Powdery mildew-induced changes in phyllosphere microbial community dynamics of cucumber.

As an important habitat for microorganisms, the phyllosphere has a great impact on plant growth and health, and changes in phyllosphere microorganisms are closely related to the occurrence of leaf diseases. However, there remains a limited understanding regarding alterations to the microbial community in the phyllosphere resulting from pathogen infections. Here, we analyzed and compared the differences in phyllosphere microorganisms of powdery mildew cucumber from three disease severity levels (0% < L1 < 30%, 30% ≤ L2 < 50%, L3 ≥ 50%, the number represents the lesion coverage rate of powdery mildew on leaves). There were significant differences in α diversity and community structure of phyllosphere communities under different disease levels. Disease severity altered the community structure of phyllosphere microorganisms, Rosenbergiella, Rickettsia, and Cladosporium accounted for the largest proportion in the L1 disease grade, while Bacillus, Pantoea, Kocuria, and Podosphaera had the highest relative abundance in the L3 disease grade. The co-occurrence network analysis of the phyllosphere microbial community indicated that the phyllosphere bacterial community was most affected by the severity of disease. Our results suggested that with the development of cucumber powdery mildew, the symbiotic relationship between species was broken, and the entire bacterial community tended to compete.

Cucumis sativus PHLOEM PROTEIN 2-A1 like gene positively regulates salt stress tolerance in cucumber seedlings.

PHLOEM PROTEIN 2-A1 like (PP2-A1) gene is a member of the PP2 multigene family, and the protein encoded by which has the function of stress defense. Based on our previous proteomic study of cucumber phloem sap, CsPP2-A1 protein expression was significantly enriched under salt stress. In this paper, we obtained CsPP2-A1 interfering (CsPP2-A1-RNAi) cucumber by Agrobacterium tumefaciens-mediated method. The phenotypic changes of wild-type (WT) cucumber, CsPP2-A1-overexpressing (OE) cucumber, and CsPP2-A1-RNAi cucumber under salt treatment were observed and compared. Furthermore, physiological indicators were measured in four aspects: osmoregulation, membrane permeability, antioxidant system, and photosynthetic system. The analysis of contribution and correlation for each variable were conducted by principal component analysis (PCA) and Pearson's correlation coefficient. The above results showed that CsPP2-A1-RNAi cucumber plants exhibited weaker salt tolerance compared to WT cucumber and CsPP2-A1-OE cucumber plants in terms of phenotype and physiological indicators in response to salt stress, while CsPP2-A1-OE cucumber always showed the robust salt tolerance. Together, these results indicated that CsPP2-A1 brought a salinity tolerance ability to cucumber through osmoregulation and reactive oxygen species (ROS) homeostasis. The results of the study provided evidence for the function of CsPP2-A1 in plant salt tolerance enhancement, and they will serve as a reference for future salt-tolerant cucumber genetic manipulation.

The phloem lectin PP2-A1 enhances aphid resistance by affecting aphid behavior and maintaining ROS homeostasis in cucumber plants.

Aphid (Aphis gossypii Glover) attack frequently results in a significant loss of output and deterioration of fruit quality in cucumber (Cucumis sativus L.). Phloem protein 2 (PP2) is conserved as a phloem lectin in plants, and few studies have been conducted on the regulatory mechanism of PP2. Based on our previous study of CsPP2-A1 in cucumber, to further investigate the biological function of CsPP2-A1, we compared the changes of selectivity, non-selectivity, colonization, reproductions of aphids, and the phenotype in wild type (WT), CsPP2-A1 overexpressing (CsPP2-A1-OE), and CsPP2-A1 interfering (CsPP2-A1-RNAi) cucumber plants after inoculation with aphids. We found that CsPP2-A1-OE cucumber plants generated resistance to aphids. The aphid colonization rate and number of reproductions of CsPP2-A1-OE cucumber plants were significantly lower than that of WT and CsPP2-A1-RNAi cucumber plants. Through Pearson's correlation and principal component analysis (PCA), it was found that CsPP2-A1 played a crucial role in the balance of reactive oxygen species (ROS) in plants. Overexpression of the CsPP2-A1 resulted in increased levels of antioxidant enzyme, eliminating ROS and preventing the damage by ROS in cucumber. Furthermore, nutritional imbalance for aphids and content of secondary metabolites were increased in overexpressed CsPP2-A1 cucumber plants, and thus preventing aphid attack. These together may improve cucumber resistance against aphids and the mechanism of CsPP2-A1 defense against aphids was preliminarily explored.

