S-ribosylhomocysteine lyase (LuxS) has been shown to regulate bacterial multicellular behaviors, typically biofilm formation. However, the mechanisms for the regulation are still mysterious. We previously identified a malonylation modification on K124 and K130 of the LuxS in the plant growth-promoting rhizobacterium B. velezensis (FZB42). In this work, we investigated the effects of the two malonylation sites on biofilm formation and other biological characteristics of FZB42. The results showed that the K124R mutation could severely impair biofilm formation, swarming, and sporulation but promote AI-2 production, suggesting inhibitory effects of high-level AI-2 on the features. All mutations (K124R, K124E, K130R, and K130E) suppressed FZB42 sporulation but increased its antibiotic production. The double mutations generally had a synergistic effect or at least equal to the effects of the single mutations. The mutation of K130 but not of K124 decreased the in vitro enzymatic activity of LuxS, corresponding to the conservation of K130 among various Bacillus LuxS proteins. From the results, we deduce that an alternative regulatory circuit may exist to compensate for the roles of LuxS upon its disruption. This study broadens the understanding of the biological function of LuxS in bacilli and underlines the importance of the two post-translational modification sites.

Phytophthora cinnamomi is a harmful microorganism that can infect Cinnamomum plants and cause the rotting of plant roots. It has been reported as infecting nearly 5000 types of plants worldwide, such as avocado trees, macadamia trees, and oak and chestnut trees, and is listed among the 10 most destructive oomycetes and the 100 most severely invasive species globally. A number of chemical agents have been applied in the control of phytophthora rot diseases because of their easy application and rapid effects. However, with the extensive use of chemical agents, P. cinnamomi has developed resistance. More importantly, it has damaged the ecological environment and affected human health. Given that biological control should be safe and effective, the screening of biocontrol strains with highly effective antagonistic effects is regarded as the primary means to control P. cinnamomi. In this study, a bacterial strain was isolated from the soil of healthy rhododendron, located near to diseased rhododendron plants, and identified as Streptomyces araujoniae, named S-03, which inhibited P. cinnamomi. The plate antagonism assay showed that S-03 could effectively bend the hyphae, reduce the number of branches, and even break them, destroying the integrity of the hyphal cell membrane to inhibit the growth of P. cinnamomi. Moreover, strain S-03 also could inhibit the activity of P. cinnamomi cell wall-degrading enzymes. To analyze the inhibitory mechanism of S-03, the effects of cell wall-degrading enzymes, secondary metabolites, and volatile substances produced by S-03 on P. cinnamomi were assessed. It was found that the fermentation broth of S-03 had a strong inhibitory effect on P. cinnamomi which means metabolites play a part in inhibition. The active substance of S-03 was initially separated and purified by ethyl acetate extraction and silica gel column chromatography and had the properties of small molecules, low polarity, and solubility in methanol. The biocontrol effect was detected in the host plants and indicated that S-03 could effectively protect rhododendron from P. cinnamomi infection. Overall, the present study findings provide compelling evidence that Streptomyces S-03 could be a biocontrol agent against plant diseases caused by P. cinnamomi.

The pine wood nematode Bursaphelenchus xylophilus parasitizes millions of pine trees worldwide each year, causing severe wilt and the death of host trees. Glycoside hydrolase 45 genes of B. xylophilus are reported to have been acquired by horizontal gene transfer from fungi and are responsible for cell wall degradation during nematode infection. Previous studies ignored the possibility of copy number variations of such genes. In this study, we determined that two of the glycoside hydrolase 45 genes evolved to maintain multiple copies with distinct expression levels, enabling the nematode to infect a variety of pine hosts. Additionally, tandem repeat variations within coding regions were also detected between different copies of glycoside hydrolase 45 genes that could result in changes in protein sequences and serve as an effective biological marker to detect copy number variations among different B. xylophilus populations. Consequently, we were able to further identify the copy number variations of glycoside hydrolase 45 genes among B. xylophilus strains with different virulence. Our results provide new insights into the pathogenicity of B. xylophilus, provide a practical marker to genotype copy number variations and may aid in population classification.