Elsevier

Scientia Horticulturae

Volume 106, Issue 2, 1 September 2005, Pages 213-227
Scientia Horticulturae

Identification, detection and frequency of lily viruses in Northern India

https://doi.org/10.1016/j.scienta.2005.03.011Get rights and content

Abstract

In a survey of lily growing fields in various parts of Himachal Pradesh, India, three viruses, Cucumber mosaic (CMV), Lily symptomless (LSV) and Lily mottle virus (LMoV) lily strain, were found quite prevalent in Asiatic (12 cultivars) and Oriental hybrids (4 cultivars) of lily, Lilium longiflorum and L. tigrinum. Apart from these viruses, Strawberry latent ringspot virus (SLRSV) was also found to infect Oriental hybrid lily. The disease incidence on the basis of symptoms observed ranged between minimum (40.7%) in cv. Alaska and maximum (83.7%) in cv. America, both Asiatic hybrid lily types. These viruses were indexed by testing outer and inner scales of bulbs and the leaves at early and flowering stages using ELISA and reverse transcription polymerase chain reaction (RT-PCR). Mechanical and insect transmission, purification, electron microscopy and sequencing of the PCR fragments were carried out for these viruses. Cloning and sequence analysis confirmed the viruses as CMV-subgroup II, LSV and LMoV. There were 96–98% nucleotide and 93–97% amino acid homology with CMV subgroup II sequences, 97–99% nucleotide and 91–92% amino acid homology with LSV sequences and 92–99% nucleotide and 85–98% amino acid homology with LMoV sequences.

Introduction

Lilies are in demand in the floriculture industry both as cut and potted flowers and ranked within the top 10 flowers in the export market. The genus Lilium includes 294 genera with 4500 species. The three commercially important divisions of lily include Easter lily (Lilium longiflorum), Asiatic and Oriental hybrids. In India, Asiatic and Oriental lilies are commonly grown in hilly areas and in mild climates. However, Easter lilies are grown with success in many parts of the country with tropical climate. Palampur is situated at latitude 32°6′N, longitude 76°5′E and altitude 1290 m, in district Kangra of Himachal Pradesh. It has a moderate climate area with temperature range of 10–30 °C. It receives 250 cm rainfall per year. The area is optimal for growing lilies commercially. Lilies can be grown from seed, bulb, scales, bulbets and bulbils. The main limiting factor in cultivation of lilies is susceptibility to virus diseases. Almost all lilies are propagated vegetatively, and infected bulbs used for forcing propagate the virus from one generation to another. These viruses affect the quality and yield of cut flowers resulting in great economic losses (Lawson, 1981, Raju and Olsen, 1985).

The most commonly occurring viruses of lily are Cucumber mosaic (CMV), Lily mottle (LMoV), Lily symptomless (LSV) and Lily X viruses (LVX) (Derks, 1995). In addition, Arabis mosaic (ArMV) (Mowat and Stefanac, 1974), Tobacco ringspot (Travis and Brierley, 1957), Tobacco rattle (McWhorter and Allenm, 1964), Tobacco mosaic, Lily virus X, Lily mild mosaic, Tomato ringspot, Narcissus mosaic (Bellardi et al., 1988, Stone, 1980, Memelink et al., 1990, Lee et al., 1996) and Strawberry latent ringspot viruses (SLRSV) (Cohen et al., 1995) have also been reported infecting lily. Ram et al. (1999) reported CMV infecting Asiatic hybrid lily cultivars on the basis of double stranded RNA (ds RNA) analysis, ELISA and host range studies.

Various lily cultivars grown in Himachal Pradesh, India, were collected and planted in the field of IHBT, Palampur, District Kangra. These plants were found to exhibit various types of symptoms, viz. mosaic, mottle, deformed leaves, deformed flowers, stunting of the plants and deformed bulbs.

The viruses were detected and characterized by bioassay, double antibody sandwich enzyme linked immunosorbent assay (DAS-ELISA), electron microscopy, immunosorbent electron microscopy, cytopathology, reverse transcription polymerase chain reaction (RT-PCR) and sequencing. Nucleotide sequences of Indian isolates were compared with other viruses reported from different parts of the world. In this way, the homologies were determined and nucleotide sequences of coat protein (cp) gene of viruses infecting lilies are presented.

Section snippets

Sample collection and establishment of virus(es) culture

The bulbs from infected plants showing symptoms were collected and planted in formalin sterilized plastic pots containing sterilized soil mixture (1 compost:2 sand:1 soil). The pots were kept in an insect proof polyhouse. The new leaves emerging from the infected bulbs were used for further analysis. Sap obtained by macerating the leaves in 0.1 M phosphate buffer, pH 7.2, followed by squeezing through double layered muslin cloth, was inoculated to Chenopodium amaranticolor, C. quinoa, Cucumis

Natural symptoms and disease incidence

Naturally infected plants of lily were found to contain either single virus or virus mixtures. When the plants were infected with a single virus, the symptoms were less pronounced and only mosaic, mottling and vein clearing was observed. In mixed infections, the plants showed severe mosaic, leaf deformation and stunting of the plants. Disease incidence on the basis of symptoms ranged from 83.7% in Asiatic hybrid cv. America to 40.7% and 40.9% in cvs. Alaska and Cavi, respectively (Table 2).

