Adaptation of cucurbit species to changes in substrate temperature: Root growth,antioxidants, and peroxidation

To investigate their response to changes in substrate temperatures, the roots from six species of cucurbit plants were exposed to 14°C, 24°C, or 34°C, while their aerial portions were maintained at natural ambient temperatures (23°C to 33°C). These species could be classified into three groups based on their stress response: Group A,Cucurbita ficifolia and C.maxima, heatsensitive but cold-tolerant; Group B,Cucumis sativus and C.melo, heat- and cold-sensitive; and Group C,Luffa cylindrica andBenincasa hispida, heat-tolerant but cold-sensitive. The highest growth rates and lowest contents of malondialdehyde (MDA) for plants in Groups A, B, and C were achieved at temperatures of 14°C, 24°C, and 24°C to 34°C, respectively. Superoxide dismutase (SOD) activity was lowest in the roots exposed to optimal growth temperatures while activities of catalase (CAT), ascorbate peroxidase (APX), and guaiacol peroxidase (G-POD) operated coordinately in a complicated manner to prevent the accumulation of reactive oxygen species (ROS) in the root cells. Moreover, all plants, regardless of species, responded to unfavorable temperatures by increasing their synthesis of ascorbate and glutathione as well as by reducing the redox ratio of those two important antioxidants.     

Resurgence of cucurbit downy mildew in the United States: Insights from comparative genomic analysis of Pseudoperonospora cubensis

Pseudoperonospora cubensis, the causal agent of cucurbit downy mildew (CDM), is known to exhibit host specialization. The virulence of different isolates of the pathogen can be classified into pathotypes based on their compatibility with a differential set composed of specific cucurbit host types. However, the genetic basis of host specialization within P. cubensis is not yet known. Total genomic DNA extracted from nine isolates of P. cubensis collected from 2008 to 2013 from diverse cucurbit host types (Cucumis sativus, C. melo var. reticulatus, Cucurbita maxima, C. moschata, C. pepo, and Citrullus lanatus) in the United States were subjected to whole‐genome sequencing. Comparative analysis of these nine genomes confirmed the presence of two distinct evolutionary lineages (lineages I and II) of P. cubensis. Many fixed polymorphisms separated lineage I comprising isolates from Cucurbita pepo, C. moschata, and Citrullus lanatus from lineage II comprising isolates from Cucumis spp. and Cucurbita maxima. Phenotypic characterization showed that lineage II isolates were of the A1 mating type and belonged to pathotypes 1 and 3 that were not known to be present in the United States prior to the resurgence of CDM in 2004. The association of lineage II isolates with the new pathotypes and a lack of genetic diversity among these isolates suggest that lineage II of P. cubensis is associated with the resurgence of CDM on cucumber in the United States.     

Effects of aqueous root extracts and hydrophobic root exudates of cucumber (Cucumis sativus L.) on nuclei DNA content and expression of cell cycle-related genes in cucumber radicles

Allelopathic interactions implicate the inhibition of cell division by allelochemicals. To examine the effects of autotoxic agents on cell cycle and plant growth, germinated cucumber seeds (Cucumis sativus L.) were incubated in solutions containing the aqueous root extracts of cucumber at 1:100, 1:50, 1:25 and 1:10 (w:v), or the hydrophobic root exudates of cucumber at 25, 50 and 100 mg·L^?1. Aqueous root extracts and hydrophobic root exudates inhibited radicle elongation by 36.47–60.18% and 38.24–62.50%, respectively. The mitosis-specific genes were down-regulated in roots exposed to aqueous root extracts and hydrophobic root exudates. Meanwhile, exposure to either aqueous root extracts or hydrophobic root exudates decreased the proportion of 2C (C-value) and increased the proportion of 8C, leading to an increased mean C-value. We conclude that autotoxic agent-induced inhibition of radicle growth was partly attributed to the down-regulation of cell cycle-related genes and endoreduplication was enhanced under our experimental condition.     

