Several plant viruses infecting floriculture crops have been characterized based on symptomatology, virus vector relationship, particle morphology, genome organization and serological relationship

  

Alstroemeria 

Alstroemeria plants were found to be infected with CMV showing mosaic symptoms  by Double Antibody Sandwich Enzyme Linked Immuno Sorbent Assay (DAS-ELISA) using CMV specific antibodies and (RT-PCR) using CMV specific primers. In RT-PCR the expected product of 540 bp was obtained in virus infected plants.

 

Carnation

 

In a survey of 61 carnation cvs. growing in Himachal Pradesh, exhibiting varied symptoms on the leaves and flowers, were screened through bioassay and DAS-ELISA. It revealed the widespread presence of Carnation vein mottle virus (CVMV) and Carnation ringspot virus (CRSV). RT-PCR further confirmed infected samples for presence of these viruses. Out of these cvs., 47 were infected with CRSV while 34 were infected with CVMV. Besides RT-PCR, Immuno Capture RT-PCR (IC-RT-PCR) was also used for specific detection of CVMV. 



Chrysanthemum 

 

Tomato aspermy virus (TAV) was detected by DAS-ELISA from chrysanthemums exhibiting mottling and deformed inflorescence grown in various states of India. Out of 15 cvs. tested, 11 (73.3%) were positive for TAV. On sap inoculation the virus produced local lesions on Chenopodium amaranticolor, C. album, C. quinoa and Cucumis sativus. Nicotiana clevelandii, N. glutinosa, N. megalosiphon and N. tabacum reacted systemically to the virus producing severe mosaic, leaf deformation and characteristic leaf enations. Tomato plants produced malformed fruits (2-3/plant) with a few seeds. Myzus persicae and Aphis gossypii transmitted the virus non-persistently. Electron microscopy of partially purified viral preparations revealed polyhedral virions (ca 29 nm

 

 

Gerbera 


 The plants showing colour breaking in the flower were infected with CMV. RT-PCR was used to detect the virus using CMV specific primers that gave an amplicon of 540 bp and sequenced.

 

Symptoms induced by Alstromeria Carlavirus on Alstromeria

Gerbera showing colour break infected with CMV

  

Gladiolus
During a survey for viruses infecting gladiolus in Palampur, some plants of 22 cvs. exhibited flower colour-break symptoms, whereas others were symptomless. The plants were found to be infected with Bean yellow mosaic virus (BYMV) when indexed by DAS-ELISA using BYMV specific antibodies. However, symptomless plants of gladiolus cv. Yellow Supreme, when indexed by DAS-ELISA using antibodies to Tobacco ringspot virus (TobRSV) were infected with TobRSV. Sap from these symptomless TobRSV-infected gladioli induced necrotic local lesions 7-10 days after inoculation followed by systemic leaf malformation characteristic of TobRSV in mechanically inoculated Nicotiana clevelendii and Vigna unguiculata seedlings. The presence of TobRSV in these plants was confirmed by DAS-ELISA. This is the first report of TobRSV occurring in gladiolus in India. TobRSV can be transmitted by contaminated tools used during harvesting of flowers and corms, by nematode vectors and gladiolus seeds. For virus management, it is recommend to use disease free planting material, good sanitation practices and vector management.

 

Iris

Iris (Iris x hollandica Hort.) growing in Kangra valley exhibiting mosaic symptoms were tested for Bean yellow mosaic virus (BYMV) by bioassay, DAS-ELISA, electron microscopy, Immunoelectron microscopy (IEM) and RT-PCR. In RT-PCR, BYMV specific primers were used that gave an amplicon of 720 bp. ELISA and IEM were able to detect BYMV only in leaf samples of the two cvs. tested while RT-PCR detected the virus in all plant parts. RT-PCR was a reliable diagnostic test for BYMV in iris bulb.

 

Iris (Iris x hollandica Hort. cv. Bluemagic and Prof. Blaauw) plants showing chlorotic stripes or mosaic patterns (typical of Iris severe mosaic virus, ISMV) were screened for presence of the virus by DAS-ELISA. Cultivar Bluemagic was severely infected in comparison to Prof. Blaauw. IEM with ISMV specific antiserum also showed presence of the virus.

