Search Results (181)

Background: Iminosugars are natural or synthetic sugar analogues with a very broad spectrum of activities, including those against the most prominent bacterial pathogens, like P. aeruginosa or S. aureus. In a series of studies, we have demonstrated that one of the synthetic iminosugars, PDIA (beta-1-C-propyl-1,4-dideoxy-1,4-imino-L-arabinitol), possesses the ability to suppress biofilm production by different pathogenic bacteria without inhibiting their growth. Thereby, PDIA is able to influence experimental skin infection caused by S. aureus. Methods: To elucidate molecular mechanisms by which PDIA impedes biofilm formation by S. aureus, a transcriptomic study was performed in which a biofilm-producing S. aureus strain was grown in the presence of PDIA for 24 and 48 h in comparison to a control without the iminosugar. The RNA was then isolated, converted into cDNA, sequenced, and data analysis was performed. Results: It appeared that PDIA caused the down-regulation of many bacteriophage genes responsible for the processes of bacterial cell lysis, and some genes responsible for cell wall degradation were also down-regulated. Among the 25 most upregulated genes were those representing the phosphotransferase system (PTS), which is required for carbohydrate uptake and control of carbon metabolism. The ranking of the most significant down-regulated genes after 24 h exposure to PDIA shows that they predominantly coded for both the synthesis and lysis of the peptidoglycan. Conclusions: We have shown here that the influence of PDIA on the expression of S. aureus genes is broad and affects many genes encoding metabolism and ribosomes. Full article

Background/Objectives: Antibiotic resistance or antimicrobial resistance (AMR) in livestock is a growing global concern that threatens both human and animal health. The overuse and misuse of antibiotics in livestock production have led to an increased propensity for the development of AMR bacterial strains in animals, which can be spread to humans through the consumption of contaminated animal products, direct contact, or environmental exposure. This review aims to summarize the development and transmission of AMR in livestock, explore its underlying mechanisms and impact on human and animal health, and discuss current practices and potential strategies for mitigation and prevention. Methods: For this narrative review, we searched articles on PubMed and Google Scholar using the terms antibiotic resistance, livestock, and environment, alone or in combination. Results: The history of antibiotic use in livestock and its link to increased AMR, along with the involved mechanisms, including the enzymatic breakdown of antibiotics, alterations in bacterial targets, horizontal gene transfer, and efflux pumps, are important. Antibiotics in livestock are used for growth promotion, disease prevention and control, and metaphylactic use. The role of livestock and the environment as reservoirs for resistant pathogens, their impact on human health, chronic infections, allergic reactions, toxicity, and the development of untreatable diseases is important to understand AMR. Conclusions: Given the widespread use of antibiotics and the potential consequences of AMR, collaborative global efforts, increased public awareness, coordinated regulations, and advancements in biological technology are required to mitigate the threat AMR poses to human and animal health. Regulatory solutions and the development of new therapeutic alternatives like antimicrobial peptides and bacteriophage therapy, and preventive measures such as DNA and mRNA vaccines, are future perspectives. Full article

Enterococcus faecalis (E. faecalis) is a major pathogen responsible for refractory apical periodontitis (RAP). It can penetrate deep into dentinal tubules, form persistent biofilms, and exhibit antibiotic resistance, thereby limiting the efficacy of conventional antimicrobial treatments. Bacteriophages (phages), due to their strong lytic activity and host specificity, have emerged as promising alternatives. In this study, a novel strictly lytic phage, ZXL-01, was isolated from lake water in Jilin, China. ZXL-01 demonstrated remarkable stability under extreme conditions, including thermal tolerance at 60 °C for 1 h and a wide pH range (4–11). Whole-genome sequencing (GenBank accession number: ON113334) revealed a genome of 40,804 bp with no virulence or tRNA genes, confirming its identity as an E. faecalis phage. Importantly, ZXL-01 exhibited potent antibiofilm activity, reducing biofilm biomass by approximately 69.4% in the inhibition group and 68.4% in the lysis group (both p < 0.001). In an in vitro root canal infection model induced by E. faecalis, scanning electron microscope (SEM) observations confirmed that ZXL-01 effectively inhibited biofilm formation and disrupted mature biofilms. These findings highlight the potential of ZXL-01 as a novel antimicrobial agent for the treatment of E. faecalis-associated apical periodontitis. Full article

