First Report of Oidium sp. Powdery Mildew and Tomato spotted wilt virus on Melampodium divaricatum

Melampodium divaricatum (Rich. ex Pers.) DC. (=M. paludosum H.B.K.), a member of the family Asteraceae and native to South America, is a recent introduction for use as a summer bedding ornamental. In September 1999, melampodium plants in multiple Baton Rouge landscapes were observed with signs of powdery mildew and symptoms of a virus-like disease. Powdery mildew spread throughout one of the plantings by late November and infected flowers and leaves. An Oidium species sporulated on both leaf surfaces but was more common on the adaxial surface. Ellipsoid conidia were produced in chains, lacked fibrosin bodies, and averaged 31 × 19 µm. No sexual stage was observed. Eight of 63 plants (cv. Derby) in one of the plantings showed virus disease symptoms that included severe leaf mosaic, leaf malformation, and stunting. Leaves from infected plants were used to sap inoculate seedling plants of melampodium and Nicotiana benthamiana. Melampodium seedlings developed typical mosaic symptoms after 48 to 56 days. N. benthamiana developed severe chlorosis and mosaic, then wilted and died after 14 days. Noninoculated plants of both species remained healthy. The virus in both plant species was identified as Tomato spotted wilt virus (TSWV) by enzyme-linked immunosorbent assay (ELISA) (Agdia, Elkhart, IN). ELISA tests for presence of Impatiens necrotic spot virus were negative. This is the first report of powdery mildew and TSWV on M. divaricatum.

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First Report of Tomato spotted wilt virus in Blackberry Lily in North America

Plant Disease ◽

10.1094/pdis.2003.87.1.102c ◽

2003 ◽

Vol 87 (1) ◽

pp. 102-102 ◽

Cited By ~ 3

Author(s):

S. Adkins ◽

L. Breman ◽

C. A. Baker ◽

S. Wilson

Keyword(s):

North America ◽

Tomato Spotted Wilt Virus ◽

Amino Acid Sequences ◽

South Florida ◽

N Gene ◽

Enzyme Linked Immunosorbent Assay ◽

Rt Pcr ◽

First Report ◽

Tomato Spotted Wilt ◽

Wilt Virus

Blackberry lily (Belamcanda chinensis (L.) DC.) is an herbaceous perennial in the Iridaceae characterized by purple-spotted orange flowers followed by persistent clusters of black fruit. In July 2002, virus-like symptoms including chlorotic ringspots and ring patterns were observed on blackberry lily leaves on 2 of 10 plants in a south Florida ornamental demonstration garden. Inclusion body morphology suggested the presence of a Tospovirus. Tomato spotted wilt virus (TSWV) was specifically identified by serological testing using enzyme-linked immunosorbent assay (Agdia, Elkhart, IN). Sequence analysis of a nucleocapsid (N) protein gene fragment amplified by reverse transcription-polymerase chain reaction (RT-PCR) with primers TSWV723 and TSWV722 (1) from total RNA confirmed the diagnosis. Nucleotide and deduced amino acid sequences of a 579 base pair region of the RT-PCR product were 95 to 99% and 95 to 100% identical, respectively, to TSWV N-gene sequences in GenBank. Since these 2-year-old plants were grown on-site from seed, they were likely inoculated by thrips from a nearby source. Together with a previous observation of TSWV in north Florida nursery stock (L. Breman, unpublished), this represents, to our knowledge, the first report of TSWV infection of blackberry lily in North America although TSWV was observed in plants of this species in Japan 25 years ago (2). References: (1) S. Adkins, and E. N. Rosskopf. Plant Dis. 86:1310, 2002. (2) T. Yamamoto and K.-I. Ohata. Bull. Shikoku Agric. Exp. Stn. 30:39, 1977.