Distribution,Host Range and Disease Severity of Pseudoperonospora cubensis on Cucurbits in the Czech Republic

Cucurbit downy mildew, caused by Pseudoperonospora cubensis, is a major cucumber disease in the Czech Republic. Disease prevalence, host range and disease severity were evaluated from 2001 to 2009. The geographical distribution of P. cubensis was assessed on ca 80–100 locations per year in two main regions of the Czech Republic (central and southern Moravia, and eastern, northern and central Bohemia). Infection by P. cubensis was observed primarily on cucumber (Cucumis sativus) but only on the leaves. During the study, disease prevalence ranged from 66 to 100%. The majority of C. sativus crops were heavily infected at the end of the growing season (second half of August). Generally, P. cubensis was present at high or very high disease severity. The loss of foliage results in the reduction in the quality and quantity of marketable yield of fruit. Pseudoperonospora cubensis was widespread across the whole area of the Czech Republic studied. Very rarely, infection was recorded in muskmelon (Cucumis melo) and Cucurbita moschata. Of other pathogens, the most frequently recorded was the cucurbit powdery mildew (Golovinomyces cichoracearum and Podosphaera xanthii).     

Detection and Location of Seed-Borne Inoculum of Didymella bryoniae and its Transmission in Seedlings of Cucumber and Pumpkin

Of the 91 tested cucurbit seed samples from thirteen countries nine from four countries were found to be infected with Didymella bryoniae. The pathogen is located on and in the seed coat including the perisperm and in the tissue of the cotyledons. Primary seedling infection occurred on the radicle, hypoctyl and cotyledons. Infection of the radicle generally caused pre-emergence tot while infection on the hypocotyl and cotyledons developed furthere inoculum for infection of the first true leaves and the stem. Experimentally, all the isolates of D. bryoniae could infect cucumber (Cucumis sativus), oriental melon (Cucumis melo var. makuwa), pumpkin (Cucurbita pepo) and watermelon (Citrullus vulgaris) at different growth stages; the susceptibility of cucumber and pumpkin was markedly influenced by previailing humid conditions.The blotter method was found more suitable for detection of seed-borne infection than the agar plate method.     

Cross pathogenicity studies with isolates of Fusarium oxysporum from either cucumber or watermelon pathogenic to both crop species*

Isolates of Fusarium oxysporum from Abaco, the Bahamas, whether obtained from wilted plants of cucumber (Cucumis sativus) or watermelon (Citrullus lanatus), were pathogenic to cucumber, watermelon, and cantaloupe (Cucumis melo var. reticulatus). The West Indian gherkin (Cucumis anguria), pumpkin (Cucurbita pepo), and three cultivars of summer squash (C. pepo var. melopepo) were not susceptible. Strains of F. oxysporum from wilted cucumber or watermelon plants from Florida were highly pathogenic only to their original host species and are regarded as different formae speciales.     

A tobacco ringspot virus-based vector system for gene and microRNA function studies in cucurbits

Cucurbits are economically important crops worldwide. The genomic data of many cucurbits are now available. However, functional analyses of cucurbit genes and noncoding RNAs have been impeded because genetic transformation is difficult for many cucurbitaceous plants. Here, we developed a set of tobacco ringspot virus (TRSV)-based vectors for gene and microRNA (miRNA) function studies in cucurbits. A TRSV-based expression vector could simultaneously express GREEN FLUORESCENT PROTEIN (GFP) and heterologous viral suppressors of RNA silencing in TRSV-infected plants, while a TRSV-based gene silencing vector could knock down endogenous genes exemplified by PHYTOENE DESATURASE (PDS) in Cucumis melo, Citrullus lanatus, Cucumis sativus, and Nicotiana benthamiana plants. We also developed a TRSV-based miRNA silencing vector to dissect the functions of endogenous miRNAs. Four representative miRNAs, namely, miR159, miR166, miR172, and miR319, from different cucurbits were inserted into the TRSV vector using a short tandem target mimic strategy and induced characteristic phenotypes in TRSV-miRNA-infected plants. This TRSV-based vector system will facilitate functional genomic studies in cucurbits.     