Small regulatory RNAs (sRNAs) play a critical role in bacterial gene expression, modulating various cellular processes, including stress responses, metabolism, virulence, and many others. While well-characterized in bacterial systems, an emerging class of phage-derived sRNAs has been identified, suggesting an underexplored regulatory network at phage–host interactions. These sRNAs, encoded within phage genomes, influence both bacterial and viral life cycles by modulating transcriptional and post-transcriptional gene expression processes. The interplay between phage-derived sRNAs and the host genome reveals a complex network of gene regulation, with an impact on bacterial fitness, pathogenesis, and horizontal gene transfer. This review explores the diverse functions of phage-encoded sRNAs, highlighting recent discoveries and their impact on bacterial physiology and phage-host interactions. Full article

We hypothesize that bacteria isolated from free-ranging animals could potentially be useful for practical applications. To meet this objective a Gram-positive bacterium was isolated from the gastrointestinal (GI) tract of a Gray Wolf (Canis lupus) using Brucella broth with hemin and vitamin K (BBHK). By small ribosomal RNA (16S) gene sequencing the bacterium was initially identified as a novel Carnobacterium maltaromaticum strain. The bacterium could be propagated both anaerobically and aerobically and was both catalase/oxidase negative and negative by the starch hydrolysis as well as negative using lipase assays. The reference whole genome sequence (WGS) was obtained using both Illumina and Nanopore sequencing. The genome assembly was 3,512,202 bp in length, encoding core bacterial genes with a GC% content of 34.48. No lysogenic bacteriophage genes were detected, although the genome harbors genes for the expression of bacteriocin and other secondary metabolites with potential antimicrobial properties. Multilocus sequence typing (MLST), WGS phylogenetics, average nucleotide identity (ANI), and single nucleotide polymorphism (SNP) analyses of the isolate’s genome indicate this bacterium is a newly identified Carnobacterium maltaromaticum sequence type (ST). Members of the Carnobacteria have anti-listeria activities, highlighting their potential functional properties. Consequently, the isolate could be a potential probiotic for canids and this is the first report on an axenic C. maltaromaticum culture from the genus Canis. Full article

Several strains of Bifidobacterium animalis subsp. lactis are blockbusters of commercial dietary supplement cocktails, widely recognized for their probiotic properties and found in various ecological niches. The present study aimed to perform an in-depth comparative genomic analysis on 71 B. animalis subsp. lactis strains isolated from diverse sources, including human and animal feces, breast milk, fermented foods, and commercial dietary supplements, to better elucidate the strain level diversity and biotechnological potential of this species. The average genome size was found to be 1.93 ± 0.05 Mb, with a GC content of 60.45% ± 0.2, an average of 1562 ± 41.3 coding sequences (CDS), and 53.4 ± 1.6 tRNA genes. A comparative genomic analysis revealed significant genetic diversity among the strains, with a core genome analysis showing that 34.7% of the total genes were conserved, while the pan-genome remained open, indicating ongoing gene acquisition. Functional annotation through EggNOG-Mapper and CAZYme clustering highlighted diverse metabolic capabilities, particularly in carbohydrate metabolism. Nearly all (70 of 71) Bifidobacterium animalis subsp. lactis strains were found to harbor CRISPR-Cas adaptive immune systems (predominantly of the Type I-E subtype), underscoring the ubiquity of this phage defense mechanism in the species. A comparative analysis of spacer sequences revealed distinct strain-specific CRISPR profiles, with certain strains sharing identical spacers that correlate with common phylogenetic clades or similar isolation sources—an indication of exposure to the same phage populations and shared selective pressures. These findings highlight a dynamic co-evolution between B. lactis and its bacteriophages across diverse ecological niches and point to the potential of leveraging its native CRISPR-Cas systems for future biotechnological applications. Our findings enhance our understanding of the genetic and functional diversity of B. animalis subsp. lactis, providing valuable insights for its use in probiotics and functional foods. Full article

