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© 2009 Plant Management Network.
Accepted for publication 3 June 2009. Published 24 August 2009.


Symptomatology of Iris yellow spot virus in Selected Indicator Hosts


Sudeep Bag and H. R. Pappu, Department of Plant Pathology, Washington State University, Pullman, WA 99164


Corresponding author: H. R. Pappu.  hrp@wsu.edu


Bag, S., and Pappu, H. R. 2009. Symptomatology of Iris yellow spot virus in selected indicator hosts. Online. Plant Health Progress doi:10.1094/PHP-2009-0824-01-BR.


Thrips-transmitted Iris yellow spot virus (genus Tospovirus, family Bunyaviridae) is an economically important pathogen of onion bulb and seed crops in the USA and other parts of the world (2,4). Studies on biological characteristics of the virus have been limited due to difficulties in obtaining consistent and reproducible mechanical transmission and lack of indicator hosts. The existence of strains of IYSV is not known though differences in symptomatology, disease severity, and yield losses could be attributed to the presence of different IYSV strains. To better understand the biological diversity of IYSV, several plant species were evaluated for their response to mechanical inoculation with IYSV under controlled greenhouse conditions.

Seeds of Arabidopsis thaliana COL 1, Capsicum annuum (Serrano pepper), Cerastium glomeratum (mouse ear chickweed), Chenopodium quinoa, Datura innooxia, D. ferox, D. stramonium, Nicotiana benthamiana, N. tabacum, Solanum melongena (egg plant), and Vigna unguiculata (Heirloom variety) were sown in small trays with insert of 36 cells each. Seeding was done in SunShine potting mix LC1. Seedlings at 2 to 6 leaf stage were transplanted to bigger pots of 6 to 10 inches in diameter. IYSV-infected onion leaf tissue (Fig. 1, a-b) from Prosser, WA, was homogenized by grinding in 0.01 M sodium phosphate buffer (pH 7.0) containing 4% β-Mercaptoethanol and the homogenate was used as the virus inocula for mechanical inoculations. Fully expanded young leaves were dusted with carborandum (600 mesh) and the virus inocula were manually applied using cotton buds. The inoculated plants were maintained at 25/18 °C (day/night) and observed for development of symptoms. Uninoculated, healthy plants of respective plants species were maintained in the same greenhouse and were tested by RT-PCR for nucleocapsid protein gene of IYSV and TSWV to make sure that they remained virus-free throughout the duration of the experiment. Each experiment was repeated at least three times. The original source plant (onion) had symptoms typical of IYSV infection and IYSV was confirmed by both ELISA and RT-PCR. None of the other known onion-infecting viruses could be detected in this plant. The experiment was performed in an insect-free, climate-controlled green house with a strict insecticidal regimen to avoid infection of test plants by any other vector borne-viruses.


 

Fig. 1. (a-b) Symptoms of Iris yellow spot virus (IYSV) infection on onion scapes; (c-j) symptoms on indicator hosts following mechanical inoculation with IYSV. (c-f) Datura stramonium; (g-j) Nicotiana benthamiana showing development of chlorotic local lesions, veinal necrosis, stem necrosis and ultimately drying of leaves and plants.

 

In D. stramonium the plants showed 25 to 30 small chlorotic local lesions initially of 2 to 5 mm (Fig.1, c and d) 10 to 12 days post inoculation (DPI). The numbers of local lesions gradually increased and spread throughout the leaves within 20 to 25 days, and as the lesions coalesced the leaves dried 35 to 40 DPI (Fig.1, e and f). The presence of IYSV in inoculated leaves was confirmed by both DAS- ELISA using a commercially available kit (Agdia Inc., Elkhart, IN) and RT-PCR (3). The virus remained localized and did not spread systematically.

In N. benthamiana, chlorotic local lesion (Fig 1, g) appeared 7 to 10 DPI which subsequently expanded leading to drying of leaves by 20 to 25 DPI. The virus also spread systematically in the inoculated plants showing severe veinal necrosis and some stem necrosis (Fig.1, h and i). Infected plants died by 40 to 50 DPI (Fig.1, j). Infection by IYSV was confirmed by DAS-ELISA and RT-PCR both in inoculated and systematically infected leaves.