Exogenously applied melatonin alleviates the damage in cucumber plants caused by Aphis goosypii through altering the insect behavior and inducing host plant resistance.

BACKGROUND: Aphis gossypii Glover is the main pest found in most cucumber-producing areas. Melatonin (MT) has been widely studied in protecting plants from environmental stresses and pathogens. However, little knowledge is available on the impact of MT on insect resistance. RESULTS: The fecundity of aphids on MT-treated cucumber leaves was inhibited. Interestingly, MT-treated plants were more attractive to aphids, which would prevent the large-scale transmission of viruses caused by the random movement of aphids. Meanwhile, MT caused varying degrees of change in enzyme activities related to methylesterified HG degradation, antioxidants, defense systems and membrane lipid peroxidation. Furthermore, transcriptomic analysis showed that MT induced 2360 differentially expressed genes (DEGs) compared with the control before aphid infection. These DEGs mainly were enriched in hormone signal transduction, MAPK signaling pathway, and plant-pathogen interaction, revealing that MT can help plants acquire inducible resistance and enhance plant immunity. Subsequently, 2397 DEGs were identified after aphid infection. Further analysis showed that MT-treated plants possessed stronger JA signal, reactive oxygen species stability, and the ability of flavonoid synthesis under aphid infection, while mediating plant growth and sucrose metabolism. CONCLUSION: In summary, MT as an environmentally friendly substance mitigated aphid damage to cucumbers by affecting the aphids themselves and enhancing plant resistance. This will facilitate exploring sustainable MT-based strategies for cucumber aphid control. © 2022 Society of Chemical Industry.

Exogenous melatonin improves the resistance to cucumber bacterial angular leaf spot caused by Pseudomonas syringae pv. Lachrymans.

Pseudomonas syringae pv. Lachrymans (Psl) is a bacterial pathogen that causes cucumber bacterial angular leaf spot (BALS). It is known that melatonin (MT), as a pleiotropic signal molecule, can improve plant stress tolerance, but less information is available about the function of MT on plant resistance to bacteria disease. Here, we investigated the effect of MT on cucumber BALS. Our results show that MT inhibited the bacteria Psl growth significantly in vitro and attenuated cucumber BALS remarkably in vivo. The concentration of bacteria in leaves treated with 0.1 mM MT was approximately 10,000 times reduced at 5 days-post-inoculation (dpi), compared to the control without MT. Transcriptomic analysis showed that 225 differentially expressed genes (DEGs) were induced in leaves after just MT treatment for 3 h. The functions of these DEGs were mainly associated with hormone signal transduction, mitogen-activated protein kinase (MAPK) signaling pathway, and photosynthesis, suggesting that MT could regulate plant growth and induce plant immunity. Moreover, 665 DEGs were induced when leaves were treated with exogenous MT in combination with the bacteria inoculation for 12 h. The functions of these DEGs were much related to plant-pathogen interaction, hormone signal transduction, and amino acids biosynthesis pathways. Many MT-induced DEGs were involved in some distinct signal transduction pathways, such as calmodulin (CaM), polyamines (PAs), nitric oxide (NO), and salicylic acid (SA). The physiological analysis shows that exogenous MT spray reduced the stomatal aperture and enhanced the activities of antioxidant and defense enzymes, which were in consistent with the results of the transcriptome analysis. In addition, MT may function in regulating the metabolic balance between plant growth and defense. In conclusion, our results demonstrate that MT could alleviate the cucumber BALS via inhibiting propagation and invasion of Psl, activating plant signaling, enhancing antioxidative and defense systems, inducing stress-related genes expression, and regulating the plant growth-defense balance.

Prokaryotic expression, purification, physicochemical properties and antifungal activity analysis of phloem protein PP2-A1 from cucumber.

