Nihat Guner

Inheritance of Resistance to Papaya Ringspot Virus-Watermelon Strain in Watermelon

Sources of resistance to the watermelon strain of papaya ringspot viruswatermelon strain (PRSV-W) have been identified within the watermelon (Citrullus lanatus) germplasm collection. Inheritance of the resistance to PRSV-W was studied in three Citrullus amarus (formerly C. lanatus var. citroides) PI accessions: PI 244017, PI 244019, and PI 485583. Three susceptible parent lines, ‘Allsweet’, ‘Calhoun Gray’, and ‘NewHampshireMidget’, were crossed with resistant PI accessions to develop F1, F2, and BC1 progenies in six families. A single recessive gene was found to control the resistance to PRSV-W in all three resistant PI accessions. Allelism tests indicated that the three PI accessions carry the same resistance allele to PRSV-W. The gene symbol ‘prv’ is proposed for PRSV-W resistance in PI 244017, PI 244019, and PI 485583 in watermelon.

Inheritance of Resistance to Zucchini Yellow Mosaic Virus in Watermelon

Sources of resistance to the Zucchini yellowmosaic virus-Florida strain (ZYMVFL) have been identified within the Citrullus genus. Inheritance of resistance to ZYMVFL was studied in PI 595203 (Citrullus mucosospermus), a resistant watermelon accession. The F1, F2, and BC1 progenies derived from the cross ‘Calhoun Gray’ (CHG) 3 PI 595203 and ‘New Hampshire Midget’ (NHM) 3 PI 595203 were used to study the inheritance of resistance to ZYMV-FL. Seedlings were inoculated with a severe isolate of ZYMV-FL at the first true leaf stage and rated weekly for at least 6 weeks on a scale of 1 to 9 on the basis of severity of viral symptoms. A single recessive gene (zymFL) was found to control the high level of resistance to ZYMV-FL in PI 595203. Watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai] is a major cucurbit crop that accounts for 7.5% of the world area devoted to vegetable crop production (FAO, 2015). In the United States, watermelon is used fresh as a dessert, or in salads. Major production states in the United States. are Florida, California, Texas, Georgia, and North Carolina (USDA, 2017). Production increased from 1.2 million Mg (metric tons) in 1987 to 5.1 million Mg in 2017, with a farm value of

