Lysis inhibition (LIN) in bacteriophage is a strategy to maximize progeny production. A clear plaque-forming mutant, CSP1C, was isolated from the turbid plaque-forming CSP1 phage. CSP1C exhibited an adsorption rate and replication dynamics similar to CSP1. Approximately 90% of the phages were adsorbed to the host cell within 12 min, and both phages had a latent period of 25 min. Burst sizes were 171.42 ± 31.75 plaque-forming units (PFU) per infected cell for CSP1 and 168.94 ± 51.67 PFU per infected cell for CSP1C. Both phages caused comparable reductions in viable E. coli cell counts at a low multiplicity of infection (MOI). However, CSP1 infection did not reduce turbidity, suggesting a form of LIN distinct from the well-characterized LIN of T4 phage. Genomic analysis revealed that a 4,672-base pairs (bp) DNA region, encompassing part of the tail fiber gene, CSP1_020, along with three hypothetical genes, CSP1_021, CSP1_022, and part of CSP1_023, was deleted from CSP1 to make CSP1C. Complementation analysis in CSP1C identified CSP1_020, CSP1_021, and CSP1_022 as a minimal gene set required for the lysis suppression in CSP1. Co-expression of these genes in E. coli with holin (CSP1_092) and endolysin (CSP1_091) resulted in lysis suppression. Lysis suppression was abolished by disrupting the proton motive force (PMF), supporting their potential role as antiholin. Additionally, CSP1_021 directly interacts with holin, suggesting that it may function as an antiholin. These findings identify new genetic factors involved in lysis suppression in CSP1, providing broader insights into phage strategies for modulating host cell lysis.

Candida albicans (C. albicans) is a common opportunistic fungal pathogen that can cause infections ranging from superficial to severe systemic diseases. This study investigates the antifungal effects of metformin on C. albicans and explores its underlying mechanisms. Growth inhibition was assessed via XTT assays, and hyphal formation and morphological changes were observed by light microscope and scanning electron microscopy (SEM). Mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) levels were measured with JC-1 and DCFH-DA probes, respectively. Gene expression related to ROS and autophagy was quantified by RT-qPCR, and autophagosomes were visualized using transmission electron microscopy (TEM). Metformin significantly inhibited C. albicans growth and hyphal formation, altered cell morphology, reduced MMP, and increased ROS levels. It activated autophagy in planktonic C. albicans but suppressed it in biofilm forms. Additionally, metformin exhibited synergistic effects with amphotericin B against planktonic C. albicans and with caspofungin against biofilms. The findings suggest that metformin exerts antifungal activity by modulating MMP, ROS levels, and autophagy-related pathways, and enhances the efficacy of specific antifungal drugs.

The increasing environmental concerns regarding conventional plastics have led to a growing demand for sustainable alternatives, such as biodegradable plastics. Yeast cell factories, specifically Saccharomyces cerevisiae and Yarrowia lipolytica, have emerged as promising platforms for bioplastic production due to their scalability, robustness, and ease of manipulation. This review highlights synthetic biology approaches aimed at developing yeast cell factories to produce key biodegradable plastics, including polylactic acid (PLA), polyhydroxyalkanoates (PHAs), and poly (butylene adipate-co-terephthalate) (PBAT). We explore recent advancements in engineered yeast strains that utilize various synthetic biology strategies, such as the incorporation of new genetic elements at the gene, pathway, and cellular system levels. The combined efforts of metabolic engineering, protein engineering, and adaptive evolution have enhanced strain efficiency and maximized product yields. Additionally, this review addresses the importance of integrating computational tools and machine learning into the Design-Build-Test-Learn cycle for strain development. This integration aims to facilitate strain development while minimizing effort and maximizing performance. However, challenges remain in improving strain robustness and scaling up industrial production processes. By combining advanced synthetic biology techniques with computational approaches, yeast cell factories hold significant potential for the sustainable and scalable production of bioplastics, thus contributing to a greener bioeconomy.

Methane gas is recognized as a promising carbon substrate for the biosynthesis of value-added products due to its abundance and low price. Methanotrophs utilized methane as their sole source of carbon and energy, thus they can serve as efficient biocatalysts for methane bioconversion. Methanotrophs-catalyzed microbial bioconversion offer numerous advantages, compared to chemical processes. Current indirect chemical conversions of methane suffer from their energy-intensive processes and high capital expenditure. Methanotrophs can be cell factories capable of synthesizing various value-added products from methane such as methanol, organic acids, ectoine, polyhydroxyalkanoates, etc. However, the large-scale commercial implementation using methanotrophs remains a formidable challenge, primarily due to limitations in gas-liquid mass transfer and low metabolic capacity. This review explores recent advancements in methanotroph research, providing insights into their potential for enabling methane bioconversion.

Phage specificity primarily relies on host cell-surface receptors. However, integrating cas genes and guide RNAs into phage genomes could enhance their target specificity and regulatory effects. In this study, we developed a CRISPR-Cas12f1 system-equipped bacteriophage λ model capable of detecting Escherichia coli target genes. We demonstrated that synthetic λ phages carrying Cas12f1-sgRNA can effectively prevent lysogen formation. Furthermore, we showcased that truncating the 3'-end of sgRNA enables precise identification of single-nucleotide variations in the host genome. Moreover, infecting E. coli strains carrying various stx2 gene subtypes encoding Shiga toxin with bacteriophages harboring Cas12f1 and truncated sgRNAs resulted in the targeted elimination of strains with matching subtype genes. These findings underscore the ability of phages equipped with the CRISPR-Cas12f1 system to precisely control microbial hosts by recognizing genomic sequences with high resolution.

Pyridoxal 5'-phosphate (PLP)-dependent enzymes participate in various reactions involved in methionine and cysteine metabolism. The representative foodborne pathogen Staphylococcus aureus expresses the PLP-dependent enzyme MccB, which exhibits both cystathionine gamma-lyase (CGL) and cysteine desulfhydrase activities. In this study, we investigated the role of Ser323 in MccB, a conserved residue in many PLP-dependent enzymes in the transsulfuration pathway. Our findings reveal that Ser323 forms a hydrogen bond with the catalytic lysine in the absence of PLP, and upon internal aldimine formation, PLP-bound lysine is repositioned away from Ser323. Substituting Ser323 with alanine abolishes the enzymatic activity, similar to mutations at the catalytic lysine site. Spectroscopic analysis suggests that Ser323 is essential for the rapid formation of the internal aldimine with lysine in wild-type MccB. This study highlights the crucial role of Ser323 in catalysis, with broader implications for other PLP-dependent enzymes, and enhances our understanding of the molecular mechanisms involved in the selective control of foodborne pathogenic bacteria.