Yinsheng Li

Antimicrobial peptides (AMPs) exhibit diverse antibacterial mechanisms. To elucidate the intracellular targets of the novel AMP, EWAMP-R, we generate EWAMP-R-resistant strains of Escherichia coli and Staphylococcus aureus. In E. coli, a nonsynonymous mutation in the clpX gene (1108 C > T) is identified, resulting in an amino acid change in ClpX (R370C). This mutation confers enhanced tolerance to EWAMP-R and induces a “hyperflagellate phenotype” in EWAMP-R-resistant E. coli, as verified by CRISPR/Cas9-generated clpX (1108 C > T) mutants. Further studies reveal that EWAMP-R can bind to the Walker A motif and inhibit the conformational changes of ClpX, suppressing its ATPase activity and the degradation of its substrates, such as LexA 1-84. The accumulation of LexA 1-84 downregulates the SOS response, inducing apoptosis-like death in E. coli. In contrast, EWAMP-R-resistant S. aureus exhibits only a metabolic slowdown. These findings establish ClpX as a novel target for AMPs and highlight EWAMP-R as a promising antibacterial agent.

Earthworm behavioral biomarkers provide more sensitive and comprehensive assessments for environmental contaminants. However, natural soil opaqueness has limited understanding of the continuous three-dimensional (3D) characteristics of earthworm burrowing activity. In this study, we developed an integrative method combining a transparent medium with a deep learning-based trajectory reconstruction algorithm to quantitatively investigate species-specific burrowing strategies. Earthworms thrived in the transparent medium, as shown by biomass increase (0.62 to 0.72 g) and >80% survival over the 28-day trial, while algorithm operational stability (83.65% coordinate extraction rate) allowed continuous tracking of the earthworm spatial coordinates in 3D locations. Comparative analysis across five earthworm species revealed species-specific patterns: Eisenia fetida, Amynthas pectiniferus, and Eudrilus eugeniae preferred surface layers and moved laterally, whereas Perionyx excavatus and Metaphire guillelmi created vertical burrows. These findings are valuable for quantifying soil health assessments using different earthworm species. Moreover, this method successfully captured avoidance behavior in arsenate-contaminated soils, as arsenic exposure inhibited cumulative burrowing length. These results establish a critical tool for advancing ecological risk assessment and management of soil pollutants by offering visual and quantitative insights into 3D earthworm burrowing dynamics, and provide potential applications in ecology, environmental pollution monitoring, and remediation.

Microbial inoculants show potential for remediating nitrate-rich, salinized soils. However, native strains often exhibit suboptimal performance under high-salinity and high-nitrate conditions, limiting their practical application. To develop a nitrate-reducing strain suitable for saline soil remediation, we adaptively acclimated a nitrate-reducing Bacillus velezensis strain BV-1 under high-nitrate conditions to enhance its salt tolerance and nitrogen metabolic capacity. The acclimatized strain exhibited significantly upregulated nar genes (4.71- to 7.56-fold) and nirD expression (1.36-fold), indicating enhanced nitrate assimilation and dissimilatory nitrate reduction to ammonium activity. In pot experiments, inoculation with this strain resulted in 46.85% nitrate removal, improved nutrient utilization (with increases of 21.86% in ammonium-N and 29.64% in available phosphorus utilization), and a 20.82% increase in lettuce fresh weight. These findings demonstrate that microbial acclimatization is an effective strategy for developing robust bioinoculants, with broad implications for sustainable agriculture and microbial strain engineering in salinized environments.

Fusarium wilt is a devastating soilborne disease that significantly reduces watermelon production worldwide. This disease is caused by Fusarium oxysporum subsp. niveum (E.F.Sm.) W.C. Snyder & H.N.Hansen. Earthworms can influence fungal populations either by consuming or dispersing fungal propagules, making them a promising candidate for the biological control of Fusarium wilt. However, the underlying mechanisms remain poorly understood. In this study, we investigated the effects of adding the local earthworm species Metaphire guillelmi (Michaelsen, 1895) on Fusarium wilt in watermelon under field conditions, laboratory pot experiments, and laboratory pot experiments with sterilized soil. The results demonstrated that, compared to the control, the earthworm addition reduced the population of F. oxysporum by approximately 10⁵ copies/mg and suppressed the incidence of Fusarium wilt by 84.4%. A correlation analysis revealed that the abundance of F. oxysporum was negatively correlated with soil organic matter (SOM), available nitrogen (AN), and available phosphorus (AP). The relative interaction index values indicated that earthworms could enhance SOM and AN levels in the soil. A two-factor network relationship analysis showed that the earthworm addition could inhibit bacteria and fungi to stimulate growth of F. oxysporum while restraining them. A metabolomics analysis revealed that most differential metabolites associated with F. oxysporum were upregulated in the presence of earthworms. Overall, M. guillelmi can reduce the occurrence of Fusarium wilt by improving soil fertility, the relationship of F. oxysporum and microorganisms, and may influence the metabolic process, which need further exploration. It is a potential pathway for the biocontrol of Fusarium wilt.

Natural rubber is an important national strategic and industrial raw material. The leaf anthracnose of rubber trees caused by the Colletotrichum species is one of the important factors restricting the yields of natural rubber. In this study, we isolated and identified strain Bacillus velezensis SF334, which exhibited significant antagonistic activity against both C. australisinense and C. siamense, the dominant species of Colletotrichum causing rubber tree leaf anthracnose in the Hainan province of China, from a pool of 223 bacterial strains. The cell suspensions of SF334 had a significant prevention effect for the leaf anthracnose of rubber trees, with an efficacy of 79.67% against C. siamense and 71.8% against C. australisinense. We demonstrated that SF334 can lead to the lysis of C. australisinense and C. siamense mycelia by causing mycelial expansion, resulting in mycelial rupture and subsequent death. B. velezensis SF334 also harbors some plant probiotic traits, such as secreting siderophore, protease, cellulase, pectinase, and the auxin of indole-3-acetic acid (IAA), and it has broad-spectrum antifungal activity against some important plant pathogenic fungi. The genome combined with comparative genomic analyses indicated that SF334 possesses most genes of the central metabolic and gene clusters of secondary metabolites in B. velezensis strains. To our knowledge, this is the first time a Bacillus velezensis strain has been reported as a promising biocontrol agent against the leaf anthracnose of rubber trees caused by C. siamense and C. australisinense. The results suggest that B. velezensis could be a potential candidate agent for the leaf anthracnose of rubber trees.