Shengbiao Hu

This study addresses the core challenge of Fusarium wilt control in agricultural production. We successfully reconstituted a functional heterologous type III secretion system (T3SS) from Photorhabdus luminescens in the biocontrol bacterium Pseudomonas protegens Pf‐5, creating an engineered molecular syringe for targeted delivery of antifungal effectors. The system is activated under low‐calcium conditions, achieved by cultivation in calcium‐limited medium followed by EGTA‐mediated chelation of residual Ca ²⁺ , enabling conditional secretion of effector proteins. By fusing the antifungal protein Bg9562 to the N‐terminal secretion signal of the T3SS effector LopT and co‐expressing it with the cognate chaperone SlcT, we obtained fluorescence‐based evidence for T3SS‐dependent delivery of Bg9562 into the hyphae of multiple Fusarium species. The engineered strain exhibited enhanced rhizosphere colonization, promoted plant growth and conferred improved protection against tomato Fusarium wilt, restoring plant height to levels approaching healthy controls. We further demonstrated the modularity of this platform by successfully transferring it into Pseudomonas koreensis D26, a strain known for its plant growth‐promoting properties, indicating broad applicability across biocontrol‐relevant pseudomonads. This work establishes a versatile T3SS‐based delivery platform for precision biocontrol, offering a generalizable strategy for engineering beneficial rhizobacteria.

Aeromonas veronii is a representative pathogen resulting in significant economic losses in aquaculture. As a potential antibacterial agent, bacteriophage has attracted wide attention. Herein, a virulent phage named Avs-1 that infects Aeromonas veronii was isolated from sewage. Avs-1 demonstrated an optimal multiplicity of infection (MOI) of 0.1, with a 20-min latent period and a 130-min burst period. Cryo-electron microscopy structural analysis revealed that Avs-1 possesses an icosahedral capsid composed of 415 copies of the major capsid protein arranged in a T = 7 lattice, along with a long non-contractile tail typical of the Siphoviridae family. Phage resistance in strain AV0110 was linked to a frameshift mutation in the manB gene encoding phosphomannomutase (PMM). The genome of phage Avs-1 consists of 44,364 bp double-stranded circular DNA, in which the endolysin (Lys40) has a modular structure with a N-acetylmuramidase domain at N-terminus and a peptidoglycan (PG)-binding domain at C-terminus. Notably, endogenous expression of Lys40 could greatly inhibit the growth of E. coli but could not lyse them. Avs-1 exhibited significant antibacterial effect in vitro and increased tilapia survival rate by 50% when used in vivo . This study provides a theoretical basis for further research on the interaction between pathogens and phages, and the prevention and treatment of Aeromonas veronii and highlights the potential of Avs-1 as a targeted biocontrol agent against Aeromonas veronii infections in aquaculture. IMPORTANCE: In this study, we present an eco-friendly approach to address a critical issue in aquaculture: Aeromonas veronii infections, which cause significant economic losses. We turned to nature’s bacterial predators—bacteriophages and viruses that specifically infect and kill bacteria. From sewage samples, we isolated a novel phage named Avs-1, which effectively targets A. veronii . Genomic and structural analyses were conducted to elucidate its mechanisms. Notably, we also identified a key gene that enabled bacteria to resist phage adsorption, which provides critical insights for future phage applications. Most importantly, in our laboratory tests, the Avs-1 bacteriophage demonstrated potent antibacterial activity and increased the survival rate of tilapia infected with A. veronii by 50%. The potential of Avs-1 bacteriophage as a targeted, environmentally friendly “biological pesticide” for controlling infections in aquaculture offers a promising alternative to conventional antibiotics.

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.

The type III secretion system (T3SS) of bacterial pathogens plays an essential role in infection and colonization processes. T3S effectors (T3SEs) are pivotal in mediating these interactions and their mechanisms of action. This study delves into the functional mechanisms of XopA, the first T3SE identified in the bacterium Xenorhabdus , which belongs to the YopJ family. XopA demonstrates cytotoxicity akin to other YopJ family members and possesses virulence determinants capable of inducing both apoptosis and autophagy. Notably, our findings reveal a complex regulatory network between XopA-induced apoptosis and autophagy. Moreover, XopA modulates the host cell’s global and inflammatory responses by targeting tubulin, thereby affecting cytoskeletal dynamics and the secretion of extracellular vesicles (EVs). The acetylation activity characteristic of the YopJ family effectors is significantly altered in HeLa cells upon XopA action, highlighting its role in post-translational modifications. Collectively, this study elucidates the multifaceted functional mechanisms of XopA, which will undoubtedly be beneficial for a better understanding of the molecular mechanisms of Xenorhabdus pathogenesis. IMPORTANCE: This study reports the groundbreaking discovery of XopA as the first type III secretion system effectors (T3SE) identified in Xenorhabdus bacteria. By demonstrating its unique ability to concurrently induce host cell apoptosis and autophagy, execute lysine acetyltransferase activity to suppress inflammatory signaling, and disrupt cytoskeletal dynamics to inhibit extracellular vesicle secretion, this work reveals a sophisticated multifunctional virulence mechanism. These findings significantly advance our understanding of bacterial pathogenesis, providing crucial insights into how T3SEs manipulate host cell processes and evade immune responses, thereby establishing a new frontier in host-pathogen interaction research.

