ABSTRACT
INTRODUCTION: Antimicrobial peptides (AMPs) are valuable alternatives to traditional antibiotics, possess a variety of potent biological activities and exhibit immunomodulatory effects that alleviate difficult-to-treat infections. Clarifying the structure-activity relationships of AMPs can direct the synthesis of desirable peptide therapeutics. OBJECTIVES: In this study, the lipopolysaccharide-binding domain (LBD) was identified through machine learning-guided directed evolution, which acts as a functional domain of the anti-lipopolysaccharide factor family of AMPs identified from Marsupenaeus japonicus. METHODS: LBDA-D was identified as an output of this algorithm, in which the original LBDMj sequence was the input, and the three-dimensional solution structure of LBDB was determined using nuclear magnetic resonance. Furthermore, our study involved a comprehensive series of experiments, including morphological studies and in vitro and in vivo antibacterial tests. RESULTS: The NMR solution structure showed that LBDB possesses a circular extended structure with a disulfide crosslink at the terminus and two 310-helices and exhibits a broad antimicrobial spectrum. In addition, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) showed that LBDB induced the formation of a cluster of bacteria wrapped in a flexible coating that ruptured and consequently killed the bacteria. Finally, coinjection of LBDB, Vibrio alginolyticus and Staphylococcus aureus in vivo improved the survival of M. japonicus, demonstrating the promising therapeutic role of LBDB for treating infectious disease. CONCLUSIONS: The findings of this study pave the way for the rational drug design of activity-enhanced peptide antibiotics.
ABSTRACT
Panax ginseng, a prized medicinal herb, has faced increasingly challenging field production due to soil degradation and fungal diseases in Northeast China. Wild-simulated cultivation has prevailed because of its sustainable soil management and low disease incidence. Despite the recognized benefits of rhizosphere microorganisms in ginseng cultivation, their genomic and functional diversity remain largely unexplored. In this work, we utilized shotgun metagenomic analysis to reveal that Pseudomonadota, Actinomycetota, and Acidobacteriota were dominant in the ginseng rhizobiome and recovered 14 reliable metagenome-assembled genomes. Functional analysis indicated an enrichment of denitrification-associated genes, potentially contributing to the observed decline in soil fertility, while genes associated with aromatic carbon degradation may be linked to allelochemical degradation. Further analysis demonstrated enrichment of Actinomycetota in 9-year-old wild-simulated ginseng (WSG), suggesting the need for targeted isolation of Actinomycetota bacteria. Among these, at least three different actinomycete strains were found to play a crucial role in fungal disease resistance, with Streptomyces spp. WY144 standing out for its production of actinomycin natural products active against the pathogenic fungus Ilyonectria robusta. These findings not only enhance our understanding of the rhizobiome of WSG but also present promising avenues for combating detrimental fungal pathogens, underscoring the importance of ginseng in both medicinal and agricultural contexts.IMPORTANCEWild-simulated ginseng, growing naturally without human interference, is influenced by its soil microbiome. Using shotgun metagenomics, we analyzed the rhizospheric soil microbiome of 7- and 9-year-old wild-simulated ginseng. The study aimed to reveal its composition and functions, exploring the microbiome's key roles in ginseng growth. Enrichment analysis identified Streptomycetes in ginseng soil, with three strains inhibiting plant pathogenic fungi. Notably, one strain produced actinomycins, suppressing the ginseng pathogenic fungus Ilyonectria robusta. This research accelerates microbiome application in wild-simulated ginseng cultivation, offering insights into pathogen protection and supporting microbiome utilization in agriculture.