 

Lily 

Electron microscopy was carried out for CMV , Lily symptomless virus (LSV)  and Tulip breaking virus (TBV)

 

Ornithogalum

Ornithogalum thyrsoides exhibited severe mosaic on the leaves. The causal virus was transmitted mechanically causing local lesions on Chenopodium album and C. amaranticolor, and systemic mosaic on Nicotiana clevelandii, N. glutinosa, N. megalosiphon and N. tabacum cv White Burley. In ELISA, the virus reacted with Cucumber mosaic virus (CMV)-specific antibodies. Leaf dip preparations from virus infected plants in electron microscopy revealed the presence of ca. 29nm isometric particles. In immuno-electron microscopy (IEM), the virus was readily trapped and decorated with CMV-specific antibodies.

Ultrastructure of virus infected cells showing cytoplasmic cylindrical inclusions, laminated aggregates, pin wheels and crystalline inclusions in lily

Ornithogalum

 

Spider lily

In a survey of spider lily (Hymenocallis littoralis), in and around Palampur, plants with leaves exhibiting curling, chlorotic striping and yellowing, characteristic of Lily  symptomless virus (LSV) were found. Leaves were tested by DAS-ELISA using LSV specific antibodies and also confirmed by RTPCR using a primer pair specific to the 3' terminal region of the virus. An expected amplicon of ~900bp was obtained.  Spider lily samples were dot blotted, probed with the cloned LSV DNA and the identity of the virus was further confirmed. Some plants that tested negative by ELISA, tested positive by dot hybridization. Pelargonium Prunus necrotic ringspot virus (PNRSV) was detected in Pelargonium sp. by ELISA, Southern hybridization  and RT-PCR.

 

Tuberose

Tuberose   (Polianthes tuberosa) showed the presence of mosaic symptoms on the leaves and flower peduncles. All the plants were heavily infected indicating its wide spread nature. Causal agent could only be transmitted mechanically to tuberose, indicating a narrow host range. Further, the virus could be transmitted by aphid vector in non-persistent manner. Positive results were obtained by employing potyvirus group specific antibody in ELISA. RT-PCR using group specific primer for its detection and non-persistent insect transmission confirmed the presence of potyvirus.

 

 Buddleja crispa: Buddleja crispa (Buddlejaceae), commonly known as Himalayan-butterfly bush, grows on open rocky places. In the course of a study to assess virus incidence in natural plant resources of high altitude areas in western Himalayan region of India, some leaves of B. crispa with mild chlorosis and mosaic symptoms were collected from Kinnaur area (altitude 2500 m amsl) of Himachal Pradesh during 2003. The infected leaves were tested by ELISA for presence of Tomato ringspot virus (DSMZ, Braunschweig, Germany), Cucumber mosaic virus and potyviruses (Agdia, Elkhart, USA). A positive result was obtained with the potyvirus group-specific antibodies.

 

Murraya koenigii: Murraya koenigii (L.) Spreng., a small, strong-smelling umbrageous tree with subcampanulate white flowers belonging to the family Rutaceae. In a survey of potyvirus incidence in the northwestern Himalaya foothills of the Kangra and Hamirpur districts in the state of Himachal Pradesh in 2004, M. koenigii plants showing mosaic symptoms on leaves, typical of a virus infection, were frequently observed. Symptomatic leaves were tested for the presence of several viruses using enzyme-linked immunosorbent assay with specific antibodies. Positive results were obtained with potyvirus group specific antibodies. To our knowledge, this is the first report of a potyvirus naturally occurring on a Murraya sp.

 

Sorbaria tomentosa: Sorbaria tomentosa syn. Spiraea sorbifolia (Family Rosaceae), common name False Spiraea, is a large, woody shrub with creamy-white flowers. During a virus survey in natural plant vegetation of the Northwest Himalaya of Himachal Pradesh, Sorbaria plants showing leaf mosaic symptoms were observed. Symptomatic leaves were screened for the presence of viruses by ELISA using specific antibodies. Positive results were obtained using antibodies specific to the potyvirus group (Agdia, Elkhart, USA).

