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Key message CRISPR/Cas9-mediated editing of Clpsk1 enhanced watermelon resistance to Fusarium oxysporum . The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system has proven to be an effective genome-editing tool for crop improvement. Previous studies described that Phytosulfokine (PSK) signalling attenuates plant immune response. In this work, we employed the CRISPR/Cas9 system to knockout Clpsk1 gene, encoding the PSK precursor, to confer enhanced watermelon resistance to Fusarium oxysporum f.sp. niveum ( FON ). Interactions between PSK and FON were analysed and it was found that transcript of Clpsk1 was significantly induced upon FON infection. Meanwhile, application of exogenous PSK increased the pathogen growth. Then, one sgRNA, which targeted the first exon of Clpsk1 , was selected for construction of pRGEB32-CAS9-gRNA-Clpsk1 expression cassette. The construct was then transformed to watermelon through Agrobacterium tumefaciens -mediated transformation method. Six mutant plants were obtained and three types of mutations at the expected position were identified based on Sanger sequencing. Resistance evaluation indicated that Clpsk1 loss-of-function rendered watermelon seedlings more resistant to infection by FON . These results indicate that CRISPR/Cas9-mediated gene modification is an effective approach for watermelon improvement.

Background Rhizosphere microorganisms and their interactions play a critical role in enhancing plant disease resistance. Here, we found that the disease severity of the resistant variety LW025 showed a decreasing trend with the increase in continuous cropping cycles. However, the mechanisms underlying the reduction in disease severity during the continuous cropping of the resistant watermelon variety LW025, particularly its relationship with the rhizosphere microbiome, remain unclear. Results In this study, the transcriptome of different watermelon varieties after continuous planting in pathogen-containing and pathogen-free soils was analyzed. The results showed that only two genes expression showed significant differences in disease-resistant variety between healthy and diseased soils. Subsequently, we analyzed the differences of rhizosphere soil microbial communities after planting different watermelon varieties for three consecutive seasons, as well as the relationship between differential microorganisms and soil physiochemical properties and soil enzyme activity. The results demonstrated continuous cropping of the disease-resistant variety LW025 formed a rhizosphere microbiome different from the initial soil and susceptible variety. Specifically, fungal changes were primarily observed in Ascomycota and Chytridiomycota, while bacterial changes were mainly observed in Cyanobacteria and Gemmatimonadetes. The bacterial functions enriched in the rhizosphere of the resistant variety LW025 after continuous cropping were primarily associated with soil nitrogen cycling. Furthermore, the plant disease index showed a significant positive correlation with the available phosphorus and potassium content in the soil, while exhibiting a significant negative correlation with soil pH and catalase activity. Conclusions Overall, the reduction in disease severity associated with continuous cropping of the disease-resistant variety LW025 was more closely related to changes in the rhizosphere microecological environment. This study explained the mechanism of the resistant variety LW025 against Fusarium infection, and provided new prospects for the development of technologies based on rhizosphere microecological environment modification to improve the resistance of watermelon to Fusarium wilt.

Root rot is an emerging disease of grafted watermelon in China that causes severe yield losses. The causal agents associated with this disease were characterized in this study. A total of 70 fungal isolates were recovered from infected roots, and the most prevalent isolates were identified as Fusarium oxysporum (31% of isolates recovered). F. oxysporum isolates induced typical root rot disease symptoms in pathogenicity tests, whereas the other isolates were nonpathogenic. On the basis of combined DNA sequence analyses, specific pathogenicity tests and root rot symptoms, the F. oxysporum was identified as F. oxysporum f. sp. radicis-lagenariae. We evaluated 37 bottle gourd rootstocks for resistance to F. oxysporum f. sp. radicis-lagenariae. The mean disease rating scores (DRSs) ranged from 1.1 to 4.0 at 20 days after inoculation. The rootstock 16S-71 was most resistant to infection. These findings provide useful information for the development of bottle gourd rootstocks with resistance to fusarium root rot and to manage this disease.