Rapid Detection of Colletotrichum lagenarium,Causal Agent of Anthracnose of Cucurbitaceous Crops,by PCR and Real‐time PCR

Rapid and accurate polymerase chain reaction (PCR) and real‐time PCR methods were developed for the detection of Colletotrichum lagenarium, the causal agent of anthracnose, in tissues of squash (Cucurbita moschata), watermelon (Citrullus lanatus), cucumber (Cucumis sativus) and muskmelon (Cucumis melo). PCR assays amplified different internal transcribed spacer sequences from C. lagenarium, so effectively detected this pathogen in infected tissues. PCR analysis with the primer co‐m‐337F1/R1 was able to differentiate C. lagenarium from other fungal pathogens, including Colletotrichum spp., Fusarium spp., Alternaria spp. and Didymella spp. An optimized real‐time PCR assay was developed to detect and monitor C. lagenarium in both infected plant tissues and soil samples. The sensitivity of real‐time PCR can detect down to 1 pg of DNA. Thus, PCR‐based analysis is a useful technique for rapid detection and diagnosis of C. lagenarium in infected plants or infested soils.     

Overview of the origins and history of the five major cucurbit crops: issues for ancient DNA analysis of archaeological specimens

The gourd family, Cucurbitaceae, contains five vegetable crops of worldwide importance, the pumpkins and squash (Cucurbita spp.), watermelons (Citrullus lanatus), melons (Cucumis melo) and cucumbers (Cucumis sativus). Here is presented a synopsis of the origin and history of these cucurbit crops. Historical records of the use of cucurbits by people take the form of archaeobotanical finds, iconography and literature. The weight of the evidence indicates that Cucurbita spp. were first cultivated in the Americas at least 10,000 years ago, and that by 1492 ce a number of cultivar-groups of pumpkins and squash had been developed by indigenous American peoples. Watermelons were cultivated in northeastern Africa at least 4,000 years ago, first probably as a source of fresh water. Melons and cucumbers are native to Asia, probably initially cultivated for the use of the young fruits as vegetables. Melons spread to eastern Africa at an early date, but cucumbers are probably a more recent domesticate and spread westwards later, reaching Europe in early medieval times. Sequencing of cucurbit plant genomes and advances in ancient DNA research offer much promise for obtaining an improved assessment of cucurbit crop origins, specifically the genetic constitution and geographical home of ancestral source populations. Next-generation genomic sequencing, if applied to an appropriate array of archaeological cucurbit remains and modern germplasm, could contribute much to the understanding of the history and evolution under domestication of cucurbit crops.     

Phylogenetic analysis of <Emphasis Type="Italic">Plectosphaerella</Emphasis> species based on multi-locus DNA sequences and description of <Emphasis Type="Italic">P. sinensis</Emphasis> sp. nov.

Species in Plectosphaerella are well known as pathogens of several plant species causing fruit, root and collar rot and collapse. In an investigation of endophytic fungi associated with cucurbit plants in China, we isolated 77 strains belonging to the genus Plectosphaerella. To identify the isolated strains, we collected the type or reference strains of all currently accepted species in Plectosphaerella except P. oratosquillae and conducted a phylogenetic analysis. Phylogenetic analysis of the partial 28S rDNA sequences showed that all species in Plectosphaerella were located in one clade of Plectosphaerellaceae. Based on multi-locus phylogenetic analysis of the ITS, CaM, EF1, TUB and morphological characteristics, all species in Plectosphaerella were well separated. Three endophytic strains from stems of Cucurbita moschata, Citrullus lanatus and Cucumis melo from North China were assigned to a new species described as P. sinensis in this paper. The new species differs morphologically from other Plectosphaerella species by irregular chlamydospores, and the dimensions of phialides and conidia. The other endophytic strains from several cucurbit plants were identified as P. cucumerina.     