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First Report of Tomato spotted wilt virus on Chrysanthemum in Serbia

Plant Disease ◽

10.1094/pdis-08-12-0778-pdn ◽

2013 ◽

Vol 97 (1) ◽

pp. 150-150 ◽

Cited By ~ 7

Author(s):

I. Stanković ◽

A. Bulajić ◽

A. Vučurović ◽

D. Ristić ◽

K. Milojević ◽

...

Keyword(s):

Tomato Spotted Wilt Virus ◽

Disease Incidence ◽

N Gene ◽

Impatiens Necrotic Spot Virus ◽

Maximum Parsimony Tree ◽

Rt Pcr ◽

First Report ◽

Tomato Spotted Wilt ◽

Negative Controls ◽

Wilt Virus

In July 2011, greenhouse-grown chrysanthemum hybrid plants (Chrysanthemum × morifolium) with symptoms resembling those associated with tospoviruses were observed in the Kupusina locality (West Bačka District, Serbia). Disease incidence was estimated at 40%. Symptomatic plants with chlorotic ring spots and line patterns were sampled and tested by double antibody sandwich (DAS)-ELISA using polyclonal antisera (Bioreba AG, Reinach, Switzerland) against the two of the most devastating tospoviruses in the greenhouse floriculture industry: Tomato spotted wilt virus (TSWV) and Impatiens necrotic spot virus (INSV) (2). Commercial positive and negative controls and extracts from healthy chrysanthemum tissue were included in each ELISA. TSWV was detected serologically in 16 of 20 chrysanthemum samples and all tested samples were negative for INSV. The virus was mechanically transmitted from ELISA-positive chrysanthemum samples to five plants each of both Petunia × hybrida and Nicotiana tabacum ‘Samsun’ using chilled 0.01 M phosphate buffer (pH 7) containing 0.1% sodium sulfite. Inoculated plants produced local necrotic spots and systemic chlorotic/necrotic concentric rings, consistent with symptoms caused by TSWV (1). The presence of TSWV in ELISA-positive chrysanthemum plants and N. tabacum‘Samsun’ was further confirmed by conventional reverse transcription (RT)-PCR. Total RNAs were extracted with an RNeasy Plant Mini Kit (Qiagen, Hilden, Germany). RT-PCR was performed with the One-Step RT-PCR Kit (Qiagen) using primers TSWVCP-f/TSWVCP-r specific to the nucleocapsid protein (N) gene (4). A Serbian isolate of TSWV from tobacco (GenBank Accession No. GQ373173) and RNA extracted from a healthy chrysanthemum plant were used as positive and negative controls, respectively. An amplicon of the correct predicted size (738-bp) was obtained from each of the plants assayed, and that derived from chrysanthemum isolate 529-11 was purified (QIAqick PCR Purification Kit, Qiagen) and sequenced (JQ692106). Sequence analysis of the partial N gene, conducted with MEGA5 software, revealed the highest nucleotide identity of 99.6% (99% amino acid identity) with 12 TSWV isolates deposited in GenBank originating from different hosts from Italy (HQ830186-87, DQ431237-38, DQ398945), Montenegro (GU355939-40, GU339506, GU339508), France (FR693055-56), and the Czech Republic (AJ296599). The consensus maximum parsimony tree obtained on a 705-bp partial N gene sequence of TSWV isolates available in GenBank revealed that Serbian TSWV isolate 529-11 from chrysanthemum was clustered in the European subpopulation 2, while the Serbian isolates from tomato (GU369723) and tobacco (GQ373172-73 and GQ355467) were clustered in the European subpopulation 1 denoted previously (3). The distribution of TSWV in commercial chrysanthemum crops is wide (2). To our knowledge, this is the first report of TSWV infecting chrysanthemum in Serbia. Since chrysanthemum popularity and returns have been rising rapidly, the presence of TSWV may significantly reduce quality of crops in Serbia. References: (1) Anonymous. OEPP/EPPO Bull. 34:271, 2004. (2) Daughtrey et al. Plant Dis. 81:1220, 1997. (3) I. Stanković et al. Acta Virol. 55:337, 2011. (4) A. Vučurović et al. Eur. J. Plant Pathol. 133:935, 2012.