In V. unguiculata, symptoms appeared as necrotic spots in inoculated leaves 5 to 6 DPI (Fig. 2, a to d), and there was no clear chlorotic lesion in the inoculated leaves of C. annuum (Fig. 2, e to g) and C. quinoa (Fig.2, h and j) developed small concentric chlorotic rings spots, which gradually increased in size, symptoms typical of tospovirus infection. Only the inoculated leaves were positive for IYSV in DAS-ELISA and RT-PCR, and no systemic infection could be seen. Other experimental hosts A. thaliana COL 1, C. glomeratum, D. innooxia, D. ferox, N. tabacum, and S. melongena tested by mechanical inoculation didn’t show any symptoms and were negative for IYSV when tested by DAS-ELISA.


 

Fig. 2. Symptoms on indicator hosts following mechanical inoculation with Iris yellow spot virus (IYSV): (a-d) symptoms on Vigna unguiculata showing diffused necrotic spots; (e-g) Capsicum annuum and (h-j) Chenopodium quinoa, both showing development of chlorotic ring spot.

 

There are no reports of natural infection of C. annuum by IYSV although C. annuum is a host for other tospoviruses such as Capsicum chlorosis virus (CaCV), Chrysanthemum stem necrosis virus (CSNV), Groundnut bud necrosis virus (GBNV), Impatiens necrotic spot virus (INSV), Tomato spotted wilt virus (TSWV), Tomato yellow ring spot virus (TYRV), Watermelon bud necrosis virus (WBNV), and Watermelon silver mottle virus (WSMoV) (1). Our studies showed that C. annum could be experimentally infected with IYSV. In areas where IYSV is present, future surveys of C. annum should include testing for IYSV.

Based on the results obtained, C. quinoa, D. stramonium, N. benthamiana, and V. unguiculata could be useful as experimental hosts for biological studies of IYSV.


Table 1. Symptoms of Iris yellow spot virus (IYSV) in various plant species in response to mechanical inoculation.

  Host Days to symptom appearance Symptoms* Response to DAS- ELISA and RT-PCR
Local Systemic
1 Arabidopsis thaliana COL 1 [-]
2 Capsicum annuum 7-10 CRS inoculated leaves [+], systemic [-]
3 Cerastium glomeratum [-]
4 Chenopodium quinoa 7-10 CRS inoculated leaves [+], systemic [-]
5 Datura stramonium 10-12 CLL, DP
[30-45 days]
  [+]
6 Datura innooxia [-]
7 Datura ferox [-]
8 Nicotiana benthamiana 7-10 CLL CLL, VN, SN, DL, DP [+]
9 Nicotiana tabacum [-]
10 Solanum melongena [-]
11 Vigna unguiculata 5-6 NS   [+]

 * Symptoms are abbreviated as: CLL = chlorotic local lesion, CRS = chlorotic ring spot, VN = veinal necrosis, SN = stem necrosis, NS = necrotic spot, DP = drying/death of plants, DL = drying of leaves.


Acknowledgments

Supported in part by funding from the USDA Western Region IPM Grant Program (No. 2007-03622), USDA-CSREES Specialty Crops Research Initiative, and the Washington State Commission for Pesticide Registration. PPNS No. 0521, Department of Plant Pathology, College of Agricultural, Human and Natural Resource Sciences, Agricultural Research Center, Project No. WNP0 0545.


Literature Cited

1. Campbell, L. R., Ullman, D. E., and Robb, K. L. 2006. The complete tospovirus resource page. Online. K-State Research and Extension, Manhattan, KS.

2. Gent, D., du Toit, L. J., Fichtner, S. F., Krishna Mohan, S., Pappu, H. R., and Schwartz. H. F. 2006. Iris yellow spot virus: An emerging threat to onion bulb and seed production. Plant Dis. 90:1468-1480.

3. Pappu, H. R., Rosales, I. M., Druffel, K. L. 2008. Serological and molecular assays for rapid and sensitive detection of Iris yellow spot virus infection of bulb and seed onion crops. Plant Dis. 92: 588–594.

4. Pappu, H. R., Jones, R. A. C., and Jain, R. K. 2009. Global status of tospovirus epidemics in diverse cropping systems: Successes gained and challenges ahead. Virus Res. 141; 219-236.