Introduction: The emergence of multi-drug resistant Shewanella xiamenensis poses a significant threat to aquaculture and public health, driving the need for alternative therapies like bacteriophages.Methods: A novel bacteriophage, SXP01, was isolated using S. xiamenensis A001 as a host. Its kinetic parameters (MOI, latent period) and stability across temperature (4-60°C), pH (6-11), and chloroform were characterized. The lysis cassette was identified genomically, and a recombinant endolysin (LysSXP-1) was expressed to test lytic activity. In vivo efficacy was evaluated in a crucian carp infection model.Results: SXP01, a Myoviridae member, showed an optimal MOI of 0.1, a 20-minute latent period, and high environmental stability. Its lysis cassette encodes two endolysins, with LysSXP-1 demonstrating direct lytic activity. In the fish model, SXP01 treatment significantly increased survival rates from 13.3% to 56.7%.Discussion: The study identifies SXP01 as a highly stable and effective bacteriophage. The synergistic action of its endolysins and its demonstrated in vivo efficacy highlight its strong potential as a targeted biocontrol agent to manage S. xiamenensis infections in aquaculture sustainably.

Engineered living materials (ELMs) integrate living cells—such as bacteria, yeast, or mammalian cells—with synthetic matrices to create responsive, adaptive systems for sensing and actuation. Among ELMs, those endowed with sensing capabilities are gaining increasing attention for applications in environmental monitoring, biomedicine, and smart infrastructure. Central to these sensing functions are synthetic gene circuits, which enable cells to detect and respond to specific signals. This mini-review focuses on recent advances in sensing ELMs empowered by synthetic gene circuits. Here, we highlight how rationally designed genetic circuits enable living materials to sense and respond to diverse inputs—including environmental chemicals, light, heat, and mechanical loadings—via programmable signal transduction and tailored output behaviors. Input signals are classified by their source and physicochemical properties, including synthetic inducers, environmental chemicals, light, thermal, mechanical, and electrical signals. Particular emphasis is placed on the integration of genetically engineered microbial cells with hydrogels and other functional scaffolds to construct robust and tunable sensing platforms. Finally, we discuss the current challenges and future opportunities in this rapidly evolving field, providing insights to guide the rational design of next-generation sensing ELMs.

Microorganisms are often likened to complex production workshops. In Saccharopolyspora spinosa (S. spinosa), the biosynthesis of spinosad is a production line within its intricate workshop. Optimizing the entire production environment and reducing unnecessary metabolic flow is essential to increasing spinosad yield. Pyruvate serves as a crucial precursor for spinosad biosynthesis. Previous studies revealed that the pyc gene is highly expressed in the gluconeogenic pathway, leading to a pyruvate shunt. By downregulating pyc, we enhanced spinosad yield, although the improvement was below expectations. We speculated that most of the accumulated pyruvate following the pyc knockdown entered some synthetic pathways unrelated to spinosad. Through metabolic pathway and qRT-PCR analyses, we found that the expression levels of gltA1 and atoB3 within the pyruvate metabolic tributary, including the TCA cycle and ethylmalonyl-CoA pathway, were significantly increased in the pyc knockdown strain. The combined knockdown of these three genes optimized the spinosad production line, increasing its yield to 633.1 ± 38.6 mg/L, representing a 199.4 % increase. This study identifies three key genes for optimizing spinosad biosynthesis and offers insights into gene screening and the efficient construction of Spinosad-producing strains.

Remodelling regulatory pathways to directionally increase the efficiency of specific promoters in chassis cells is an effective strategy for the rational construction of expression systems. However, the repeated utilization of one regulator to modify the host cell to improve expression motif efficiency has a limited effect. Therefore, it is preferable to identify new regulatory factors to activate specific pathways and thus further improve the efficiency of target elements. Heterochromatin protein 1 (HP1) is considered a main factor responsible for heterochromatin maintenance; it binds DNA and thus forms a tight structure to repress gene expression in fungi. This study revealed that the overexpression of HepA (a homologue of HP1) increased amylase expression in Penicillium oxalicum. Furthermore, HepA was overexpressed in two engineered strains in which the endoglucanase TaEG and amylase Amy15B were recombinantly expressed under the control of the amylase promoter Pamy15A, resulting in increased production of these two enzymes. Therefore, HepA could be used as a novel facilitator to modify Penicillium chassis cells, in which the efficiency of expression motifs located in the amylase pathway can be further strengthened.