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First Report of Tomato spotted wilt virus on Soybean in Iran

Plant Disease ◽

10.1094/pdis.2001.85.12.1290b ◽

2001 ◽

Vol 85 (12) ◽

pp. 1290-1290 ◽

Cited By ~ 12

Author(s):

A. R. Golnaraghi ◽

N. Shahraeen ◽

R. Pourrahim ◽

Sh. Ghorbani ◽

Sh. Farzadfar

Keyword(s):

Glycine Max ◽

Tomato Spotted Wilt Virus ◽

Chenopodium Quinoa ◽

Datura Stramonium ◽

Enzyme Linked Immunosorbent Assay ◽

Mechanical Inoculation ◽

First Report ◽

Tomato Spotted Wilt ◽

Soybean Leaves ◽

Wilt Virus

During the summers of 1999 and 2000, 3,110 soybean (Glycine max) leaf samples were randomly collected from soybean fields in the Ardebil, Goletan, Khuzestan, Lorestan, and Mazandaran provinces of Iran. Tomato spotted wilt virus (TSWV) was detected in leaf samples by TSWV-specific polyclonal antibody (As-0526 and As-0580, DSMZ, Braunschweig, Germany) in double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). Mechanical inoculation of 26 plant species (10 plants per species) and cultivars with extracts of positive leaf samples produced necrotic local lesions in Beta vulgaris, Chenopodium quinoa, C. amaranticolor, Gomphrena globosa, Phaseolus vulgaris cv. Talash, Vicia faba, and Vigna unguiculata cv. Mashad; produced systemic chlorosis followed by necrosis in Datura stramonium, D. metel, Nicotiana rustica, N. tabacum cv. Samsun, N. glutinosa, N. bentamiana, and Glycine max cv. Hill; and produced chlorosis, stunting, and bud necrosis in Arachis hypogaea (peanut). Plants developing these symptoms following mechanical inoculation with extracts from original soybean leaves were positive in ELISA for TSWV. ELISA results indicate that the overall incidence of TSWV on soybean in the five provinces was 5.4%. TSWV has been reported in potato (2) and tomato (1) from Iran, but to our knowledge, this is the first report of the occurrence of TSWV on soybean in Iran. References: (1) K. Bananej et al. Iran. J. Plant Pathol. 34:30, 1998. (2) R. Pourrahim et al. Plant Dis. 85:442, 2001.

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First Report of Tomato spotted wilt virus in Soybean (Glycine max) in Georgia

Plant Disease ◽

10.1094/pd-90-0524b ◽

2006 ◽

Vol 90 (4) ◽

pp. 524-524 ◽

Cited By ~ 12

Author(s):

C. Nischwitz ◽

S. W. Mullis ◽

R. D. Gitaitis ◽

A. S. Csinos

Keyword(s):