Relative incidence,spatial distribution and genetic diversity of cucurbit viruses in eastern Spain

Viral diseases that could cause important economic losses often affect cucurbits, but only limited information on the incidence and spatial distribution of specific viruses is currently available. During the 2005 and 2006 growing seasons, systematic surveys were carried out in open field melon (Cucumis melo), squash and pumpkin (Cucurbita pepo), watermelon (Citrullus lanatus) and cucumber (Cucumis sativus) crops of the Spanish Community of Valencia (eastern Spain), where several counties have a long standing tradition of cucurbit cultivation and production. Surveyed fields were chosen with no previous information as to their sanitation status, and samples were taken from plants that showed virus‐like symptoms. Samples were analysed using molecular hybridisation to detect Beet pseudo‐yellows virus (BPYV), Cucurbit aphid‐borne yellows virus (CABYV), Cucumber mosaic virus (CMV), Cucumber vein yellowing virus (CVYV), Cucurbit yellow stunting disorder virus (CYSDV), Melon necrotic spot virus (MNSV), Papaya ring spot virus (PRSV), Watermelon mosaic virus (WMV) and Zucchini yellow mosaic virus (ZYMV). We collected 1767 samples from 122 independent field plots; out of these, approximately 94% of the samples were infected by at least one of these viruses. Percentages for the more frequently detected viruses were 35.8%, 27.0%, 16.5% and 7.2% for CABYV, WMV, PRSV and ZYMV, respectively, and significant deviations were found on the frequency distributions based on either the area or the host sampled. The number of multiple infections was high (average 36%), particularly for squash (more than 57%), with the most frequent combination being WMV + PRSV (12%) followed by WMV + CABYV (10%). Sequencing of WMV complementary DNA suggested that ‘emerging’ isolates have replaced the ‘classic’ ones, as described in southern regions of France, leading us to believe that cucurbit cultivation could be severely affected by these new, emerging isolates.     

Transferability of Cucurbita SSR markers for genetic diversity assessment of Turkish bottle gourd (Lagenaria siceraria) genetic resources

The genetic diversity present in crop landraces represents a valuable genetic resource for breeding and genetic studies. Bottle gourd (Lagenaria siceraria) landraces in Turkey are highly genetically diverse. However, the limited genomic resources available for this crop hinder the molecular characterization of Turkish bottle gourd germplasm for its adequate conservation and management. Therefore, we evaluated the efficacy of 40 SSR markers from major cucurbit crops (Cucurbita pepo L. and Cucurbita moschata L.) in 30 bottle gourd landraces, together with 16 SRAP primer combinations. In addition, we compared the genetic relationship between bottle gourd and 31 other cucurbit accessions (11 Cucurbita maxima, 3 C. moschata, 5 C. pepo subsp. ovifera, 10 C. pepo and 2 Luffa cylindrica). Twenty-seven Cucurbita SSR markers showed transferability to bottle gourd. SSR markers amplified 59 alleles, in bottle gourd genome with an average of 1.64 alleles per locus. Together, SSR and SRAP markers amplified 453 fragments across the 61 accessions, and clearly discriminated L. siceraria and L. cylindrica from the other cucurbit species. Genetic diversity analysis separated edible cucurbit from ornamentals, while population structure analysis classified L. siceraria in two subpopulations defined by fruit shape, rather than geographical origin. The results indicated that the genomic resources available for Cucurbita species are valuable to study and preserve the genetic diversity of bottle gourd in Turkey.     

