The fungal plant pathogen Verticillium dahliae is the causal agent of vascular wilt, a disease that can seriously diminish cotton fiber yield. The pathogenicity mechanism and the identity of the genes that interact with cotton during the infection process still remain unclear. In this study, we investigated the low-pathogenic, non-microsclerotium-producing mutant vdpr3 obtained in a previous study from the screening of a T-DNA insertional library of the highly virulent isolate Vd080; the pathogenicity-related gene (VdPR3) in wild-type strain Vd080 was cloned. Knockout mutants (ΔVdPR3) showed lower mycelium growth and obvious reduction in sporulation ability without microsclerotium formation. An evaluation of carbon utilization in mutants and wild-type isolate Vd080 demonstrated that mutants-lacking VdPR3 exhibited decreased cellulase and amylase activities, which was restored in the complementary mutants (ΔVdPR3-C) to levels similar to those of Vd080. ΔVdPR3 postponed infectious events in cotton and showed a significant reduction in pathogenicity. Reintroduction of a functional VdPR3 copy into ΔVdPR3-C restored the ability to infect cotton plants. These results suggest that VdPR3 is a multifunctional gene involved in growth development, extracellular enzyme activity, and virulence of V. dahliae on cotton.

Verticillium dahliae Kleb., the causal pathogen of vascular wilt, can seriously reduce the yield and quality of many crops, including cotton (Gossypium hirsutum). To control the harm caused by V. dahliae, considering the environmental pollution of chemical fungicides and their residues, the strategy of plant nutrition regulation is becoming increasingly important as an eco-friendly method for disease control. A new compound micronutrient fertilizer (CMF) found in our previous study could reduce the damage of cotton Verticillium wilt and increase yield. However, there is little information about the mode of action of CMF to control this disease. In the present study, we evaluated the role of CMF against V. dahliae and its mechanism of action in vitro and in vivo. In the laboratory tests, we observed that CMF could inhibit hyphal growth, microsclerotia germination, and reduce sporulation of V. dahliae. Further studies revealed that the biomass of V. dahliae in the root and hypocotyl of cotton seedlings treated with CMF were significantly reduced compared with the control, and these results could explain the decline in the disease index of cotton Verticillium wilt. Furthermore, those key genes involved in the phenylpropanoid metabolism pathway, resistance-related genes defense, and nitric oxide signaling pathway were induced in cotton root and hypocotyl tissue when treated with CMF. These results suggest that CMF is a multifaceted micronutrient fertilizer with roles in inhibiting the growth, development, and pathogenicity of V. dahliae and promoting cotton growth.

Verticillium wilt, caused by the notorious fungal pathogen Verticillium dahliae, is one of the main limiting factors for cotton production. Due to the stable dormant structure microsclerotia, long-term variability and co-evolution with host plant, its pathogenicity mechanism is very complicated, and the interaction mechanism between pathogen and host plant is also unclear. So identification and functional analysis of the genes involved in the pathogenicity or virulence of this fungus will benefit to uncover the molecular pathogenic mechanism of V. dahliae. In this review, many multifunction genes covering microsclerotia development, pathogen infection, effector proteins, transcription factors, horizontal gene transfer and trans-kingdom RNA silencing have been summarized to provide a theoretical basis to deep understand the molecular pathogenicity mechanism of V. dahliae and promote to effectively control Verticillium wilt. Furtherly, these pathogenicity-related genes may be considered as targets for effective control of Verticillium wilt in cotton.

Glycoside hydrolase (GH) family members act as virulence factors and regulate plant immune responses during pathogen infection. Here, we characterized the GH28 family member endopolygalacturonase VdEPG1 in Verticillium dahliae. VdEPG1 acts as a virulence factor during V. dahliae infection. The expression level of VdEPG1 was greatly increased in V. dahliae inoculated on cotton roots. VdEPG1 suppressed VdNLP1‐mediated cell death by modulating pathogenesis‐related genes in Nicotiana benthamiana. Knocking out VdEPG1 led to a significant decrease in the pathogenicity of V. dahliae in cotton. The deletion strains were more susceptible to osmotic stress and the ability of V. dahliae to utilize carbon sources was deficient. In addition, the deletion strains lost the ability to penetrate cellophane membrane, with mycelia showing a disordered arrangement on the membrane, and spore development was affected. A jasmonic acid (JA) pathway‐related gene, GhOPR9, was identified as interacting with VdEPG1 in the yeast two‐hybrid system. The interaction was further confirmed by bimolecular fluorescence complementation and luciferase complementation imaging assays in N. benthamiana leaves. GhOPR9 plays a positive role in the resistance of cotton to V. dahliae by regulating JA biosynthesis. These results indicate that VdEPG1 may be able to regulate host immune responses as a virulence factor through modulating the GhOPR9‐mediated JA biosynthesis.