Bacteriophage therapy is a promising approach for targeting antibiotic-resistant bacteria and modulating gut microbiota in metabolic diseases such as obesity. This study evaluated the impact of a two-phage cocktail on an ex vivo colonic simulation model of gut microbiota derived from obese individuals, both in its normalized state and after enrichment with Enterobacter cloacae, an obesity-related bacteria. Microbiological analyses confirmed that the phage cocktail remained active throughout the colonic regions over three digestion cycles and effectively reduced enterobacterial populations in the enriched microbiota. Metabarcoding of the 16S rRNA gene revealed that phage therapy did not significantly alter the abundance of dominant genera, but selectively reduced E. cloacae across all colonic regions. Alpha diversity was significantly affected only in the enriched microbiota, while beta diversity analysis indicated significant compositional shifts during therapy, with reduced dispersion in the final treatment stage. Short-chain fatty acid profiling demonstrated region- and group-specific metabolic responses, with increased lactic and butyric acid concentrations in the ascending colon of the enriched microbiota following phage treatment. This study provides the first ex vivo evidence that a two-phage cocktail can selectively eliminate E. cloacae while preserving overall microbiota structure and functionality. These findings establish a foundation for future in vivo studies exploring the role of phage therapy in reshaping gut microbial communities and metabolic profiles, highlighting its potential as a precision tool for managing gut dysbiosis in metabolic disorders. Full article

Enzymes play an essential role in many different industries; however, their operating conditions are limited due to the loss of enzyme activity in the presence of proteases and at temperatures significantly above physiological conditions. One way to improve the stability of these enzymes against high temperatures and proteases is to encapsulate them in protective shells or virus-like particles. This work presents a streamlined, three-step, cell-free protein synthesis (CFPS) procedure that enables rapid in vitro enzyme production, targeted encapsulation in protective virus-like particles (VLPs), and facile purification using a 6× His-tag fused to the VLP coat protein. This process is performed in under 12 h and overcomes several limitations of enzyme encapsulation, such as the control of packing density, speed, and complexity of the process. Here, we encapsulate the enzyme Candida antarctica lipase B in the VLP from the bacteriophage Qβ, while in the presence of a linking RNA aptamer. The encapsulated enzymes largely retained their activity in comparison to the free enzymes. Additionally, when subjected to 90 °C temperatures or 5 h incubation with proteases, the encapsulated enzymes maintained their activity, whereas the free enzymes lost their activity. In this work, we also demonstrate control over packing density by achieving packing densities of 4.7 and 6.5 enzymes per VLP based off the concentration of enzyme added to the encapsulation step. Full article

Six novel Microbacterium phages belonging to the Tectiviridae family were isolated using Microbacterium testaceum as a host. Phages MuffinTheCat, Badulia, DesireeRose, Bee17, SCoupsA, and LuzDeMundo were purified from environmental samples by students participating in the Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) program at Alliance University, New York. The phages have linear dsDNA genomes 15,438–15,636 bp with 112–120 bp inverted terminal repeats. Transmission electron microscopy (TEM) imaging analysis revealed that the six novel phages have six-sided icosahedral double-layered capsids with an internal lipid membrane that occasionally forms protruding nanotubules. Annotation analysis determined that the novel Microbacterium phages all have 32–34 protein-coding genes and no tRNAs. Like other Tectiviridae, the phage genomes are arranged into two segments and include three highly conserved family genes that encode a DNA polymerase, double jelly-roll major capsid protein, and packaging ATPase. Although the novel bacteriophages have 91.6 to 97.5% nucleotide sequence similarity to each other, they are at most 58% similar to previously characterized Tectiviridae genera. Consequently, these novel Microbacterium phages expand the diversity of the Tectiviridae family, and we propose they form the sixth genus, Zetatectivirus. Full article