Glycine Max ◽

Large Scale ◽

Tomato Spotted Wilt Virus ◽

The United States ◽

Enzyme Linked Immunosorbent Assay ◽

First Report ◽

Nucleocapsid Gene ◽

Fta Cards ◽

Tomato Spotted Wilt ◽

Wilt Virus

Tomato spotted wilt virus (TSWV) is a member of the family Bunyaviridae and has a wide host range including important crops such as tomato, pepper, tobacco, peanut, and onion. In areas of Georgia, soybean (Glycine max) is double cropped between two onion crops and as a rotation crop with peanuts. Soybeans do not show any TSWV symptoms, and therefore, have not been tested on a large scale for the virus. However, because symptomless weed and crop plants provide a reservoir for TSWV and the thrips vectors (2), a survey was conducted during the summer of 2005 to evaluate the occurrence of TSWV in soybean. The survey took place in seven counties in southern Georgia with field sizes ranging between 0.4 and 20 ha (1 and 50 acres). Soybean cultivars included Haskell, DP7220, DP6770, Pioneer 97B52, and Vigoro V622NRR. Of 848 randomly selected plants tested using the double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) (Agdia, Inc., Elkhart, IN), 6.6% tested positive for TSWV. Plants testing positive ranged from seedling to the pod-setting stages. Leaves and roots of several plants tested positive, indicating a systemic infection. Soybean plants testing positive using ELISA were blotted onto FTA cards (Whatman Inc., Brentford, UK) to bind viral RNA for preservation, and the blotted samples were processed according to the manufacturer's protocol. Reverse transcription-polymerase chain reaction using punch-outs from the FTA cards and TSWV nucleocapsid gene specific forward and reverse primers (5′-TTAAGCAAGTTCTGTGAG-3′ and 5′-ATGTCTAAGGTTAAGCTC-3′), respectively (4), confirmed the identity of TSWV. TSWV has been found in soybean in other parts of the world (1) but has only been reported in the United States in a survey from Tennessee (3). To our knowledge, this is the first report of the occurrence of TSWV in soybean in Georgia. The role soybean plays as a reservoir or green bridge for thrips and TSWV is currently unknown. References: (1) A. R. Golnaraghi et al. Plant Dis. 88:1069, 2004. (2) R. L. Groves et al. Phytopathology 91:891, 2001. (3) B. S. Kennedy and B. B. Reddick. Soybean Genet. Newsl. 22:197, 1995. (4) H. R. Pappu et al. Tob. Sci. 40:74, 1996.

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First Report of Tomato spotted wilt virus in Leek (Allium porrum) in the United States

Plant Disease ◽

10.1094/pd-90-0525a ◽

2006 ◽

Vol 90 (4) ◽

pp. 525-525 ◽

Cited By ~ 2

Author(s):

C. Nischwitz ◽

S. W. Mullis ◽

R. D. Gitaitis ◽

A. S. Csinos ◽

S. M. Olson

Keyword(s):

Tomato Spotted Wilt Virus ◽

The United States ◽

Close Relative ◽

Enzyme Linked Immunosorbent Assay ◽

Allium Porrum ◽

First Report ◽

Nucleocapsid Gene ◽

Fta Cards ◽

Tomato Spotted Wilt ◽

Wilt Virus

Tomato spotted wilt virus (TSWV) is a member of the family Bunyaviridae. It has many important crop hosts including tomato, pepper, tobacco, peanut, and onion. In Georgia, Vidalia onions (Allium cepa), a close relative of leek, can be infected by TSWV and Iris yellow spot virus (IYSV), which is another thrips-vectored tospovirus (2). For this reason, samples of leek transplants with virus-like symptoms in one field at the border of Georgia and Florida were tested for the presence of TSWV and IYSV. The transplants had been grown from seed in a greenhouse at the same location. The sampled plants exhibited extended bleaching of leaf tips and necrotic lesions. These symptoms were also seen on onion plants infected with TSWV and IYSV. The only natural infections of leek with IYSV have been reported thus far only from Reunion Island (4) and Slovenia (1), but to our knowledge, TSWV has not been reported as a pathogen of leek. Green tissue near the necrotic lesions and bleached tips of one symptomatic leaf per plant was sampled and analyzed using a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) (Agdia, Inc., Elkhart, IN). Of 90 plants tested, eight were positive for TSWV and none were positive for IYSV. Leek samples testing positive using ELISA were blotted onto FTA cards (Whatman Inc., Brentford, UK) to bind viral RNA for preservation and then processed according to the manufacturer's protocol. Punch-outs from the FTA cards were used for reverse transcription polymerase chain reaction (RT-PCR) with the TSWV-specific forward primer (5′-TTAAGCAAGTTCTGTGAG-3′) and reverse primer (5′-ATGTCTAAGGTTAAGCTC-3′) (3) to confirm the identity of TSWV. The primers are specific to the viral nucleocapsid gene. An amplicon of the expected size (774 bp) was produced from TSWV ELISA-positive leek plants, but not from healthy controls. TSWV has been found in many plants worldwide, but to our knowledge this is the first report of TSWV infecting leek. The effect that TSWV has on leek production is currently unknown. References: (1) D. A. Benson et al. Nucleic Acids Res. 1:32 (Database issue):D23-6, 2004. (2) S. W. Mullis et al. Plant Dis. 88:1285, 2004. (3) H. R. Pappu et al. Tob. Sci. 40:74, 1996. (4) I. Robène-Soustrade et al. Online publication. New Dis. Rep. 11, 2005.

