37/150), with a significant variation observed regarding the contamination levels of the tested products. 36.3% of the isolates demonstrated robust biofilm production, with 82.3% displaying multi-drug resistance (MDR). Isolates revealed complete susceptibility to amikacin (100%) and gentamycin (82%). Quinolone resistance was observed in 23% and 17% of the isolates for ciprofloxacin and levofloxacin, respectively. 80% (4/5) among the confirmed isolates were enclosed the plasmid qnr genes. The genes pslA and gyrA were detected in 40% and 60%, respectively. EOW, particularly when combined with PAA, reveals strong antibacterial activity against P. aeruginosa and greatly decreases its count. Moreover, it can eliminate its biofilm within 20 min of exposure and significantly decrease the expression levels of quinolone-resistant genes. In conclusion, PAA and EOW are effective agents for biofilm control on SS surfaces, particularly when used in combined form, and could be more effective in combating resistant bacterial infections and controlling the spread of PMQR.

The capability of mycobacteria to survive in adverse environments is crucial for successful infection, yet the underlying mechanisms remain unclear. A novel sRNA, ncBCG427, was previously identified in intracellular versus extracellular mycobacteria, with predicted targets clustering in lipid metabolism pathways in Mycobacterium smegmatis (M. sm). This study aimed to investigate how ncBCG427 regulates the survival of M. sm through lipid metabolism. Using lipidomics, metabolites from the ncBCG427-expressing strain (MS_ncBCG427) and the control strain (MS_Vector) were screened, revealing enrichment in lipid-associated pathways. The gene MSMEG_4757 (Fas) was identified as critical to this pathway and confirmed as a target of ncBCG427. Western blot analysis demonstrated that ncBCG427 increased Fas expression in THP-1 cells post-infection. Additionally, Oil Red O staining indicated that both ncBCG427 and MSMEG_4757 enhanced lipid droplet formation in A549 cells. Both MS_ncBCG427 and MS_4757 exhibited increased biofilm formation and enhanced survival under various adverse conditions, including carbon starvation, acid stress, membrane stress, and exposure to drugs such as rifampicin and streptomycin. In contrast, low-expression strains (MS_sh4757 and MS_ncBCG427_sh4757) showed reduced survival. In conclusion, ncBCG427 targets MSMEG_4757 to regulate lipid metabolism, enhancing biofilm formation and survival in adverse environments, revealing a novel mechanism of mycobacterial survival and potential antimicrobial targets.