Grafting can improve the resistance of watermelon to soil-borne diseases. However, the molecular mechanism of defense response is not completely understood. Herein, we used a proteomic approach to investigate the molecular basis involved in grafted watermelon leaf defense against Fusarium oxysporum f.sp. niveum ( FON ) infection. The bottle gourd rootstock-grafted (RG) watermelon seedlings were highly resistant to FON compared with self-grafted (SG) watermelon plants, with a disease incidence of 3.4 and 89%, respectively. Meanwhile, grafting significantly induced the activity of pathogenesis-related proteases under FON challenge. Proteins extracted from leaves of RG and SG under FON inoculation were analyzed using two-dimensional gel electrophoresis. Thirty-nine differentially accumulated proteins (DAPs) were identified and classified into 10 functional groups. Accordingly, protein biosynthetic and stress- and defense-related proteins play crucial roles in the enhancement of disease resistance of RG watermelon seedlings, compared with that of SG watermelon seedlings. Proteins involved in signal transduction positively regulated the defense process. Carbohydrate and energy metabolism and photosystem contributed to energy production in RG watermelon seedlings under FON infection. The disease resistance of RG watermelon seedlings may also be related to the improved scavenging capacity of reactive oxygen species (ROS). The expression profile of 10 randomly selected proteins was measured using quantitative real-time PCR, among which, 7 was consistent with the results of the proteomic analysis. The functional implications of these proteins in regulating grafted watermelon response against F. oxysporum are discussed.

Auxin response factors (ARFs) are pivotal transcription factors involved in many aspects of auxin-dependent developmental processes. While functions of ARFs have been extensively studied in Arabidopsis, their distinct role in cucumber remains unclear. In this study, a cucumber auxin response factor homolog, CsARF10a, was cloned and overexpressed in tomato plants. RT-qPCR analysis indicated that the expression abundance of CsARF10a was significantly decreased in cucumber leaves and female flowers, and the expression level of CsARF10a was relatively low in pollinated fruits and hormone-treated fruits compared with that in unpollinated fruits. Moreover, the overexpression of CsARF10a in tomato resulted in multiple phenotypic changes, including a wider leaf blade, delayed fruit ripening, and parthenocarpic fruit set in CsARF10a-OE lines. Taken together, our research shed light on the regulatory importance of CsARF10a in regulating various phenotype alterations and laid a solid foundation for further functional studies.

The bottle gourd [Lagenaria siceraria (Molina) Standl.] is often utilized as a rootstock for watermelon grafting. This practice effectively mitigates the challenges associated with continuous cropping obstacles in watermelon cultivation. The lower ground temperature has a direct impact on the rootstocks’ root development and nutrient absorption, ultimately leading to slower growth and even the onset of yellowing. However, the mechanisms underlying the bottle gourd’s regulation of root growth in response to low root zone temperature (LRT) remain elusive. Understanding the dynamic response of bottle gourd roots to LRT stress is crucial for advancing research regarding its tolerance to low temperatures. In this study, we compared the physiological traits of bottle gourd roots under control and LRT treatments; root sample transcriptomic profiles were monitored after 0 h, 48 h and 72 h of LRT treatment. LRT stress increased the malondialdehyde (MDA) content, relative electrolyte permeability and reactive oxygen species (ROS) levels, especially H2O2 and O2−. Concurrently, LRT treatment enhanced the activities of antioxidant enzymes like superoxide dismutase (SOD) and peroxidase (POD). RNA-Seq analysis revealed the presence of 2507 and 1326 differentially expressed genes (DEGs) after 48 h and 72 h of LRT treatment, respectively. Notably, 174 and 271 transcription factors (TFs) were identified as DEGs compared to the 0 h control. We utilized quantitative real-time polymerase chain reaction (qRT-PCR) to confirm the expression patterns of DEGs belonging to the WRKY, NAC, bHLH, AP2/ERF and MYB families. Collectively, our study provides a robust foundation for the functional characterization of LRT-responsive TFs in bottle gourd roots. Furthermore, these insights may contribute to the enhancement in cold tolerance in bottle gourd-type rootstocks, thereby advancing molecular breeding efforts.

Watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai] is an economically important vegetable crop grown extensively worldwide. To facilitate the identification of agronomically important traits and provide new information for genetic and genomic research on this species, a high-density genetic linkage map of watermelon was constructed using an F₂ population derived from a cross between elite watermelon cultivar K3 and wild watermelon germplasm PI 189225. Based on a sliding window approach, a total of 1,161 bin markers representing 3,465 SNP markers were mapped onto 11 linkage groups corresponding to the chromosome pair number of watermelon. The total length of the genetic map is 1,099.2 cM, with an average distance between bins of 1.0 cM. The number of markers in each chromosome varies from 62 in chromosome 07 to 160 in chromosome 05. The length of individual chromosomes ranged between 61.8 cM for chromosome 07 and 140.2 cM for chromosome 05. A total of 616 SNP bin markers showed significant (P < 0.05) segregation distortion across all 11 chromosomes, and 513 (83.3 %) of these distorted loci showed distortion in favor of the elite watermelon cultivar K3 allele and 103 were skewed toward PI 189225. The number of SNPs and InDels per Mb varied considerably across the segregation distorted regions (SDRs) on each chromosome, and a mixture of dense and sparse SNPs and InDel SDRs coexisted on some chromosomes suggesting that SDRs were randomly distributed throughout the genome. Recombination rates varied greatly among each chromosome, from 2.0 to 4.2 centimorgans per megabase (cM/Mb). An inconsistency was found between the genetic and physical positions on the map for a segment on chromosome 11. The high-density genetic map described in the present study will facilitate fine mapping of quantitative trait loci, the identification of candidate genes, map-based cloning, as well as marker-assisted selection (MAS) in watermelon breeding programs.