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First Report of Tomato Spotted Wilt Virus on Potatoes in Iran

Plant Disease ◽

10.1094/pdis.2001.85.4.442b ◽

2001 ◽

Vol 85 (4) ◽

pp. 442-442 ◽

Cited By ~ 14

Author(s):

R. Pourrahim ◽

Sh. Farzadfar ◽

A. A. Moini ◽

N. Shahraeen ◽

A. Ahoonmanesh

Keyword(s):

Tomato Spotted Wilt Virus ◽

Chenopodium Quinoa ◽

Plant Viruses ◽

Thrips Tabaci ◽

Enzyme Linked Immunosorbent Assay ◽

Leaf Extracts ◽

Systemic Necrosis ◽

First Report ◽

Tomato Spotted Wilt ◽

Wilt Virus

Severe leaf and stem necrosis before flowering was observed in potato (Solanum tuberosum) fields of Firouzkoh Province, Iran, during the summer of 1998. Infected plants died before the end of the growing season. Necrosis was more severe in cv. Agria than in cvs. Ajaxs and Arinda. A high population of Thrips tabaci was observed in August and September. Tomato spotted wilt virus (TSWV) (1) was detected in affected potatoes by using specific TSWV-IgG (from Bioreba) in double-antibody sandwich enzyme linked immunosorbent assay and by indicator plant reactions. Mechanical inoculation of indicator plants with leaf extracts of symptomatic potatoes produce necrotic local lesions in Chenopodium quinoa, C. amaranticolor, Gomphrena globosa, Vicia faba, Vigna sinensis, Phaseolus aureus var. Gohar, P. vulgaris, and Petunia hybrida. The virus caused systemic necrosis in Capsicum frutescens, Datura stramonium, D. metel, Nicotiana glutinosa, N. rustica, and Trapaeolum majus, preceded by systemic chlorotic spots. TSWV was reported from ornamental crops in Tehran and Absard areas near to Firouzkoh province (2), but this is the first report of TSWV occurrence on potatoes in Iran. References: (1) T. S. Ie. Descriptions of Plant Viruses. No. 39, 1970. (2) A. A. Moeini, et al. Iran. J. Plant Pathol. (In press.)

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First Report of Tomato spotted wilt virus in Common Agapanthus

Plant Disease ◽

10.1094/pdis.2000.84.4.491b ◽

2000 ◽

Vol 84 (4) ◽

pp. 491-491 ◽

Cited By ~ 4

Author(s):

C. R. Wilson ◽

A. J. Wilson ◽

S. J. Pethybridge

Keyword(s):

