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Kai-Shu Ling and Amnon Levi

One-hundred ninety U.S. PIs of bottlegourd [Lagenaria siceraria (Mol.) Standl.] were evaluated for their resistance to the Florida strain of Zucchini yellow mosaic virus (ZYMV-FL). Seedlings in the first leaf stage were mechanically inoculated with freshly prepared ZYMV-FL tissue extract in a greenhouse. Four weeks postinoculation, plants were visually evaluated for symptom expression and tissue samples from upper noninoculated leaves were collected for serological analysis with enzyme-linked immunosorbent analysis (ELISA). A combination of symptom expression and ELISA value was considered in determining the resistance or susceptibility for each accession. Of the 190 L. siceraria PIs screened, 36 accessions were in complete resistance (no disease symptom with negative ELISA on all tested plants), 64 PIs showed partial resistance (some of the tested plants were resistant, whereas others were susceptible), and 90 PIs were susceptible (severe symptom and positive ELISA on all tested plants). The ZYMV-FL resistance exists mostly among L. siceraria PIs collected in India. Thirty-three of the 36 L. siceraria PIs showing ZYMV-FL resistance were collected in India, one in Indonesia, one in South Africa, and one in Zimbabwe. To rule out any potential escapes in the primary screening, a repeated test using representative accessions, including 10 susceptible, three partially resistant, and three completely resistant PIs, was done to confirm the ZYMV-FL resistance. Furthermore, the resistance to ZYMV-FL was shown to be heritable in progenies generated through self-pollination of single plants in each of five resistant PIs as well as in three F1 hybrids.

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Amnon Levi, Karen R. Harris-Shultz, and Kai-shu Ling

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W. Patrick Wechter, Amnon Levi, Kai-Shu Ling, Chandrasekar Kousik, and Charles C. Block

Bacterial fruit blotch (BFB) caused by the bacterium Acidovorax avenae subsp. citrulli (Aac) is a seed-borne disease that threatens most cucurbit crops. Although limited resistance has been found in a small number of Plant introductions (PIs) in watermelon (Citrullus spp.), there are no reports of high levels of resistance in germplasm lines of Cucumis spp. In this study, 332 Cucumis spp. PIs were screened for resistance to Aac using a newly developed seed vacuum–infusion assay. Significant differences in the reaction of the PI to BFB were observed. The majority of lines were found to be extremely susceptible to the disease. However, several PIs with lower levels of resistance were also identified. Variability in the reaction of plants within each PI was also observed. Of the 332 PI tested, 16 were selected for additional evaluation using a standard spray inoculation tests. PI 353814, PI 381171, PI 536573, and PI 614401, all belonging to C. melo, and PI 504558 (C. ficifolius) were found to have significantly greater levels of resistance than susceptible control cultivars or other PIs in two independent spray inoculation tests. Germplasm lines developed from these PIs may be useful as sources of resistance to BFB in Cucumis breeding programs.

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Chandrasekar S. Kousik, Amnon Levi, Kai-Shu Ling, and W. Patrick Wechter

Powdery mildew (Podosphaera xanthii) can cause severe damage to cucurbit crops grown in open fields and greenhouses. In recent years, there has been an increased interest in the United States in grafting watermelon plants onto various cucurbit rootstocks. Bottle gourd plants (Lagenaria siceraria) are being used throughout the world as rootstocks for grafting watermelon. Although gourd plants are beneficial, they may still be susceptible hosts to various soilborne and foliar diseases. Bottle gourd plant introductions (PI) resistant to diseases and pests can be a valuable source of germplasm in rootstock breeding programs. We evaluated 234 U.S. PIs of L. siceraria for tolerance to powdery mildew in two greenhouse tests. Young seedlings were inoculated by dusting powdery mildew spores of melon race 1 on the cotyledons. Plants were rated 2 weeks after inoculation using a 1 to 9 scale of increasing disease severity. Although none of the L. siceraria PIs were immune to powdery mildew, several PIs had significantly lower levels of powdery mildew severity compared with susceptible watermelon cultivar Mickey Lee. The experiment was repeated with 26 select PIs on whole seedlings and cotyledon disks. Significant variability in the level of resistance to powdery mildew on plants within PI was observed. Moderate resistance in several PIs to powdery mildew was confirmed. PI 271353 had consistently lower ratings in the various tests and can be considered the most resistant to P. xanthii race 1 among the L. siceraria accessions evaluated in this study. A few other PIs with moderate resistance to powdery mildew included PI 271357, PI 381840, and PI 273663. These results suggest that novel sources of resistance could be developed by careful selection and screening of several of the PIs with moderate resistance described in our study.