The Genes of Watermelon

579 million (USDA, 2017). Plant diseases incited by viruses are a major limiting factor in commercial watermelon production worldwide. More than 10 viruses are known to be a problem in watermelon field production (Ali et al., 2012; Wang et al., 2017). The major viruses affecting watermelon in the United States are Zucchini yellow mosaic virus (ZYMV), Papaya ringspot virus-watermelon strain (PRSV-W), and Watermelon mosaic virus (WMV). All three viruses are nonpersistently transmitted by several species of aphids, and mixed infections are common (Ali et al., 2012; Chen et al., 2008; Guner et al., 2018; Morata and Puigdom enech, 2017; Wehner et al., 2001). Chemical control of the vectors is not usually an efficient method of controlling the disease. Cultural controls such as treatment with mineral oil sprays and light-reflective surfaces, and cross-protection with weak ZYMV isolates showed limited effectiveness and required additional input costs. Therefore, genetic resistance remains the simplest, most effective, and most efficient means of limiting losses to these diseases (Ali et al., 2012; Levi et al., 2016a; Nagendran et al., 2017; Yu et al., 2011). ZYMV was first described in 1981 in squash grown in northern Italy and France, where it was named Muskmelon yellow stunt virus (Lisa and Dellvalle, 1981; Tiwari and Rao, 2014). ZYMV infects all the agriculturally important species of Cucurbitaceae family (e.g., C. lanatus, Cucumis sativus, Cucumis melo, and Cucurbita spp.) and is considered the most destructive virus in watermelon production worldwide (Nagendran et al., 2017; Provvidenti, 1991). ZYMV is transmitted in a nonpersistent manner by several aphid species (e.g., Aphis gossypii Glover) and easily transmitted mechanically. In areas where cucurbit crops are not grown continuously, the virus overwinters on wild species. Natural infection appears to be limited to species of the Cucurbitaceae, but members of 11 families of dicotyledons are considered diagnostic hosts (Tiwari and Rao, 2014). ZYMV, with flexuous particles 750 nm long containing a single strand of RNA, belongs to the genus Potyvirus in the family Potyviridae (Romay et al., 2014; Tiwari and Rao, 2014). At least 25 strains of ZYMV have been identified (Desbiez and Lecoq, 1997). Provvidenti et al. (1984) reported the occurrence of Connecticut (CT) and Florida (FL) strains of ZYMV, with the FL strain occurring more widely in the United States. In the 1990s, Provvidenti also identified a new ZYMV strain infecting cucurbit fields around Beijing, China, Zucchini yellow mosaic virus-China strain (ZYMV-CH). Plants infected with any of the ZYMV strains reduce their photosynthetic capacity and display stunted growth, deformed fruit, and early mortality (Guner and Wehner, 2008). Symptoms of severe ZYMV infection in cucurbit crops include yellowmosaic, stunting, blistering, and laminar reduction on leaves, and fruit remaining small, developing knobby areas, greatly malformed, and mottled (Nagendran et al., 2017). Researchers have screened the germplasm resources of several cucurbit species for resistance to ZYMV, and the inheritance of the resistance has been reported in cucumber (C. sativus), melon (C. melo), and squash (Cucurbita spp.). ZYMV resistance is controlled by a single recessive gene in cucumber (Cardoso et al., 2010; Kabelka and Grumet, 1997), a single dominant gene in melon (Park et al., 2004; Pitrat and Lecoq, 1984), and a single dominant gene in squash (Paris and Brown, 2005). The watermelon germplasm collection has been screened for resistance to ZYMV, and several resistant PI accessions have been identified (Boyhan et al., 1992; Guner, 2004; Provvidenti, 1991). Provvidenti (1991) reported ZYMV resistance in four PI accessions of watermelon from Zimbabwe (PI 482322, PI 482299, PI 482261, and PI 482308). The ZYMV resistance in the four resistant watermelon PI accessions appeared to be specific to the Florida strain (ZYMVFL). Some accessions of egusi watermelon (C. mucosospermus) originating in Nigeria (PI 494528 and PI 494532) were reported resistant to ZYMV, and the resistance was not specific to the virus strain. However, the resistance was temperature dependent and was expressed best in warm or hot climates (Provvidenti, 1986). Provvidenti (1991) reported that resistance in PI 482261 (Citrullus amarus) was conferred by a single recessive gene, which he named zym. Boyhan et al. (1992) reported additional sources of resistance to ZYMV in PI 386026, PI 386025 (both Citrullus colocynthis), and the egusi PI 595203 (C. mucosospermus). They also confirmed ZYMV resistance in PI 482261 and PI 494528. Guner (2004) screened 1613 PI accessions along with 41 watermelon cultivars for resistance to ZYMV. A high level of resistance to ZYMV was found in several PI accessions. These new resistant PI accessions were PI 386019, PI 490377, PI 596662, PI 485580, PI 560016, PI 494528, PI 386016, PI 482276, and PI 595201. The study also confirmed the resistance of PI 595203 (egusi), PI 386025, PI 386026, and PI 494528. However, the resistance of PI 482322, PI 482299, PI 482261, and PI 482308 as reported by Provvidenti (1991) was not confirmed. Those PI accessions had only moderate resistance to the Florida strain (ZYMV-FL) used in the study of Guner (2004). Although it was reported that the inheritance of the resistance to ZYMV-CH in PI 595203 is controlled by a single recessive gene (Xu et al., 2004), there is not a formal report on the inheritance of the resistance to ZYMV-FL in PI 595203. Whether the ZYMV-FL and ZYMV-CH strains are the same has not been determined (Harris et al., 2009; Levi and Ling, 2017; Levi et al., 2016b; Ling et al., 2009). Therefore, the Received for publication 12 Mar. 2018. Accepted for publication 26 Apr. 2018. Corresponding author. E-mail: lariver2@ncsu. edu. HORTSCIENCE VOL. 53(8) AUGUST 2018 1115 objective of this study was to determine the inheritance of ZYMV-FL resistance in PI 595203 (C. mucosospermus), an egusi watermelon type that was also identified as the source of resistance to ZYMV-CH strain, a geographically distant isolate. Materials and Methods Plant material. The parental lines used in this study were CHG and NHM, highly susceptible to ZYMV, and an egusi accession (PI 595203), our best source of resistance. Two families were developed by crossing PI 595203 with CHG and NHM. All crosses were made using hand-pollination in the greenhouse. Six generations (susceptible parent, resistant parent, F1, F2, backcross to the susceptible parent, and backcross to the resistant parent) were developed for the study of inheritance of resistance. For each of the two crosses, 180 plants were tested: 5 Ps, 5 Pr, 30 BC1Ps, 30 BC1Pr, 10 F1, and 100 F2 plants were used to test the inheritance from each cross. Planting and management. All experiments were performed in the greenhouse of the Department of Plant Pathology at North Carolina State University in Raleigh, NC. Greenhouse temperatures ranged from 23 to 43 C (day) and from 12 to 24 C (night). We seeded directly in plastic pots (100 · 100 mm size, 600-mL volume) filled with a soilless mix (Canadian sphagnum moss, perlite, vermiculite, and processed pine bark). We planted two seeds per pot and thinned to one to ensure a uniform experiment (Gusmini et al., 2017). Inoculum preparation. The virus isolate was obtained from E. Hiebert, University of Florida, Gainesville. The Florida strain of ZYMV was used to study the inheritance of resistance. Isolate 2088, a severe isolate of ZYMV-FL, was described by Wisler et al. (1995). The ZYMV isolate used for our research was a subculture of isolate 2088, maintained on ‘Gray Zucchini’ squash (Cucurbita pepo L.) from Seminis Vegetable Seeds (Woodland, CA). The inoculum was prepared by grinding infected ‘Gray Zucchini’ leaves using a mortar and pestle in 0.02M phosphate buffer (Fig. 1), pH 7.0. The leaf to buffer ratio was 1:5 (1 g infected leaf to 5 mL buffer). To maintain the ZYMV-FL isolate and increase the inoculum, we used the rub method (Guner et al., 2002). Squash plants were inoculated by dusting one leaf on each 3-week-old plant with 800-mesh carborundum and then applying the inoculum to the leaf with a pestle, which was rotated in a circular motion eight to 10 times as if painting the leaf with inoculum. After inoculation, carborundum was rinsed off the leaves to improve light interception, and the plants were maintained in aphid-proof cages. All ‘Gray Zucchini’ plants were seeded in Metro-Mix 200 (Scotts-Sierra Horticultural Products Company, Marysville, OH) in 160 mm diameter (1550 mL volume) clay pots. Plants were fertilized weekly with 150 mg·kg Peters Professional 20–20–20 N–P–K (Scotts-Sierra Horticultural Products Company). Inoculation