RNA nanoparticles, derived from the packaging RNA three-way junction motif (pRNA-3WJ) of the bacteriophage phi29 DNA packaging motor, have been demonstrated to be thermodynamically and chemically stable, with promise as a nanodelivery system. Background/Objectives: A previous study showed that RNA nanoparticles with antiangiogenic aptamers (anti-vascular endothelial growth factor (VEGF) and anti-angiopoietin-2 (Ang2) aptamers) inhibited cell proliferation via WST-1 assay. To further investigate the antiangiogenic potential of these RNA nanoparticles, a modified three-dimensional (3D) spheroid sprouting assay model of human umbilical vein endothelial cells was utilized in the present study. Methods: Three groups of RNA nanoparticles were evaluated, namely, pRNA-3WJ series, RNA square series (polygon-type RNA nanoparticles), and 8WJ series (multiple-way junction RNA nanoparticles), which were conjugated with a single anti-VEGF, the combination of one anti-VEGF and one anti-Ang2, or multiple anti-VEGF aptamers. The core scaffold RNA nanoparticles (without aptamers) were used as the references, and bevacizumab was used as the positive control. Results: The results demonstrated the inhibition effects of the RNA nanoparticles on endothelial cell tube formation at 67 nM in a 3D spheroid sprouting model. The results in the 3D spheroid sprouting assay are consistent with those of the WST-1 proliferation assays. Conclusions: Among the RNA nanoparticles evaluated, 3WJ-3VEGF and SQR-VEGF-Ang2 had inhibition effects equivalent to bevacizumab and were promising for anti-angiogenesis treatment. Full article

As natural predators of bacteria, tailed bacteriophages can be used in biocontrol applications, including antimicrobial therapy. Also, phage lysis is a detrimental factor in technological processes based on bacterial growth and metabolism. The spectrum of bacteria bacteriophages interact with is known as the host range. Phage science produced a vast amount of host range data. However, there has been no attempt to analyse these data from the viewpoint of modern phage and bacterial taxonomy. Here, we performed a meta-analysis of spotting and plaquing host range data obtained on strains of production host species. The main metric of our study was the host range value calculated as a ratio of lysed strains to the number of tested bacterial strains. We found no boundary between narrow and broad host ranges in tailed phages taken as a whole. Family-level groups of strictly lytic bacteriophages had significantly different median plaquing host range values in the range from 0.18 (Drexlerviridae) to 0.70 (Herelleviridae). In Escherichia coli phages, broad host ranges were associated with decreased efficiency of plating. Bacteriophage morphology, genome size, and the number of tRNA-coding genes in phage genomes did not correlate with host range values. From the perspective of bacterial species, median plaquing host ranges varied from 0.04 in bacteriophages infecting Acinetobacter baumannii to 0.73 in Staphylococcus aureus phages. Taken together, our results imply that taxonomy of bacteriophages and their bacterial hosts can be predictive of intraspecies host ranges. Full article

Salmonella is one of the main foodborne pathogens. Irrational antibiotic management has led to an increase in the incidence of multidrug-resistant strains. Bacteriophages may be an alternative method of food biopreservation and contribute to reducing the number of food poisonings requiring pharmacotherapy. This study aimed to isolate a bacteriophage (phage) targeting indigenous multidrug-resistant (MDR) Salmonella strains, followed by their biological, morphological, and genomic characterization. In this study we isolated Salmonella phage KKP_3822, targeting MDR Salmonella Manchester strain KKP 1213. Salmonella phage KKP_3822 retained high activity in the temperature range from −20 °C to 40 °C and active acidity from pH 3 to 11. Temperatures of 70 °C and 80 °C and extreme pH values (2 and 12) significantly reduced the phage titer. Its activity decreased proportionally to the time of UV exposure. Genome analysis (linear dsDNA with a length of 114,843 bp) revealed the presence of 27 tRNA genes. Proteins encoded by the vB_Sen-IAFB3822 phage were divided into functional modules related to (i) phage structure/assembly, (ii) DNA replication/modification/regulation, (iii) phage lysis, and (iv) DNA packaging into the capsid. No genes associated with antibiotic resistance or integration into the host genome, markers of temperate bacteriophages, were annotated in the Salmonella phage KKP_3822 genome. Based on morphological features and whole-genome sequence analysis, the newly isolated Salmonella phage KKP_3822 shows the greatest similarity to representatives of tailed phages from the Caudoviricetes class, Demerecviridae family, and Epseptimavirus genus. Genome analysis confirmed the virulent nature of the Salmonella phage KKP_3822, making it a potential candidate for food biocontrol. Full article