Tomato Spotted Wilt Virus ◽

Protein Gene ◽

Enzyme Linked Immunosorbent Assay ◽

First Report ◽

Tomato Spotted Wilt ◽

Nucleocapsid Protein Gene ◽

Polymerase Chain ◽

Wilt Virus ◽

Line Patterns ◽

Leaf Symptoms

Common agapanthus (Agapanthus praecox subsp. orientalis), native to South Africa, is a popular ornamental flowering bulb species belonging to the Amaryllidaceae and is commonly found in residential gardens. Roots from some Agapanthus sp. also are used in traditional medicine in Africa. Common agapanthus collected from a residential property in Hobart, Tasmania, Australia, showed leaf symptoms of concentric ring and line patterns, irregular chlorotic blotches, and streaks. Symptomatic plants were severely stunted and failed to flower. Symptomatic leaves prematurely senesced, but young foliage subsequently produced was symptomless. Similar symptoms have been reported in other members of the Amaryllidaceae and are associated with infection by Tomato spotted wilt virus (TSWV; e.g., Nerine and Hippeastrum spp.) or Cucumber mosaic virus (CMV; e.g., Hippeastrum sp.) (2). The presence of TSWV and absence of CMV in symptomatic plants of common agapanthus was determined by enzyme-linked immunosorbent assay. Confirmation of TSWV infection was provided by reverse-transcription polymerase chain reaction assay with primers specific to the nucleocapsid protein gene of TSWV, with nucleic extracts from symptomatic plants producing an expected ≈800-bp amplicon (1). This is the first report of TSWV infection of any species within the Amaryllidaceae in Australia and the first report of the occurrence of TSWV in common agapanthus. References: (1) R. K. Jain et al. Plant Dis. 82:900, 1998. (2) G. Loebenstein et al. 1995. Virus and Virus-like Diseases of Bulb and Flower Crops. John Wiley & Sons, Chichester, U.K.

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Outbreak of Tomato spotted wilt virus in Tomato in Kenya

Plant Disease ◽

10.1094/pdis.2001.85.10.1123b ◽

2001 ◽

Vol 85 (10) ◽

pp. 1123-1123 ◽

Cited By ~ 6

Author(s):

A. W. Wangai ◽

B. Mandal ◽

H. R. Pappu ◽

S. Kilonzo

Keyword(s):

Host Range ◽

Tomato Spotted Wilt Virus ◽

Virus Disease ◽

N Gene ◽

Enzyme Linked Immunosorbent Assay ◽

Rt Pcr ◽

Tomato Spotted Wilt ◽

Wide Host Range ◽

Test Kit ◽

Wilt Virus

Tomato spotted wilt virus (TSWV) of the genus Tospovirus, family Bunyaviridae (1), causes an economically important virus disease in tomato in several parts of the world. The virus has a wide host range that includes numerous crops and weeds and is transmitted by at least seven species of thrips. Tomato crops in the Subukia, Bahati, and Kabazi areas of the Nakuru District in Kenya were affected by a disease suggestive of TSWV infection during the November 1999 to March 2000 tomato-growing season. Farmers reported up to 80% losses of their potential yields. Characteristic symptoms were noticed on fruits, especially when they were green. Distinct concentric rings on fruits, which later turned into brown, uneven ripening, were the most visible symptoms. Foliage did not develop pronounced symptoms, but mild bronzing was observed in a few cultivars. However, foliage senesced prematurely, starting with older leaves. Foliar symptoms were mistaken for blight infection, and as a result, excessive fungicides were applied that failed to manage the disease. To test for TSWV infection, tomato leaf samples collected from the fields were tested initially with a TSWV test kit (HortiTech, Horticulture Research International, Wellesbourne, UK), and the results were confirmed by double-antibody sandwich-enzyme-linked immunosorbent assay with antibodies from Agdia Inc. (Elkhart, IN). Further molecular characterization was done using reverse transcription-polymerase chain reaction (RT-PCR). Total RNA was extracted from symptomatic leaves of tomato cv. Money Maker using the RNeasy mini kit (Qiagen Inc., Valencia, CA). Using primers 5′ TTAAGC AAGTTCTGTGAG 3′ and 5′ ATGTCTAAGGTTAAGCTC 3′ specific to the nucleoprotein (N) gene of TSWV, the N gene was amplified by RT-PCR (2). A 777-bp product of the expected size was obtained from symptomatic plants, whereas no amplification was obtained from noninfected tomato. The PCR product was cloned into pGEM-T Easy (Promega, Madison, WI) and sequenced. A search of GenBank revealed a sequence identity of 95 to 99% with the N genes of known TSWV isolates. To our knowledge, this is the first report TSWV infection of tomato in Kenya. Considering its wide host range, future surveys should be directed toward estimating its incidence in tomato and other TSWV-susceptible crops, such as Irish potatoes, pepper, peanut (groundnut), beans, and a wide variety of ornamental cut flowers in Kenya. References: (1) J. W. Moyer. Tospoviruses (Bunyaviridae). Pages 1803–1807 in: Encyclopedia of Virology. A. Granoff and R. G. Webster, eds. Academic Press, San Diego, CA, 1999. (2) Jain et al. Plant Dis. 82:900, 1998.