Phloem protein 2 (PP2) is a protein having lectin properties that can be isolated from the phloem sap. Based on our previous proteomic study of phloem sap of Cucumis sativus, it was found that the expression of PP2 A1-like was significantly up-regulated under salt stress, which may be a molecular mechanism of plant adaptation to stress. This paper carried out the expression and purification of the CsPP2-A1 gene in E. coli for further characteristic analysis. The results demonstrated that the CsPP2-A1 in shake flask cultures was mainly expressed in the soluble form at 15 °C or in inclusion bodies at 37 °C. Secondly, Ni-IDA affinity chromatography and SDS-PAGE were employed to yield highly purified CsPP2-A1 protein. The purified CsPP2-A1 was then subjected to Western blot and MALDI-TOF-MS analysis for protein identification. The biological activity analysis results showed that CsPP2-A1 had hemagglutinating activities to rabbit erythrocytes, and Chitotetraose may be the specific inhibitory sugar of CsPP2-A1. The optimal hemagglutination activity of CsPP2-A1 protein was achieved between pH 5-9, and between 20 and 60 °C. Moreover, CsPP2-A1 had significant inhibitory effects on Botrytis cinerea and Phytophthora infestans, and the inhibitory effect on B. cinerea was better than that on P. infestans.

Understanding of the postgerminative development response to salinity and drought stresses in cucumber seeds by integrated proteomics and transcriptomics analysis.

The postgerminative development is a complex, genetically programmed process, and also the most dangerous period before the developing seedlings reach the autotrophy state. To obtain a comprehensive understanding of postgerminative development mechanism, the study focuses on an integrative analysis on transcriptome, proteome, and microRNA in cucumber seeds under drought and salt stress. Drought and salt stress caused differential expression of 4197 mRNAs, 36 microRNAs and 768 proteins compared with the control, and 827 mRNAs, 364 proteins, and 12 microRNAs were shared by the two stresses. Numerous common differentially expressed genes and proteins participated the signal transduction of plant hormone, photosynthesis, and argine and proline metabolism. We noted the correlation among nitric oxide, polyamine, proline, and ethylene metabolism, thereby helping to elucidate the role of these substances, which are derived either directly or indirectly from arginine, in the regulation of abiotic stress and provide a basis for building better network-based molecular models in further research. Above findings contribute to new and useful information regarding the common molecular mechanisms during cucumber seedling development under drought and salt stress. SIGNIFICANCE: Water scarcity and high salt are two of the most destructive and wide stress factors which limit the growth and progression of plants by affecting a variety of vital physiological and biochemical processes. Our study focuses on an integrative analysis on transcriptome, proteome, and microRNA for confirming the essential regulators as well as pathways using cucumber postgerminative development under drought and salt stress. Arginine metabolism is a vital response to abiotic stress during cucumber seed germination.

[Advances in Agrobacterium tumefaciens-mediated transgenic cucumber].

Cucumber (Cucumis sativus) is an important vegetable crop in the world. Agrobacterium-mediated transgenic technology is an important way to study plant gene functions and improve varieties. In order to further accelerate the transgenic research and breeding process of cucumber, we described the progress and problems of Agrobacterium tumefaciens-mediated transgenic cucumber, from the influencing factors of cucumber regeneration ability, genetic transformation conditions and various additives in the process. We prospected for improving the genetic transformation efficiency and safety selection markers of cucumber, and hoped to provide reference for the research of cucumber resistance breeding and quality improvement.

Response of proteome and morphological structure to short-term drought and subsequent recovery in Cucumis sativus leaves.

Drought is the primary limitation to plant growth and yield in agricultural systems. Cucumber (Cucumis sativus) is one of the most important vegetables worldwide and has little tolerance for water deficit. To understand the drought stress response strategy of this plant, the responses of cucumber to short-term drought and rewatering were determined in this study by morphological structure and proteomic analyses. The leaf relative water content was significantly decreased under drought, and the cell structure was altered, while rewatering obviously alleviated the symptoms of water shortage and cell damage. A total of 320 and 246 proteins exhibiting significant abundance changes in response to drought and recovery, respectively, were identified. Our proteome analysis showed that 63 co-regulated proteins were shared between drought and rewatering, whereas most of the responsive proteins were unique. The proteome is adjusted through a sequence of regulatory processes including the biosynthesis of secondary metabolites and the glutathione metabolism pathway, which showed a high correlation between protein abundance profile and corresponding enzyme activity. Drought and recovery regulated different types of proteins, allowing plants to adapt to environmental stress or restore growth, respectively, which suggests that short-term drought and recovery are almost fully uncoupled processes. As an important component of the antioxidant system in plants, glutathione metabolism may be one of the main strategies for regulating antioxidant capacity during drought recovery. Our results provide useful information for further analyses of drought adaptability in cucumber plants.