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Amnon Levi, John Coffey, Laura Massey, Nihat Guner, Elad Oren, Yaakov Tadmor, and Kai-shu Ling

The bitter desert watermelon, Citrullus colocynthis (L.) Schrad is a wild species valuable for biotic and abiotic stress resistance that could be exploited for improving watermelon cultivars [Citrullus lanatus (Thunberg) Matsum & Nakai var. lanatus]. The objective of this study was to survey and identify C. colocynthis accessions displaying resistance to the Papaya ringspot virus-watermelon strain (PRSV-W). Thirty-one accessions of C. colocynthis, collected in Africa, the Middle East, southwest Asia, and India were evaluated for PRSV-W resistance. Of these 31 accessions, 4 U.S. Plant Introduction (PI) accessions, including 525080 (collected in Qena, Egypt) and PI 537277, PI 652554, and Griffin 14201 (collected at the northern Indian desert of Rajasthan and the neighboring region of Punjab, Pakistan) showed high resistance to PRSV-W. Plants of these four resistant PIs were self-pollinated to produce S1 and S2 seeds that continued to maintain the high levels of PRSV resistance. Since there is a wide genetic distance between watermelon cultivars and C. colocynthis, we performed crosses and backcrosses with watermelon cultivars, including ‘Charleston Gray’ and ‘Sugar Baby’ to produce viable seed that would be useful in the development of genetic populations and in introducing the resistance into watermelon cultivars.

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Haiying Zhang, Guoyi Gong, Shaogui Guo, Yi Ren, Yong Xu, and Kai-Shu Ling

Because of the growing threat of global warming, drought stress could severely affect the normal growth and development of crop plants. To alleviate such an adverse effect, there is a need to screen watermelon germplasm collections to identify genetic sources for potential drought tolerance. In the present study, 820 accessions of USDA's Citrullus PIs and 246 watermelon breeding lines were evaluated for their drought tolerance at the seedling stage under extreme water stress conditions in a greenhouse. Significant variations in drought tolerance were observed in the Citrullus germplasm collections. Using fast clustering analysis, the tested watermelon materials could be assigned into four groups, including tolerant, intermediate tolerant, moderately sensitive, and sensitive, respectively. The most drought-tolerant Citrullus germplasm, including 13 Citrullus lanatus var. lanatus and 12 C. lanatus var. citroides accessions, were originated from Africa. These genetic materials could be used for rootstock breeding or for developing drought-tolerant watermelon cultivars.

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Chandrasekar S. Kousik, Kai-Shu Ling, Scott Adkins, Craig G. Webster, and William W. Turechek

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Karen R. Harris, Kai-Shu Ling, W. Patrick Wechter, and Amnon Levi

Zucchini yellow mosaic virus (ZYMV) is one of the most economically important viruses affecting watermelon [Citrullus lanatus (Thunb.) Matsun & Nakai var. lanatus] in the United States. The ZYMV-Florida strain (ZYMV-FL) is considered a major limitation to commercial watermelon production in the United States. Inheritance of resistance to ZYMV-FL is conferred by a recessive gene. This report describes the identification of single-reaction, polymerase chain reaction-based markers linked to the ZYMV-FL resistance gene in watermelon. In this study, we identified a marker ZYMV-resistant polymorphism (ZYRP) linked to the ZYMV-FL resistance gene locus (genetic distance of 8 cM) in an F2 population, and in a backcross one to the resistant parent population (BC1R) (genetic distance of 13 cM). The identification of a single nucleotide polymorphism within the ZYRP marker for the parental genotypes allowed the development of a sequence-characterized amplification region marker linked to the ZYMV-FL resistance gene. Experiments using a BC2F2 population derived from the U.S. Plant Introduction 595203 (C. lanatus var. lanatus) and the recurrent parent ‘Charleston Gray’ indicated that the ZYRP marker can be used in marker-assisted selection to identify genotypes containing the gene conferring ZYMV-FL resistance in watermelon.

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Carol Gonsalves, Baodi Xue, Marcela Yepes, Marc Fuchs, Kaishu Ling, Shigetou Namba, Paula Chee, Jerry L. Slightom, and Dennis Gonsalves

A single regeneration procedure using cotyledon explants effectively regenerated five commercially grown muskmelon cultivars. This regeneration scheme was used to facilitate gene transfers using either Agrobacterium tumefaciens (using `Burpee Hybrid' and `Hales Best Jumbo') or microprojectile bombardment (using `Topmark') methods. In both cases, the transferred genes were from the T-DNA region of the binary vector plasmid pGA482GG/cp cucumber mosaic virus-white leaf strain (CMV-WL), which contains genes that encode neomycin phosphotransferase II (NPT II), β-glucuronidase (GUS), and the CMV-WL coat protein (CP). Explants treated with pGA482GG/cpCMV-WL regenerated shoots on Murashige and Skoog medium containing 4.4 μm 6-benzylaminopurine (BA), kanamycin (Km) at 150 mg·liter-1 and carbenicillin (Cb) at 500 mg·liter-1. Our comparison of A. tumefaciens- and microprojectile-mediated gene transfer procedures shows that both methods effectively produce nearly the same percentage of transgenic plants. R0 plants were first tested for GUS or NPT II expression, then the polymerase chain reaction (PCR) and other tests were used to verify the transfer of the NPT II, GUS, and CMV-WL CP genes. This analysis showed that plants transformed by A. tumefaciens contained all three genes, although co-transferring the genes into bombarded plants was not always successful. R1 plants were challenge inoculated with CMV-FNY, a destructive strain of CMV found in New York. Resistance levels varied according to the different transformed genotypes. Somaclonal variation was observed in a significant number of R0 transgenic plants. Flow cytometry analysis of leaf tissue revealed that a significant number of transgenic plants were tetraploid or mixoploid, whereas the commercial nontransformed cultivars were diploid. In a study of young, germinated cotyledons, however, a mixture of diploid, tetraploid, and octoploid cells were found at the shoot regeneration sites.