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First Report of Tomato spotted wilt virus on Gloxinia in Bosnia and Herzegovina

Plant Disease ◽

10.1094/pdis-08-12-0777-pdn ◽

2013 ◽

Vol 97 (3) ◽

pp. 429-429 ◽

Cited By ~ 2

Author(s):

V. Trkulja ◽

J. Mihić Salapura ◽

B. Ćurković ◽

I. Stanković ◽

A. Bulajić ◽

...

Keyword(s):

Bosnia And Herzegovina ◽

Tomato Spotted Wilt Virus ◽

Amino Acid Identity ◽

N Gene ◽

Impatiens Necrotic Spot Virus ◽

Post Inoculation ◽

Rt Pcr ◽

First Report ◽

Tomato Spotted Wilt ◽

Wilt Virus

In June and July 2012, symptoms resembling those caused by a tospovirus infection were observed on the greenhouse-grown gloxinia (Sinningia speciosa Benth. and Hook.) in the Lijevče polje, in the vicinity of Banja Luka (Bosnia and Herzegovina). Infected plants exhibited chlorotic ring spots and chlorotic and necrotic patterns followed by necrosis and distortion of leaves. Disease symptom incidence was estimated at 30% out of 400 inspected plants. Symptomatic leaves were collected and tested by double-antibody sandwich (DAS)-ELISA test using commercial polyclonal antisera (Bioreba AG, Reinach, Switzerland) for two of the most important tospoviruses in the greenhouse production of ornamentals: Tomato spotted wilt virus (TSWV) and Impatiens necrotic spot virus (INSV) (2). TSWV was detected serologically in 27 out of 30 tested gloxinia samples, and all were negative for INSV. Symptomatic leaves of five selected ELISA-positive gloxinia plants were separately ground in chilled 0.01 M phosphate buffer (pH 7) containing 0.1% w/v sodium sulphite and were mechanically inoculated on five plants of Petunia × hybrida. All inoculated plants produced typical symptoms of TSWV (1), necrotic spots on inoculated leaves in 2 to 5 days post-inoculation. For further confirmation of TSWV infection, total RNAs were extracted using the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany) from all 27 infected gloxinia plants and tested by reverse transcription (RT)-PCR assay. A 738-bp fragment of TSWV nucleocapsid (N) gene was amplified with One-Step RT-PCR Kit (Qiagen) using primer pairs TSWV CP-f and TSWV CP-r (4). Total RNAs from Serbian tobacco TSWV isolate (GenBank Accession No. GQ373173) and RNA extract from healthy gloxinia plants were used as positive and negative controls, respectively. Amplicons of the expected size were obtained from all 27 naturally infected gloxinia plants, while no amplification products were obtained from the healthy control. After the purification with QIAquick PCR Purification Kit (Qiagen), the RT-PCR product obtained from one selected isolate 160-12 was sequenced directly in both directions and submitted to GenBank (JX468079). Sequence analysis of the partial N gene, conducted by MEGA5 software (3), from isolate 160-12 showed the highest nucleotide identity of 99.7% (100% amino acid identity) with eight pepper isolates of TSWV from Spain (FR693229, FR693231, FR693152-153, FR693078, FR693081, FR693089, and FR693092). To our knowledge, this is the first report on the occurrence of TSWV in Bosnia and Herzegovina. The presence of this harmful pathogen into a new area could have a serious threat to intensive and increasing production of ornamentals and numerous other TSWV susceptible species in Bosnia and Herzegovina. The discovery of TSWV on gloxinia should prompt more surveys, thorough inspections, and subsequent testing of other TSWV susceptible plants cultivated in Bosnia and Herzegovina. References: (1) Anonymous. OEPP/EPPO Bull. 34:271, 2004. (2) Daughtrey et al. Plant Dis. 81:1220, 1997. (3) K. Tamura et al. Mol. Biol. Evol. 28:2731, 2011. (4) A. Vučurović et al. Eur. J. Plant Pathol. 133:935, 2012.

