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2005 Plant Management Network.
Accepted for publication 22 November 2004. Published 5 January 2005.


First Report of Hibiscus latent Fort Pierce virus in New Mexico


Jean E. Allen, New Mexico State University, Department of Entomology, Plant Pathology, and Weed Science, Las Cruces 88003; Ivanka Kamenova and Scott Adkins, USDA-ARS, U.S. Horticultural Research Laboratory, Fort Pierce, FL 34945; and Stephen F. Hanson, New Mexico State University, Department of Entomology, Plant Pathology, and Weed Science, Las Cruces 88003


Corresponding author: Stephen F. Hanson. shanson@nmsu.edu


Allen, J. E., Kamenova, I., Adkins, S., and Hanson, S. F. 2005. First report of Hibiscus latent Fort Pierce virus in New Mexico. Online. Plant Health Progress doi:10.1094/PHP-2005-0105-01-HN.


Hibiscus spp. are common landscape and potted ornamental plants throughout the southern United States. Two new tobamovirus species have recently been isolated from Hibiscus rosa-sinensis plants with diffuse chlorotic spots and rings and an overall chlorotic mottle (1,4). One of these viruses was first identified in Florida, and it was named Hibiscus latent Fort Pierce virus (HLFPV) to reflect the location and host from which it was isolated (3). The other virus was first identified in Singapore and was named Hibiscus latent Singapore virus (HLSV) (4). During the summer of 2003, foliar symptoms including chlorotic spots and chlorotic mottling were observed on H. rosa-sinensis and H. syriacus plants in and around Las Cruces, NM (Fig. 1).


 




Fig. 1. Representative New Mexico Hibiscus spp. plants tested for Hibiscus latent Fort Pierce virus (HLFPV). (A) Virus-like symptoms including diffuse chlorotic spots and chlorotic mottle in a leaf from an H. rosa-sinensis plant that tested positive for HLFPV by TBIA. (B) Leaf from a Hibiscus rosa-sinensis plant that tested negative for HLFPV by TBIA. (C and D) H.syriacus plants that tested negative and positive, respectively, for HLFPV by TBIA. (E and F) Individual leaves from plants shown in C and D, respectively.

 

Fifty Hibiscus spp. plants including indoor potted plants, landscape plants, and local nursery stock were sampled from eight different locations, including three local nurseries. Twenty-eight of the 50 samples had virus-like symptoms, whereas the remaining samples were from randomly selected asymptomatic plants. Twenty-three of the 28 symptomatic plants had mild symptoms, including chlorotic spots (Fig. 1A), which were consistent with previous reports of HLFPV, whereas five of the plants had more severe symptoms such as leaf distortion, stunted growth, and a lack of flowering (Fig. 1C-F). Initial testing for HLFPV was by tissue blot immunoassay (TBIA) using IgG prepared to HLFPV virions as previously described (2). No cross reaction of this IgG with HLSV-infected hibiscus has previously been observed in double antibody sandwich enzyme-linked immunosorbent assays. TBIA identified 16 HLFPV-infected samples (Fig. 2), all of which came from plants with virus-like symptoms. HLFPV was not detected by TBIA in any of the 22 asymptomatic plants. Electron microscopic analysis of leaf dips from symptomatic leaves revealed rigid, rod-shaped particles with dimensions of ~15 nm in width and ~250 to 300 nm in length (Fig. 3), consistent with tobamovirus virions and supporting the HLFPV diagnosis by TBIA. Similar particles were not observed in leaf dips from asymptomatic leaves.


 

Fig. 2. Tissue blot immunoassay (TBIA) of hibiscus samples from New Mexico. TBIA was performed as previously described (2). Results from a typical blot testing ten independent Hibiscus rosa-sinensis samples are shown. Samples 2 and 3 were judged to be positive.

