Bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the most devastating diseases, resulting in significant yield losses in rice. The extensive use of chemical antibacterial agents has led to an increase the environmental toxicity. Nanotechnology products are being developed as a promising alternative to control plant disease with low environmental impact. In the present study, we investigated the antibacterial activity of biosynthesized chitosan nanoparticles (CSNPs) and zinc oxide nanoparticles (ZnONPs) against rice pathogen Xoo. The formation of CSNPs and ZnONPs in the reaction mixture was confirmed by using UV-vis spectroscopy at 300–550 nm. Moreover, CSNPs and ZnONPs with strong antibacterial activity against Xoo were further characterized by scanning and transmission electron microscopy, Fourier-transform infrared spectroscopy, and X-ray diffraction. Compared with the corresponding chitosan and ZnO alone, CSNPs and ZnONPs showed greater inhibition in the growth of Xoo, which may be mainly attributed to the reduction in biofilm formation and swimming, cell membrane damage, reactive oxygen species production, and apoptosis of bacterial cells. Overall, this study revealed that the two biosynthesized nanoparticles, particularly CSNPs, are a promising alternative to control rice bacterial disease.

Plants grow together with microbes that have both negative and positive impacts on the host, while prokaryotes are in turn also hosts for viruses, co-evolving together in a complex interrelationship. Most research focuses on the interaction of either bacterial pathogens interacting with the plant host, or the impact on viruses on their pathogenic bacterial hosts. Few studies have investigated the co-evolution of bacterial pathogens with their host plants as well as with their bacterial viruses. In this work, we aimed to identify the genes that were associated with both phage sensitivity and host pathogenicity of the bacterium Xanthomonas oryzae pv. oryzae (Xoo), which is the most important bacterial rice pathogen. Using the Tn5 transposon mutation technology, we created a library of Xoo strain C2 comprising 4524 mutants, which were subsequently tested for phage infectability. The phage infection tests showed that less than 1% of the mutants (n = 36) were resistant to phage infection, which was attributed to the Tn5 insertion in 19 genes. Interestingly, three out of 19 genes that conveyed resistance to the phage resulted in reduced pathogenicity to rice seedlings compared to the wild type. We identified three genes involved in both phage infection and bacterial virulence, which were studied by knockout mutants and complementation experiments. All of the three knockout mutants were resistant to infection by phage X2, while the complemented strains restored the susceptibility to the bacterial virus. Surprisingly, the genes are also essential for pathogenicity, which we confirmed by single knockout mutants corresponding to the Tn5 mutants. All three genes are involved in lipopolysaccharide synthesis, thus changing the cell envelope surface molecule composition. Our work shows a possible balance in terms of the connection between bacterial virulence and phage resistance, supporting the deployment of phages for the biocontrol of plant pathogens.

Rhizoctonia solani is the causative agent of rice sheath blight disease. In a previous study, we found that the growth of R. solani was inhibited by Burkholderia seminalis strain R456. Therefore, the present study was conducted to identify the genes involved in the antifungal activity of B. seminalis strain R456 by using a Tn5 transposon mutation method. Firstly, we constructed a random insertion transposon library of 997 mutants, out of which 11 mutants showed the defective antifungal activity against R. solani. Furthermore, the 10 antagonism-related genes were successfully identified based on analysis of the Tn5 transposon insertion site. Indeed, this result indicated that three mutants were inserted on an indigenous plasmid in which the same insertion site was observed in two mutants. In addition, the remaining eight mutants were inserted on different genes encoding glycosyl transferase, histone H1, nonribosomal peptide synthetase, methyltransferase, MnmG, sulfate export transporter, catalase/peroxidase HPI and CysD, respectively. Compared to the wild type, the 11 mutants showed a differential effect in bacteriological characteristics such as cell growth, biofilm formation and response to H₂O₂ stress, revealing the complexity of action mode of these antagonism-related genes. However, a significant reduction of cell motility was observed in the 11 mutants compared to the wild type. Therefore, it can be inferred that the antifungal mechanism of the 10 above-mentioned genes may be, at least partially, due to the weakness of cell motility. Overall, the result of this study will be helpful for us to understand the biocontrol mechanism of this bacterium.

Bacteriophage (phage) AP1 has been reported to effectively lyse Acidovorax oryzae, the causative agent of bacterial brown stripe in rice. However, phage AP1 exhibits strain-specific lysis patterns. In order to enhance the potential of phages for biological control of rice bacterial brown stripe, this study investigated the possible mechanism of strain-specific infection by characterizing phage AP1 and its susceptible (RS-2) and resistant (RS-1) strains. Based on the current classification standards and available database information, phage AP1 was classified into the class Caudoviricetes, and it is a kind of podophage. Comparative analysis of the susceptible and resistant strains showed no significant differences in growth kinetics, motility, biofilm formation, or effector Hcp production. Interestingly, the resistant strain demonstrated enhanced virulence compared to the susceptible strain. Prokaryotic expression studies indicated that six putative structural proteins of phage AP1 exhibited varying degrees of binding affinity (1.90–9.15%) to lipopolysaccharide (LPS). However, pull-down assays and bacterial two-hybrid analyses revealed that only gp66 can interact with four host proteins, which were identified as glycosyltransferase, RcnB, ClpB, and ImpB through immunoprecipitation and mass spectrometry analyses. The role of LPS in the specific infection mechanism of phage AP1 was further elucidated through the construction of knockout mutant strains and complementary strains targeting a unique gene cluster (wbzB, wbzC, wbzE, and wbzF) involved in LPS precursor biosynthesis. These findings provide novel insights into the mechanisms of phage-host specificity, which are crucial for the effective application of phage AP1 in controlling rice bacterial brown stripe.