Aims Fusarium oxysporum is a causal disease that threatens watermelon production, but little information on the molecular mechanisms involved in host defense is available. To understand the defense response, a proteome-level changes that occur in watermelon roots during F. oxysporum infection were investigated. Methods We utilized two-dimensional gel electrophoresis (2-DE) to compare changes in the root proteome profiles and validated their expression using real-time PCR. Results A total of 690 spots were detected, and 32 proteins had significant changes in abundance and were further identified by mass spectrometry. These proteins were mainly involved in metabolism, stress and defense and amino acid biosynthesis. RT-PCR analysis revealed that transcripts corresponding to the nine randomly selected proteins could be significantly induced, their expression patterns were consistent with the proteomic results except for Apx and Tdh. The involvement of these proteins in regulating watermelon response against F. oxysporum is discussed. Conclusions The reprogrammed proteins were involved in several biological processes, which indicates that watermelon can directly alter the abundance of these proteins to establish a defense response. This work helps us understand the basic processes during the watermelon-F. oxysporum interaction and may contribute to improve resistance breeding toward this pathogen.

Watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai var. lanatus] is an economically important vegetable belonging to the Cucurbitaceae family. Genotypes that exhibit agronomically important traits are selected for the development of elite cultivars. Understanding the genetic diversity and the genotype population structure based on molecular markers at the genome level can speed up the utilization of diverse genetic resources for varietal improvement. In the present study, we carried out an analysis of genetic diversity based on 3882 SNP markers across 37 core watermelon genotypes, including the most widely used watermelon varieties and wild watermelon. Based on the SNP genotyping data of the 37 watermelon genotypes screened, gene diversity and polymorphism information content values across chromosomes varied between 0.03–0.5 and 0.02–0.38, with averages of 0.14 and 0.13, respectively. The two wild watermelon genotypes were distinct from cultivated varieties and the remaining 35 cultivated genotypes were differentiated into three major clusters: 20 genotypes were grouped in cluster I; 11 genotypes were grouped in cluster II; three advanced breeding lines of yellow fruit flesh and genotype SW043 were grouped in cluster III. The results from neighbour-joining dendrogram, principal coordinate analysis and STRUCTURE analysis approaches were consistent, and the grouping of genotypes was generally in agreement with their origins. Here we reveal the genetic relationships among the core watermelon genotypes maintained at the Jiangsu Academy of Agricultural Sciences, China. The molecular and phenotypic characterization of the existing core watermelon genotypes, together with specific agronomic characteristics, can be utilized by researchers and breeders for future watermelon improvement.

Background Patients with insulo-Sylvian gliomas continue to present with severe morbidity in cognitive functions primarily due to neurosurgeons’ lack of familiarity with non-traditional brain networks. We sought to identify the frequency of invasion and proximity of gliomas to portions of these networks. Methods We retrospectively analyzed data from 45 patients undergoing glioma surgery centered in the insular lobe. Tumors were categorized based on their proximity and invasiveness of non-traditional cognitive networks and traditionally eloquent structures. Diffusion tensor imaging tractography was completed by creating a personalized brain atlas using Quicktome to determine eloquent and non-eloquent networks in each patient. Additionally, we prospectively collected neuropsychological data on 7 patients to compare tumor-network involvement with change in cognition. Lastly, 2 prospective patients had their surgical plan influenced by network mapping determined by Quicktome. Results Forty-four of 45 patients demonstrated tumor involvement (< 1 cm proximity or invasion) with components of non-traditional brain networks involved in cognition such as the salience network (SN, 60%) and the central executive network (CEN, 56%). Of the seven prospective patients, all had tumors involved with the SN, CEN (5/7, 71%), and language network (5/7, 71%). The mean scores of MMSE and MOCA before surgery were 18.71 ± 6.94 and 17.29 ± 6.26, respectively. The two cases who received preoperative planning with Quicktome had a postoperative performance that was anticipated. Conclusions Non-traditional brain networks involved in cognition are encountered during surgical resection of insulo-Sylvian gliomas. Quicktome can improve the understanding of the presence of these networks and allow for more informed surgical decisions based on patient functional goals.