 

Fig. 3. Electron microscopic detection of tobamovirus-like particles in symptomatic hibiscus plants. Hibiscus leaves were ground in water and spotted onto EM grids. After drying, samples were stained with 2.5% aqueous uranyl acetate and examined in a Hitachi H7000 transmission electron microscope. Image shown was taken at 80,000x magnification, scale bar = 100 nm. Image shows tobamovirus-like rods that were typically seen in symptomatic leaves but not in extracts of asymptomatic leaves.


The presence of HLFPV was confirmed by amplification of the capsid protein gene by immunocapture reverse-transcription polymerase chain reaction (IC-RT-PCR) using previously described methods and primers specific for HLFPV (2). The expected size product (535 bp) was amplified from a TBIA-positive, symptomatic H. syriacus sample (Fig. 4, lane 5). Sequence analysis of the IC-RT-PCR DNA fragment revealed 100% nucleotide identity with the corresponding portion of the HLFPV capsid protein gene. This finding supports the identification of HLFPV in New Mexico, and distinguishes it from HLSV, which only shares 68% nucleotide identity with the HLFPV capsid protein gene (1). No DNA fragments were amplified by IC-RT-PCR from uninfected Nicotiana tabacum, TBIA negative H. rosa-sinensis and H. syriacus leaves (Fig. 4, lanes 1-4), or HLSV-infected H. rosa-sinensis leaves (data not shown).


 

Fig. 4. Detection of Hibsicus latent Fort Pierce virus (HLFPV) by immunocapture reverse-transcription polymerase chain reaction (IC-RT-PCR). Samples of uninfected Nicotiana tabacum leaves (lane 1), asymptomatic leaves of Hibiscus rosa-sinensis (lanes 2 and 3) and H. syriacus (lane 4), and a symptomatic leaf of H. syriacus were tested for HLFPV infection by IC-RT-PCR with specific primers, analyzed by gel electrophoresis in a 1% agarose gel and imaged with a Kodak Image Station 2000. Comparison with DNA markers included on the gel (not shown) was used to determine the size in base pairs of the PCR products. All asymptomatic leaves (lanes 2-4) tested negative for HLFPV by tissue blot immunoassay, whereas symptomatic leaves (lane 5) tested positive.

 

Symptoms alone are not reliable diagnostic indicators for HLFPV or HLSV infection of hibiscus because as a vegetatively propagated crop, hibiscus can accumulate multiple viruses over time. Based on surveys in Florida (1), it is possible (and perhaps even probable) that some of the plants sampled in the current work are co-infected with one or more additional viruses which complicate symptomatology. For this reason, we used multiple methods (TBIA, electron microscopy, IC-RT-PCR, and sequence analysis) to conclude that HLFPV, previously identified in Florida (1) and Thailand (Adkins and Chiemsombat, unpublished), is also present in multiple locations and environments in Las Cruces, New Mexico. Detection of HLFPV in numerous H. rosa-sinensis and H. syriacus samples in New Mexico suggests that it may be widely distributed in this state, as is the case in Florida (1) and Thailand (Adkins and Chiemsombat, unpublished). This represents the first report of HLFPV in the western United States. Movement of ornamental plants could increase the geographic distribution of HLFPV.


Acknowledgment

This work was supported in part by USDA grant #2004-06129 and the Fred C. Gloeckner Foundation, Inc.


Literature Cited

1. Adkins, S., Kamenova, I., Achor, D., and Lewandowski, D. J. 2003. Biological and molecular characterization of a novel tobamovirus with a unique host range. Plant Dis. 87:1190-1196.

2. Kamenova, I., and Adkins, S. 2004. Comparison of detection methods for a novel tobamovirus isolated from Florida hibiscus. Plant Dis. 88:34-40.

3. Kamenova, I., and Adkins, S. 2004. Transmission, in planta distribution, and management of Hibiscus latent Fort Pierce virus, a novel tobamovirus isolated from Florida hibiscus. Plant Dis. 88:674-679.

4. Srinivasan, K. G., Narendrakumar, R., and Wong, S. M. 2002. Hibiscus virus S is a new subgroup II tobamovirus: evidence from its unique coat protein and movement protein sequences. Arch. Virol. 147:1585-1598.