In this study, the effects of the starter culture Lactobacillus sakei on pH, the total lactic acid concentration, nitrite depletion and histamine reduction of chili sauce were investigated. The nitrite concentration in the CPN sample was significantly lower (p < 0.05) than that of the CPS sample (as control), and decreased by 30.87%.The putrescine and histamine concentration in starter culture inoculated chili sauce were significantly lower (p < 0.05) than that of the control sample, and were decreased by 38.08% and 49.88%, respectively. These results revealed that L. sakei could be used as a potential starter culture for nitrite depletion and biogenic amines reduction of chili sauce with good sensory attributes.

Verticillium wilt, mainly caused by a soil-inhabiting fungus Verticillium dahliae, can seriously reduce the yield and quality of cotton. The complex mechanism underlying cotton resistance to Verticillium wilt remains largely unknown. In plants, reactive oxygen species (ROS) mediated by Rbohs is one of the earliest responses of plants to biotic and abiotic stresses. In our previous study, we performed a time-course phospho-proteomic analysis of roots of resistant and susceptible cotton varieties in response to V. dahliae, and found early differentially expressed protein burst oxidase homolog protein D (GhRbohD). However, the role of GhRbohD-mediated ROS in cotton defense against V. dahliae needs further investigation. In this study, we analyzed the function of GhRbohD-mediated resistance of cotton against V. dahliae in vitro and in vivo. Bioinformatics analysis showed that GhRbohD possessed the conservative structural attributes of Rbohs family, 12 members of RbohD out of 57 Rbohs in cotton. The expression of GhRbohD was significantly upregulated after V. dahliae inoculation, peaking at 6 hpi, and the phosphorylation level was also increased. A VIGS test demonstrated that ROS production, NO, H2O2 and Ca2+ contents of GhRbohD-silenced cotton plants were significantly reduced, and lignin synthesis and callose accumulation were damaged, important reasons for the impairment of GhRbohD-silenced cotton’s defense against V. dahliae. The expression levels of resistance-related genes were downregulated in GhRbohD-silenced cotton by qRT-PCR, mainly involving the lignin metabolism pathway and the jasmonic acid signaling pathway. However, overexpression of GhRbohD enhanced resistance of transgenic Arabidopsis to V. dahliae challenge. Furthermore, Y2H assays were applied to find that GhPBL9 and GhRPL12C may interact with GhRbohD. These results strongly support that GhRbohD activates ROS production to positively regulate the resistance of plants against V. dahliae.

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Background In our previous study, a strain EBS03 with good biocontrol potential was screened out of 48 strains of cotton endophyte Bacillus subtilis by evaluating the controlling effect against cotton Verticillium wilt. However, its mechanism for controlling Verticillium wilt remains unclear. The objective of this study was to further clarify its controlling effect and mechanism against cotton Verticillium wilt. Results The results of confrontation culture test and double buckle culture test showed that the inhibitory effects of EBS03 volatile and nonvolatile metabolite on mycelium growth of Verticillium dahliae were 70.03% and 59.00%, respectively; the inhibitory effects of sporulation and microsclerotia germination were 47.16% and 70.06%, respectively. In the greenhouse test, the EBS03 fermentation broth root irrigation had the highest controlling effect at 87.11% on cotton Verticillium wilt, and significantly promoted the growth of cotton seedlings. In the field experiment, the controlling effect of EBS03 fermentation broth to cotton Verticillium wilt was 42.54% at 60 days after cotton sowing, and the boll number per plant and boll weight in EBS03 fermentation broth seed soaking, root irrigation, and spraying treatments significantly increased by 19.48% and 7.42%, 30.90% and 2.62%, 15.99% and 9.20%, respectively. Furthermore, EBS03 improved the resistance of cotton leaves against the infection of V. dahliae, and induced the outbreak of reactive oxygen species and accumulation of callose. In addition, the results of real time fluorescent quantitative polymerase chain reaction (RT-qPCR) detection showed that EBS03 significantly induced upregulation expression level of defense-related genes PAL, POD, PPO, and PR10 in cotton leaves, enhanced cotton plant resistance to V. dahliae, and inhibited colonization level of this fungal pathogen in cotton. Conclusion Bacillus subtilis EBS03 has a good biological defense capability, which can inhibit the growth and colonization level of V. dahliae, and activate the resistance of cotton to Verticillium wilt, thus increase cotton yield.