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© 2007 Plant Management Network.
Accepted for publication 25 September 2007. Published 19 November 2007.


First Report of Monarda didyma Infection with Impatiens necrotic spot virus in Washington State


Rayapati A. Naidu, Olufemi J. Alabi, Gandhi Karthikeyan, and Mark Nelson, Irrigated Agriculture Research and Extension Center, Department of Plant Pathology, Washington State University, Prosser 99350; and Anne Morrell, Northwest Horticulture LLC, Mabton, WA 98935


Corresponding author: Rayapati A. Naidu.  naidu@wsu.edu


Naidu, R. A., Alabi, O. J., Karthikeyan, G., Nelson, M., and Morrell, A. 2007. First report of Monarda didyma infection with Impatiens necrotic spot virus in Washington State. Online. Plant Health Progress doi:10.1094/PHP-2007-1119-02-BR.


Monarda didyma is an herbaceous perennial widely used as a garden ornamental plant throughout USA. During summers of 2006 and 2007, nearly 90% of M. didyma cv. ‘Gardenview Scarlet’ plants in a one-year-old, 0.1 acre commercial nursery showed general chlorosis of leaves, chlorotic rings and necrotic spots and blotches on the leaves (Fig. 1), tip necrosis of shoots (Fig. 2), and stunting and deformed leaves. Stems of infected plants displayed water-soaked lesions that expanded with time into irregular-shaped necrotic patches (Fig. 3). Incidence of symptomatic plants was high and we did not observe edge effect or other clustering of symptomatic plants. It is unlikely that these symptoms were due to soil-borne pathogens, since the field was fumigated with Vapam (70 gal/acre at 298 lb of Metam Sodium a.i./acre) before planting. No aphids and whiteflies were found on symptomatic plants, but western flower thrips (Frankliniella occidentalis, Pergande) were observed in flowers of symptomatic plants.


 

Fig. 1. M. didyma plants infected with INSV showing leaf symptoms of chlorosis, chlorotic spots, and necrotic spots.

 

     
 

Fig. 2. INSV-infected M. didyma branch showing necrotic blotches on young leaves.

 

Fig. 3. M. didyma plant transplanted from a symptomatic plant in the field showing stem necrosis symptoms. The necrotic patches are distributed on the main stem as well as on side shoots. In advanced stages, the branches die.

 

Symptomatic leaves were tested for Tomato spotted wilt virus (TSWV) and Impatiens necrotic spot virus (INSV) using ImmunoStrip tests (Agdia Inc., Elkhart, IN). Only INSV was detected in diseased plants. To further confirm the presence of INSV, total RNA was extracted from symptomatic leaves using a RNeasy plant minikit (Qiagen Inc., Valencia, CA) and used for reverse transcription-polymerase chain reaction (RT-PCR) amplification (RT at 52°C for 30 min and 35 cycles of PCR with each cycle consisting of 30 sec denaturation at 94°C, 45 sec annealing at 56°C, and 60 sec extension at 72°C followed by 7 min final extension step at 72°C) of the nonstructural protein (NSs) gene of INSV. The sequences of the forward and reverse primers used for RT-PCR were 5’-ATGTCTAGTGCAATGTATGAAAC-3’ (identical to nt 63-85 of INSV S-RNA, GenBank Accession no. NC003624) and 5’-GTTAGTTTAAATCTAATTTAG-3’ (complementary to nt 1393-1415), respectively. These primers directed the amplification of a PCR product of about 1300 nucleotides from total RNA from symptomatic leaves (Fig. 4). No product was amplified from total RNA from leaves of non-symptomatic plants. To verify that the amplified products were derived from INSV RNA, the PCR fragment was cloned into pCR2.1 vector (Invitrogen Corp, Carlsbad, CA) and three independent clones were sequenced in both directions. Sequence analyses of these clones showed 100 per cent nucleotide sequence identity among themselves (GenBank accession number: EU095193) and 98 percent nucleotide sequence identity with corresponding sequences of INSV isolates from Japan (AB109100), Italy (DQ425096), and the Netherlands (NC003624).


 

Fig. 4. RT-PCR analysis of total RNA extracted from leaves of four field-grown M. didyma plants: two asymptomatic and apparently healthy plants (H) and two plants with symptoms consistent with INSV infection (I). A DNA fragment of approximately 1.3 kilobase pairs specific to the NSs gene of INSV was amplified only from symptomatic plants in Figs. 1 and 2. DNA molecular weight markers (Mr) are shown on either side.

 

We did not determine whether the original infection of INSV came from outside sources or through the infected transplanted cuttings. Infection of M. didyma by INSV in commercial greenhouse settings has been reported only in Pennsylvania in 1992 (2). Therefore, this is the first report of INSV infecting a M. didyma in the field.

This report adds to existing knowledge on distribution of INSV in ornamentals (1) with implications for the nursery industry in Washington State. Cuttings from infected plugs may serve as source of virus when distributed to retail garden centers and landscapers. Once introduced into a new location, resident vector thrips species could spread INSV to other ornamental and nursery stock plants. Primary propagators should insure that the original source plants are free of INSV before propagation and shipment to wholesale and retail nurseries within and outside the state.


Literature Cited

1. Daughtrey, M. L., Jones, R. K., Moyer, J. W., Daub, M. E., and Baker, J. R. 1997. Tospoviruses strike the greenhouse industry: INSV has become a major pathogen of flower crops. Plant Dis. 81:1220-1230.

2. Hausbeck, M. K., Welliver, R. A., Derr, M. A., and Gildow, F. E. 1992. Tomato spotted wilt virus survey among greenhouse ornamentals in Pennsylvania. Plant Dis. 76:795-800.