Cucurbit Grafting

Due to limited availability of arable land and high market demand for off-season vegetables, cucurbits (plants in the family Cucurbitaceae) are continuously cultivated under unfavorable conditions in some countries. These conditions include environments that are too cold, wet, or dry, or are cool low-light winter greenhouses. Successive cropping can increase salinity, the incidence of cucurbit pests, and soilborne diseases like fusarium wilt caused by Fusarium spp. These conditions cause various physiological and pathological disorders leading to severe crop loss. Chemical pest control is expensive, not always effective, and can harm the environment. Grafting can overcome many of these problems. In fact, in many parts of the world, grafting is a routine technique in continuous cropping systems. It was first commonly used in Japan during the late 1920s by grafting watermelon [Citrullus lanatus (Thunb.) Matsum. and Nakai] onto pumpkin [Cucurbita moschata Duchesne ex. Poir] rootstocks. Soon after, watermelons were grafted onto bottle gourd [Lagenaria siceraria (Molina) Standl.] rootstocks. This practice helped control declining yield due to soilborne diseases. China produces more than half the world's watermelons and cucumbers (Cucumis sativus L.), and approximately 20% of these are grafted. Use of rootstocks can enhance plant vigor through vigorous attainment of soil nutrients, avoidance of soil pathogens and tolerance of low soil temperatures, salinity, and wet-soil conditions. The type of rootstock affects cucurbit plant growth, yield, and fruit quality. Cucurbit grafting is rare in the United States, but with continued loss of quality disease-free farmland along with the phase-out of methyl bromide, the U.S. cucurbit industry sees grafting as an attractive option. Some seed companies now offer watermelon transplants grafted onto squash or bottle gourd rootstocks, and some transplant facilities offer grafting services. There have been thorough analyses of cucurbit grafting in other countries, but the literature in English is limited. This review summarizes the state of the cucurbit grafting industry on a global level, translating work published in many languages.     

Natural Infection by Tospoviruses of Cucurbitaceous and Fabaceous Vegetable Crops in India

Natural infection of tospoviruses on three cucurbitaceous (Cucumis sativus, cucumber; Luffa acutangula, ridge gourd; Citrullus lanatus, watermelon) and three fabaceous (Vigna unguiculata, cowpea; Phaseolus vulgaris, French bean; Dolichos lablab, sem) vegetable crops in India was identified on the basis of nucleocapsid protein (NP) gene characteristics. The complete NP gene of the cowpea isolate from Kerala and the sem isolate from Tamil Nadu was 831 nucleotides long, encoding a protein of 276 amino acids. For other Tospovirus isolates from cucumber, French bean, ridge gourd and watermelon, the partial NP gene (291 nt) was sequenced. Comparative NP gene sequence analyses revealed that fabaceous isolates shared maximum identity both at the nucleotide (92–97%) and amino acid (93–97%) levels with the corresponding region of Groundnut bud necrosis virus (GBNV), whereas cucurbitaceous isolates shared maximum identity both at nucleotide (93–99%) and amino acid (95–98%) levels with the corresponding region of Watermelon bud necrosis virus (WBNV), results suggesting that the Tospovirus isolates infecting fabaceous hosts should be regarded as strain of GBNV, whereas those infecting cucurbitaceous hosts as a strain of WBNV. Nucleocapsid protein gene was conserved both in GBNV and WBNV isolates originating from different hosts and locations.     

Evidence for graft transmission of structural phloem proteins or their precursors in heterografts of Cucurbitaceae

Interspecific and intergeneric grafts of Cucurbitaceae were used to study the mobility of structural P-proteins in the phloem. When Cucumis sativus L. scions were grafted onto Cucurbita rootstocks, at least nine additional proteins appeared on sodium dodecyl sulfate-polyacrylamide electrophoresis gels of scion exudate, 9–11 d after grafting. These proteins corresponded exactly to those of the respective Cucurbita sp. rootstock, including the filament-forming phloem protein PP1 and the phloem lectin PP2, as shown by the apparent molecular weights and peptide maps. According to probing at three sites, the additional proteins were evenly distributed within the scion. The appearance of additional proteins was correlated with the establishment of phloem bridges across the graft union. The developmental coincidence establishes that the structural proteins or their precursors are translocated in the phloem. This translocation was a universal phenomenon in Cucurbitaceae as shown by a comparative screening for additional proteins in eleven graft combinations, using Benincasa hispida (Thunb.) Cogn., Citrullus colocynthis (L.) Schrad., Cucumis melo L, C. sativus, Cucurbita ficifolia Bouché, Cucurbita maxima Duchesne ex Lam., and Trichosanthes cucumerina var. lobata Roxb. According to this screening, the direction of transmission of additional proteins depended upon the combination tested. While some graft partners failed to show exchange, some behaved as “donor” for additional proteins and still others could be both “donor” or “acceptor”. However, whether used as scion or stock, C. sativus was consistently identified as an acceptor. The occurrence of additional proteins in heterografts is discussed with regard to the transport mechanism of structural P-proteins in the phloem and its relationship to assimilate transport. Received: 18 February 1998 / Accepted: 12 May 1998     