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First Report of Tomato spotted wilt virus and Impatiens necrotic spot virus in Slovenia

Plant Disease ◽

10.1094/pdis.2001.85.12.1288a ◽

2001 ◽

Vol 85 (12) ◽

pp. 1288-1288 ◽

Cited By ~ 11

Author(s):

I. Mavrič ◽

M. Ravnikar

Keyword(s):

Tomato Spotted Wilt Virus ◽

Polyclonal Antiserum ◽

Impatiens Necrotic Spot Virus ◽

Necrotic Spot ◽

Spot Virus ◽

First Report ◽

Tomato Spotted Wilt ◽

Systemic Symptoms ◽

Wilt Virus ◽

Das Elisa

In July 2000, concentric necrotic rings and patterns were observed on greenhouse-grown pepper (Capsicum anuum L. ‘Blondi’). Symptoms were present only on lower leaves, not on young leaves or fruits. Typical tospovirus particles using electron microscopy were observed in leaf-dip preparations of symptomatic leaves. Impatiens necrotic spot virus (INSV) was detected in symptomatic but not in asymptomatic tissues using double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) with polyclonal antiserum (Loewe Biochemica, Sauerlach, Germany). Nicotiana benthamiana, N. rustica, and Petunia sp. were mechanically inoculated with sap of symptomatic leaves. Local and systemic symptoms were observed only on N. benthamiana. Tomato spotted wilt virus (TSWV) infections were later confirmed in some pepper and tomato plants with distinct systemic symptoms and in greenhouse-grown chrysanthemums, calla lilies, cyclamen and spatiphylum using DAS-ELISA with polyclonal antiserum. Severe systemic symptoms were observed only on some chrysanthemum cultivars, where the infection rate of TSWV was between 80 and 100%. Such TSWV-infected plants were not marketable. Symptomless infections by the same virus were also found. At one location, calla lilies were heavily infected by TSWV, but only local symptoms on leaves were observed. INSV was also confirmed using DAS-ELISA on different chrysanthemum cultivars. Mixed infections of TSWV and INSV were detected using DAS-ELISA on calla lilies with local necrotic rings and patterns on leaves and on Impatiens walerana with systemic necrosis. A limited number of weeds from the vicinity of greenhouses containing symptomatic plants were tested for the presence of TSWV using DAS-ELISA, and only Artemisia vulgaris was infected. Both viruses, TSWV from chrysanthemum and INSV from pepper, were isolated on test plants, and their identity was confirmed using DAS-ELISA. For further verification of TSWV and INSV infection, immunocapture reverse-transcription polymerase chain reaction was performed using general tospovirus primers (1). Amplification products of the expected size were detected and sequenced. Comparison of nucleic acid sequences of amplification products with viral sequence databases confirmed the identities of both viruses. To our knowledge, this is the first report of TSWV and INSV infection in ornamental and vegetable plants in Slovenia. Reference: (1) R. J. Weekes et al. Acta Hortic. 431:159, 1996.

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