Microbial polyethylene terephthalate (PET) degradation has emerged as an environmentally friendly approach to reducing plastic pollution, but microbial activity is often limited by the high temperatures (60–70 °C) required for efficient degradation. In this study, six endogenous thermotolerant genes were identified in the PET-degrading strain Bacillus thermoamylovorans JQ3. Overexpression of the genes hrcA, hsp20A, and hsp20B significantly enhanced thermotolerance in Escherichia coli, which increased cell viability by > 24.2% at temperatures above 40 °C. Similarly, overexpression of hsp20A in B. thermoamylovorans (B. th_Hsp20A) improved its thermotolerance, increasing cell viability by 46.8% at 50 °C and 69.8% at 60 °C. The hsp20A-encoded protein was identified as a small heat shock protein (sHSP). B. th_Hsp20A exhibited significantly improved PET degradation at 60 °C compared with 50 °C, which released 282 μg of terephthalic acid after 7 days of incubation, representing an 84.3% increase compared with the wildtype strain (153 μg). These results highlight that the overexpression of thermotolerant proteins, particularly sHSPs, significantly enhances microbial PET degradation under high-temperature conditions, offering a novel pathway for enhancing microbial PET waste degradation.

Aspertetranones are a unique class of marine fungal meroditerpenoids characterized by a highly oxygenated, linear 6/6/6/6 tetracyclic core fused to an 𝛼-pyrone scaffold. Although the pathway of aspertetranone biosynthesis in Aspergillus ochraceopetaliformis has been partially elucidated, the full potential of these compounds remains untapped. The structural diversity and enzyme promiscuity of tailoring reactions offer unexplored opportunities for the generation of bioactive derivatives through combinatorial biosynthesis. In this study, we identified the atn biosynthetic gene cluster responsible for aspertetranone production in deep-sea-derived Aspergillus versicolor ADS-F20. Through the systematic heterologous expression of 12 key genes in Aspergillus oryzae, the full pathway reconstitution and targeted biosynthesis of 17 metabolites were achieved, thus expanding the known chemical space of meroterpenoids. Notably, bioactivity screening identified compound 6 as having potent antibacterial and antifungal activities against Vibrio vulnificus ATCC 27562 (MIC = 4.50 𝜇g/mL) and Phytophthora nicotianae (MIC = 9.01 𝜇g/mL). Compound 11 demonstrated broad-spectrum anticancer and cytotoxic effects against the K-562, MCF7, and PATU8988T cell lines. This study underscores the power of pathway reprogramming and catalytic network engineering as versatile strategies for expanding the structural and functional diversity of biosynthetic pathway components.

Gene knockdown is a pivotal genetic manipulation technique, particularly when targeting lethal genes or genes involved in product synthesis pathways, where complete gene knockout is not a viable option. This approach is particularly valuable in multinucleate species, such as Fusarium fujikuroi, where generating homogeneous gene knockouts is notoriously difficult. To address these limitations, we first screened a set of repression domains, and then leveraged the optimal candidates to construct a CRISPR/dCas9-mediated knockdown platform for F. fujikuroi. By targeting erg9, which encodes squalene synthase, the first committed enzyme in the mevalonate pathway for ergosterol biosynthesis, we successfully diverted a portion of the metabolic flux from sterol production to gibberellic acid (GA) biosynthesis. This strategy minimizes carbon loss to competing pathways while retaining phenotypically normal growth. Additionally, CRISPR/dCas9-mediated knockdown of the dehydrogenase gene des enhanced GA4 production by 2.62-fold and eliminated the intermediate GA7, generating a GA3 + 4-producing strain and fine-tuning its metabolic profile. Using our CRISPRi system, we achieved a 70–89 % reduction in erg9 mRNA levels and a 67– 84 % reduction in des mRNA levels. Our findings establish a tailored CRISPRi platform for effective gene repression in F. fujikuroi.