Response of Cucumber Cultivars and Other Cucurbitaceous Species to Infection by Hop Stunt Viroid1

The response of 8 different species within the Cucurbitaceae to hop stunt viroid (HSV) was examined. Some of these plants developed several kinds of systemic symptoms depending upon species, variety or cultivar infected. Among 12 out of 26 cucumber cultivars become infected severely, so visual symptoms are reliable for diagnosis because symptom severity is correlated with viroid content. Other cucurbitaceous species except Citrullus lanatus, Cucurbita maxima and Cucurbita moschata were also susceptible. Among them, Benincasa hispida was found to be a more suitable bioassay plant than Suuyou cucumber which has been hitherto used.     

A comparative analysis of phloem exudate proteins from Cucumis melo,Cucumis sativus and Cucurbita maxima by polyacrylamide gel electrophoresis and isoelectric focusing

Summary Proteins in sieve tube exudate from Cucumis melo L., Cucumis sativus L. and Cucurbita maxima Duch. were analysed by gel electrophoresis and isoelectric focusing. Estimated molecular weights and isoelectric points for the major and minor proteins from each plant species are presented. Electrophoresis revealed striking differences between the protein complements of exudatc from the two genera investigated. Similarly, although a few exudate proteins from the two species of Cucumis possessed identical molecular weights, several major proteins were peculiar to each species. Isoelectric focusing of proteins in exudate samples from the three plants confirmed the marked differences in their protein complements. Furthermore, focusing also revealed differences between cultivars of Cucumis sativus. Both Cucumis sativus and Cucurbita maxima possessed relatively large amounts of basic proteins; these were absent in exudate from Cucumis melo. The implications of these results are discussed in relation to present concepts regarding the interrelationships and possible functional roles of P-proteins.Abbreviations DTT dithiothreitol - Bis N,N-methylenebisacrylamide - SDS Sodium dodecyl sulphate - TCA trichloroacetic acid     

Taxonomic significance of trichomes micromorphology in cucurbits

Studies on trichomes micromorphology using Scanning Electron Microscope (SEM) were undertaken in 23 species with one variety under 13 genera of the family Cucurbitaceae (viz., Benincasa hispida (Thunb.) Cogn., Citrullus lanatus (Thunb.) Matsum. & Nakai, Cucumis melo var. agrestis Naudin, Cucumis sativus L., Diplocyclos palmatus (L.) C. Jeffrey, Edgaria dargeelingensis C.B. Clarke, Gynostemma burmanicum King ex Chakr., Gynostemma pentaphyllum (Thunb.) Makino, Gynostemma pubescens (Gagnep.) C.Y. Wu, Hemsleya dipterygia Kuang & A.M. Lu, Lagenaria siceraria (Molina) Standl., Luffa acutangula (L.) Roxb., Luffa cylindrica M. Roem., Luffa echinata Roxb., Melothria heterophylla (Lour.) Cogn., Melothria leucocarpa (Blume) Cogn., Melothria maderspatana (L.) Cogn., Sechium edule (Jacq.) Sw., Thladiantha cordifolia (Blume) Cogn., Trichosanthes cucumerina L., T. cucumerina var. anguina (L.) Haines, Trichosanthes dioica Roxb., Trichosanthes lepiniana (Naudin) Cogn. and T. tricuspidata Lour.). The trichomes in the family Cucurbitaceae vary from unicellular to multicellular, conical to elongated, smooth to ridges, with or without flattened disk at base and cyctolithic appendages, thin to thick walled, curved at apices to blunt. Trichomes micromorphology in the family Cucurbitaceae was found significant taxonomically.