Background/Objectives: Antimicrobial resistance (AMR) is a major public health threat, which is exacerbated by the lack of new antibiotics and the emergence of multidrug-resistant (MDR) superbugs. Comprehensive efforts and alternative strategies to combat AMR are urgently needed to prevent social, medical, and economic consequences. Pseudomonas aeruginosa is a pathogen responsible for a wide range of infections, from soft tissue infections to life-threatening conditions such as bacteremia and pneumonia. Bacteriophages have been considered as a potential therapeutic option to treat bacterial infections. Our aim was to isolate phages able to infect MDR P. aeruginosa strains. Methods: We isolated two lytic phages, using the conventional double layer agar technique (DLA), from samples obtained from the influent of a wastewater treatment plant in Concepción, Chile. The phages, designated as PaCCP1 and PaCCP2, were observed by electron microscopy and their host range was determined against multiple P. aeruginosa strains using DLA. Moreover, their genomes were sequenced and analyzed. Results: Phage PaCCP1 is a member of the Septimatrevirus genus and phage PaCCP2 is a member of the Pbunavirus genus. Both phages are tailed and contain dsDNA. The genome of PaCCP1 is 43,176 bp in length with a GC content of 54.4%, encoding 59 ORFs, one of them being a tRNA gene. The genome of PaCCP2 is 66,333 bp in length with a GC content of 55.6%, encoding 102 non-tRNA ORFs. PaCCP1 is capable of infecting five strains of P. aeruginosa, whereas phage PaCCP2 is capable of infecting three strains of P. aeruginosa. Both phages do not contain bacterial virulence or AMR genes and contain three and six putative Anti-CRISPR proteins. Conclusions: Phages PaCCP1 and PaCCP2 show promise as effective treatments for MDR P. aeruginosa strains, offering a potential strategy for controlling this clinically important pathogen through phage therapy. Full article

Different viruses are abundant in aquatic ecosystems. There has been limited research on the viral communities in the upper reaches of the Yangtze River. Yellow catfish (Pelteobagrus fulvidraco), an important economic fish that is widely distributed in the upper reaches of the Yangtze River, was selected as the research object. Using RNA sequencing, we identified 11 viruses belonging to the Adintoviridae, Tombusviridae, Caudovirales, Microviridae, Picornavirales, and other bacteriophage families. The predominant viral families/order in Luzhou (LZ), Fuling (FL), and Wanzhou (WZ) were Caudovirales, Adinoviridae, and Microviridae, respectively. The virome from WZ had a unique community composition, with a high abundance of Picornavirales compared with LZ and FL. In LZ, the predominant double-stranded RNA virus family was Siphoviridae. Phylogenetic analyses showed that viruses presented high genetic diversity. Phylogenetically, Wenling pleuronectiformes picornavirus was close to the family Caliciviridae, which includes yellow catfish calicivirus (YcCV), responsible for the massive mortality of yellow catfish in 2020. This study provides insights into the viral community composition in yellow catfish in the upper reaches of the Yangtze River, revealing a diverse and unique river water virome and providing clues for future research on the origin of viral pathogens. Full article

To better understand host–phage interactions and the genetic bases of phage resistance in a model system relevant to potential phage therapy, we isolated several spontaneous mutants of the USA300 S. aureus clinical isolate NRS384 that were resistant to phage K. Six of these had a single missense mutation in the host rpoC gene, which encodes the RNA polymerase β’ subunit. To examine the hypothesis that mutations in the host RNA polymerase affect the transcription of phage genes, we performed RNA-seq analysis on total RNA samples collected from NRS384 wild-type (WT) and rpoC_G17D mutant cultures infected with phage K, at different timepoints after infection. Infection of the WT host led to a steady increase of phage transcription relative to the host. Our analysis allowed us to define 53 transcriptional units and to categorize genes based on their temporal expression patterns. Predicted promoter sequences defined by conserved −35, −10, and, in some cases, extended −10 elements, were found upstream of early and middle genes. However, in many cases, sequences upstream of late genes did not contain clear, complete, canonical promoter sequences, suggesting that factors in addition to host RNA polymerase are required for their expression. Infection of the rpoC_G17D mutant host led to a transcriptional pattern that was similar to that of the WT at early timepoints. However, beginning at 20 min after infection, transcription of late genes (such as phage structural genes and host lysis genes) was severely reduced. Our data indicate that the rpoC_G17D mutation prevents the expression of phage late genes, resulting in a failed infection cycle for phage K. In addition to illuminating the global transcriptional landscape of phage K throughout the infection cycle, this study will inform our investigations into the basis of phage K’s control of its transcriptional program as well as mechanisms of phage resistance. Full article