 

Tulip: Different varieties of Tulip grown in H.P. and J&K (India) have been found to exhibit various symptoms: yellowing, chlorotic striping, vein clearing and deformed flower. Asiatic and Oriental hybrid lily were originally imported from the Netherlands and have been under cultivation for more then 10 years. Tulip was procured from Shrinagar, Northern India. The lily and tulip were investigated for the presence of LSV by DAS-ELISA and RT-PCR, and the coat protein sequence was characterized and the variability examine.

 

Rose 

Rose plants were found to be infected with a viral disease exhibiting mosaic, line pattern symptoms.  The virus was identified as Prunus necrotic ringspot virus (PNRSV) by DAS-ELISA using PNRSV specific antibodies.

 

 

Symptoms induced by PNRSV on Begonia 

 

Survey & Identification of Viruses Infecting mandate crops:

 

 

» Aglaonema: Dasheen mosaic potyvirus (DsMV)

 

» Alstromeria: Alstroemeria mosaic potyvirus (AlMV), Tospovirus, Cucumber mosaic  virus (CMV) and Alstroemeria carlavirus

 

» Asparagus: Asparagus 1 potyvirus, Asparagus 2 ilarvirus, Asparagus 3 potexvirus

 

» Begonia:Carnation mottle carmovirus (CarMV), Tospovirus, Prunus necrotic  ringspot virus (PNRSV)

 

» Carnation:Carnation latent carlavirus (CLV), Carnation ringspot dianthovirus (CRSV), Carnation etched ring caulimovirus (CERV), CarMV, Carnation necrotic fleck closterovirus (CNFV), Carnation vein mottle  potyvirus (CVMV)

 

» Chrysanthemum: Chrysanthemum B carlavirus (CVB), Chrysanthemum aspermy  cucumovirus (CAV), CMV, Chrysanthemum stunt viroid (CSVd)

 

» Gladiolus: Bean yellow mosaic potyvirus (BYMV), CMV, PNRSV, Tobacco mosaic tobamovirus (TMV), Tobacco rattle (TRV),  Tobacco ringspot nepovirus (TRSV), Tospovirus Iris: BYMV, Iris mild mosaic potyvirus (IMMV), Iris severe mosaic potyvirus   (ISMV), 

            

» Lily: Arabis mosaic virus (ArMV), CMV, Lily symptomless carlavirus (LSV), Lily  mottle potyvirus (LmoV), Lily X potexvirus (LVX), Strawberry latent ringspot   nepovirus (SLRSV), Tulip breaking potyvirus (TBV)

 

» Rose:             Apple mosaic ilarvirus (ApMV), ArMV, PNRSV, SLRSV

 

» Gerbera: CMV

 

» Orchids: Cymbidium mosaic potexvirus (CymMV), Odontoglossum ringspot  tobamovirus (ORSV)

 

Electron Microscopy

EM studies with respect to particle morphology and inclusion bodies were carried out for 19 viruses:

 BYMV (gladiolus), BYMV (iris), CarMV, CMV (chrysanthemum), CMV (gladiolus), CMV (lily), CymMV, CNFV, CRSV, CVB, CVMV, DsMV, IMMV, ISMV, LMoV, LSV, ORSV, TAV and TBV

 

 

Fig 9 Electron microscopy of culture3 showing (a) particles of size 650nm (b) Clumping of virions with LSV antiserum (C) virus aggregation in cytoplasm as (V) cytoplasmicinvasion in chloroplast as (Cy I) (d) virus aggregation near nucleolus wall and pore formation (P). Bar =250nm

Immunosorbent Electron Microscopy for 15 viruses:
BYMV(gladiolus), BYMV (iris), CMV(chrysanthemum), CMV (gladiolus), CMV (lily), CymMV, CVB, DsMV, IMMV, ISMV, LMov, LSV, ORSV, TAV and TBV,