Swimming pools represent a unique aquatic environment, with continuous bacterial inoculation from swimmers and high disinfectant levels to control microbial growth. Whether, how, and which groups of bacteria can survive this strong disinfectant stress and pose a potential threat to swimmer health remain poorly understood. This study leveraged the unique opportunity of a newly opened swimming pool to address these questions. Analysis of bacterial community structures in water revealed that disinfectants substantially and temporarily simplified bacterial communities, with only a few dominant specialized groups surviving under disinfectant stress. Among the culturable pathogens, such as Escherichia coli, Staphylococcus, and Acinetobacter, only specific strains possessing genetic adaptations, such as stress-tolerant plasmids, persisted. Although most of the isolated strains did not exhibit high levels of antibiotic resistance, the presence of resistant pathogens was confirmed, one of which carried a unique complex Class I integron. This study demonstrates how bacteria emerge and persist in heavily managed, disinfected aquatic environments, such as swimming pools. Consequently, there is an urgent need to shift focus from merely reducing bacterial populations to developing sustainable strategies for managing persistent bacteria to protect public health.

Archaea of the order Sulfolobales are the earliest extremophiles discovered. They inhabit acidic terrestrial hot springs worldwide. Since their discovery, these microbes have attracted the attention of scientists studying molecular biology and microbial resources. Their evolutionary closeness to Asgard archaea and eukaryotes, the availability of genetic toolboxes for several species, and unique metabolic pathways make them ideal microbes for studying the biology of archaea and the origin of eukaryotic features, as well as platforms for synthetic biology and potential industrial applications, such as biomass degradation and bioleaching. This review summarizes recent advances in the study of Sulfolobales biology and discusses their biotechnological applications with a specific focus on biomass degradation using obligate heterotrophic Sulfolobales species.

Although ansamycins are clinically important macrolactams, their pentaketide subset exhibits limited biological activities. Genome mining of rhizosphere-derived Streptomyces sp. LR53 uncovered a cryptic pentaketide ansamycin gene cluster (tpm). Activation via 3-amino-5-hydroxybenzoic acid (AHBA) feeding, promoter replacement, and deletion of the cytochrome P450 (CYP450) gene tpm16 enabled the discovery of seven novel metabolites, tropansamycins A–G (1– 7), establishing the seventh distinct pentaketide ansamycin scaffold. Notably, congeners 5 and 6, predominantly accumulated in the Δtpm16 mutant, exhibited potent activity against gram-negative pathogenic bacterium Xanthomonas oryzae and gram-positive bacterium Staphylococcus aureus, with MICs ranging from 2 to 8 μg/mL and were toxic to the producer strain cultivated on LB agar medium. This reveals that the CYP450 enzyme Tpm16 functions as a modification enzyme modulating the bioactivity profile of pathway intermediates. These findings not only expand the structural diversity of ansamycins but also highlight the potential of these pathogen-active ansamycins as promising leads.

Actinobacteria represent a prolific source of bioactive natural products. However, the complex transcriptional regulatory networks in these bacteria, particularly the interplay between transcription factors (TFs) and their regulatory ligands (TF-RLs), remain poorly characterized and lack dedicated resources. In this context, we introduce the Actinobacteria Transcription Factor Database (Actinobacteria TFDB), a comprehensive repository that systematically integrates TF-centric data across 25 representative species. The current version encompasses 629 TFs, classified into 69 families, documents 11,776 TF-target relationships and 28 TF posttranslational modification sites. Uniquely, it features a dedicated collection of 54 experimentally validated TF-RL interactions. Beyond providing standardized annotations, sequence and structural features, and regulatory networks, Actinobacteria TFDB incorporates a specialized TF-RL module that enables interactive exploration and visualization of allosteric regulatory mechanisms. By consolidating multi-dimensional TF data from diverse sources, this resource empowers systems-level analyses and facilitates the rational design of regulatory strategies to activate silent biosynthetic gene clusters and optimize metabolite production. The database is publicly available at http://mingleadgene.com:9315/#/home.