Discussion

Viruses are the major problem in lily growing fields. They often decrease the yield and quality of flowers. Lily growing fields in Himachal Pradesh, India, were found to have a high percentage of virus-infected plants. In our survey 16 cultivars of Asiatic and Oriental hybrids, L. longifolium and L. tigrinum were found to be infected with the virus diseases. The highest disease incidence on the basis of symptoms 83.7% was recorded in Asiatic hybrid cv. America and the lowest 40.7% was found in

Acknowledgements

We express gratitude to the Director and Mr. P. Navaghare, Institute of Himalayan Bioresource Technology, Palampur, for facilities and technical help, respectively. Financial grant (No. BT/PR/1421/SPD/11/099/98) from Department of Biotechnology, Govt. of India, for this study is gratefully acknowledged.

References (32)

  • A.F.L.M. Derks et al.

    Symptomen en andre schade door lelievirus X in lelies. LVX veroorzaakt minder schade dan LSV

    Vakwerk

    (1990)
  • A.F.L.M. Derks et al.

    Purification of Tulip breaking virus and production of antisera for use in ELISA

    Neth. J. Plant Pathol.

    (1982)
  • G.D. Foster

    Carlavirus isolation and RNA extraction

  • I.D. Garg et al.

    Morphological changes in flexuous potato viruses upon decoration in immunosorbent electron microscopy

    Acta Virol.

    (1993)
  • J.Y. Kim et al.

    Occurrence of lily viruses and damage of viral diseases in Oriental lilies

    RDA J. Crop Prot.

    (1998)
  • S.A. Langeveld et al.

    Identification of potyviruses using the polymerase chain reaction with degenerate primers

    J. Gen. Virol.

    (1991)
  • Cited by (45)

    • Simultaneous detection of three lily viruses using Triplex IC-RT-PCR

      2017, Journal of Virological Methods
      Citation Excerpt :

      Unfortunately lilies are often infected with viral diseases causing degeneration in the quality of many elite varieties. These infections can cause severe economic losses in terms of quantity and quality of flower and bulb production (Sharma et al., 2005; Wang et al., 2010a). More than ten different viruses have been reported to infect lilies worldwide(Ryu et al., 2002).

    • Rapid and sensitive detection of Lily symptomless virus by reverse transcription loop-mediated isothermal amplification

      2016, Journal of Virological Methods
      Citation Excerpt :

      As such, it is necessary to develop a rapid, sensitive, and specific detection method to prevent the spread of LSV. Several methods have been reported for LSV detection, such as real-time reverse transcription (RT-)PCR (Wei et al., 2012), quantitative RT-PCR (Sun et al., 2014), RT-PCR combined with enzyme-linked immunosorbent assay (ELISA) (Sharma et al., 2005), immunocapture RT-PCR combined with indirect ELISA (Kim et al., 2012), microsphere-based fluorescent immunoassay (Zou et al., 2011), and dot- or tissue-blot immunoenzyme assay (Makkouk et al., 1993). However, PCR-based detection methods and microsphere-based fluorescent immunoassay are time-consuming and require expensive instruments and skilled technicians.

    • Purification and immuno-gold labeling of lily mottle virus from lily leaves

      2016, Journal of Virological Methods
      Citation Excerpt :

      Symptoms may also be very mild, or plants maybe symptomless during early growth stages. Enzyme-linked immunosorbent assay (ELISA; Sharma et al., 2005; Tong et al., 2010) is commonly used to detect LMoV in lily. However, the method is time consuming, and requires technical expertise and specialized laboratory equipment.

    • Production of virus-free plants of Lilium spp. from bulbs obtained in vitro and ex vitro

      2015, Scientia Horticulturae
      Citation Excerpt :

      The variation between genotypes in the percentage of plant regeneration from meristematic tips recorded in the present study is in agreement with previous research (Cai et al., 1988) in which the survival rate of meristematic tips cultured in vitro was highly dependent on the genotype and varied from 42% to 88% in Dahlia spp. Previous studies showed that the RT-PCR reaction in adult plants offers higher sensitivity than DAS-ELISA, thus allowing detection of viruses in serology-negative plants (Niimi et al., 2003; Sato et al., 2002; Sharma et al., 2005). Moreover, previous research that compared RT-PCR with DAS-ELISA after a treatment for LSV elimination from Lilium (Nesi et al., 2009) reported similar results to those collected in the present study, i.e. DAS-ELISA negative plants were also negative when analyzed by RT-PCR.

    • Development of an immunochromatographic strip test for rapid detection of lily symptomless virus

      2015, Journal of Virological Methods
      Citation Excerpt :

      A special cellulose-based pad will be suitable for this purpose. Currently, laboratory detection of LSV is based on various methods including ELISA (Sharma et al., 2005), RT-PCR (Niimi et al., 2003; Zhang et al., 2010) and real-time PCR (Nesi et al., 2013). The sensitivity of a real-time PCR assay was found to be greater than that of the RT-PCR for the detection of LSV (Nesi et al., 2013).

    View all citing articles on Scopus

    Gene sequences for Cucumber mosaic virus (CMV), Lily symptomless virus (LSV) and Lily mottle virus (LMoV) of this study have been submitted to EMBL Data base with accession numbers AJ585086, AJ585052 and AJ549932, respectively.

    View full text