Phloem sap proteome studied by iTRAQ provides integrated insight into salinity response mechanisms in cucumber plants.

Cucumber is an economically important crop as well as a model system for plant vascular biology. Salinity is one of the major environmental factors limiting plant growth. Here, we used an iTRAQ-based quantitative proteomics approach for comparative analysis of protein abundances in cucumber phloem sap in response to salt. A total of 745 distinct proteins were identified and 111 proteins were differentially expressed upon salinity in sensitive and tolerant cultivars, of which 69 and 65 proteins changed significantly in sensitive and tolerant cultivars, respectively. A bioinformatics analysis indicated that cucumber phloem employed a combination of induced metabolism, protein turnover, common stress response, energy and transport, signal transduction and regulation of transcription, and development proteins as protection mechanisms against salinity. The proteins that were mapped to the carbon fixation pathway decreased in abundance in sensitive cultivars and had no change in tolerant cultivars under salt stress, suggesting that this pathway may promote salt tolerance by stabilizing carbon fixation and maintaining the essential energy and carbohydrates in tolerant cultivars. This study leads to a better understanding of the salinity mechanism in cucumber phloem and provides a list of potential gene targets for the further engineering of salt tolerance in plants.

Effect of short-term water deficit stress on antioxidative systems in cucumber seedling roots.

BACKGROUND: Cucumber is one of the most popular vegetables, and have little tolerance for water stress. The antioxidant defense system is one of major drought defense and adaptive mechanisms in plants, however, relatively few data are available regarding antioxidant systems in responses of cucumber to water deficit. The effect of short-term drought stress on the antioxidant system, lipid peroxidation and water content in cucumber seedlings roots was investigated. RESULTS: The results showed that polyethylene glycol (PEG) induced water stress markedly decreased water content of cucumber seedling roots after treatment of 36 h, and caused excessive generation of reactive oxygen species (ROS) including superoxide (O2.-), hydrogen peroxide (H2O2). Meanwhile, malondialdehyde (MDA) content increased. Antioxidant enzymes including superoxide dismutases (SOD), peroxidases (POD), catalase (CAT) and ascorbate peroxidase (APX) activities increased in different time and different extent under water stress, while ascorbate (AsA) and glutathione (GSH) content, glutathione reductase (GR), dehydroascorbate reductase (DHAR) and monodehydroascorbate reductase (MDHAR) activities all decreased when compared to control. CONCLUSIONS: Therefore, it can be concluded that water stress strongly disrupted the normal metabolism of roots and restrained water absorption, and seemingly enzymatic system played more important roles in protecting cucumber seedling roots against oxidative damage than non-enzymatic system in short-term water deficit stress.

Proteomic analysis of cucumber seedling roots subjected to salt stress.

To understand metabolic modifications in plants under salt stress, the physiological and biochemical responses of cucumber (Cucumis sativus L. cv. Jinchun No. 2) seedlings to salt stress was investigated. The dry weight and fresh weight of cucumber seedling roots were significantly reduced by treatment with NaCl; Na(+) and Cl(-) were increased, while K(+) and K(+)/Na(+) ratio were decreased. To identify components of salt stress signaling, we compared the high resolution two-dimensional gel electrophoresis (2-DE) protein profiles of control and NaCl-treated roots, and the intensity of 34 protein spots varied. Of these spots, the identities of 29 (21 up-regulated and 8 down-regulated protein spots induced after salt stress) were determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and liquid chromatography electro-spray ionization tandem mass spectrometry (LC-ESI-MS/MS). The majority of the proteins had functions related to metabolism, energy and transport, and are involved in regulating reactions and defending against stress. A semi-quantitative reverse transcriptional-polymerase chain reaction (PCR) approach based on peptide sequences was used to compare transcript and protein accumulation patterns for 10 candidate proteins. Of these proteins, 8 patterns of induced transcript accumulation were consistent with those of induced protein accumulation. It is therefore likely that the response of the plant's proteome to NaCl stress is complex, and that the identified proteins may play an important role in regulating adaptation activities following exposure to NaCl stress in order to facilitate ion homeostasis.