The escalating crisis of antimicrobial resistance, particularly driven by ESKAPE pathogens, poses a serious threat to global public health. With the stagnant pipeline of novel antibiotic discovery, repurposing herbal extracts as antibiotic adjuvants to revive existing therapies is a promising strategy. This review systematically elaborates the multi-faceted mechanisms by which herbal extracts and their bioactive constituents counteract multi-drug resistance in ESKAPE pathogens. These mechanisms include the inhibition of antibiotic-degrading enzymes, alteration of bacterial membrane permeability to facilitate antibiotic uptake, blockade of major efflux pumps that expel multiple drug classes, and counteraction of antibiotic target modification. Furthermore, the roles of phytochemicals in inhibiting and dispersing biofilms, attenuating virulence factors, disrupting quorum sensing, and promoting reactive oxygen species accumulation are discussed. Despite the potent synergistic potential demonstrated by various lead compounds (such as andrographolide, quercetin, berberine, thymol, curcumin, eugenol, and oleanolic acid) in preclinical models, their progression through the translational pipeline is frequently hindered by inconsistent manufacturing standards and the inherent complexity of elucidating their polypharmacological mechanisms. Advancing the clinical utility of these herbal adjuvants necessitates a multidimensional approach, including artificial intelligence-driven screening, in-depth mechanistic studies of multi-component synergy, and comprehensive clinical assessments. Accordingly, this review aimed to establish a rigorous foundation for designing next-generation anti-infective therapies based on herb–antibiotic synergy.

Despite the considerable potential of lactic acid bacteria (LAB) as probiotics, there is a fundamental gap between the functional limitations of wild-type strains and the complex demands of aquaculture. Modular and intelligent engineering strategies are the primary avenues for bridging this gap. This article systematically reviews the strategies and advances in the application of genetically engineered LAB. Technologies, including clustered regularly interspaced short palindromic repeat (CRISPR)/Cas systems, Red/ET recombination, and functional modifications have significantly enhanced the targeted delivery, environmental tolerance, and multiple probiotic functions of LAB, successfully yielding engineered strains such as oral vaccine strains expressing pathogen antigens, antimicrobial peptide-high-yielding antibacterial strains, and nitrite-degrading water-improving strains. These engineered strains have demonstrated superior performance in disease prevention, growth promotion, and environmental remediation compared to wild-type strains in the farming of tilapia, shrimp, and shellfish. However, challenges, such as plasmid instability, biosafety risks, and regulatory barriers, remain unresolved. Future research should focus on multi-omics-guided precision design, development of environmentally responsive genetic circuits, and full-cycle risk assessment, promoting engineered LAB as a core solution for sustainable aquaculture through collaboration across industries, academia, and research.

Infections caused by bacteria pose a risk to humanity as drugs become increasingly ineffective as resistance to bacterial strains emerge along with biofilm and persister formation. This review critically evaluates host defense peptides, rational design strategies that have guided next-generation antimicrobial peptide (AMP) discovery, and their current limitations. We also highlight optimization approaches including sequence engineering and chemical modification, synergistic combinations of antibiotics or adjuvants, and nanoscale delivery platforms that enhance stability, targeted delivery, and biofilm penetration. We also discuss the key chemical properties, delivery kinetics, and stimuli-responsive drug delivery for antibacterial and antibiofilm actions as well as the toxic effects of organic- and inorganic-based AMP delivery platforms. This underlines the importance of diverse modification techniques and artificial intelligence (AI)-assisted designs to improve the antibacterial activity, stability, and biocompatibility of AMPs. This study examines the latest advances in the combination of AMPs with drug delivery systems to improve clinical outcomes. Finally, the review discusses the clinical status, research gaps, current obstacles, and prospects of AMPs in antimicrobial resistance (AMR) therapy, offering key findings for the development of innovative AMPs with significant antibacterial activity, stability, and safety for AMR treatment.