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Papaya Ringspot Virus

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Papaya Ringspot
Kate Stokes and Andrew
• Papaya ringspot virus-type P
• Occurs as 2 strains
– Type P infects both papaya and cucurbits such
as squash, pumpkin, cucumber, and watermelon
– Type W infects watermelon only
General Characteristics
• Member of potyvirus family
• Long, flexous rod-shaped particles about
800-900 nm in length
• Non-enveloped filamentous virions
• Single-stranded linear RNA genome 12 kb
total, encapsulated by a coat protein
• 5’ terminus of RNA has VPg
• Yellowing and vein-clearing on young leaves
• Yellow mottling of the leaves, severe blistering
and leaf distortion
• Dark-green streaks and rings in the leafstalks
and stems
• Concentric rings and spots or C-shaped
markings, a darker green than the backgroundgreen fruit color
• Can affect vigor of fruit and trees and fruit
• Occurs in nearly every region where
papaya is grown except for Africa,
including Hawaii, Taiwan, Brazil,
Thailand, the Caribbean islands and
the Philippines.
• Particularly severe in Thailand,
Taiwan, the Philippines, and the
southern region China
• Carried from plant to plant by aphids
during feeding probes
• Not spread by other insects and does
not survive in soil or dead plant
• Also spread by movement of infected
papaya plants and cucurbit seedlings
• Not usually seed-transmitted but
there is one case from the Philippines
• Brazil
– 1969 PRSV-p found in two main
growing regions, Sao Paulo
and Rio de Janeiro
– By 1984 73% of the
industry had moved to
remote regions to evade
the virus
• Hawaii
– Severely affected the papaya industry in the 1950s.
Subsequently, the papaya industry was relocated to Puna
– In May 1992, PRSV was discovered in Puna, the area
where 95% of Hawaii’s papaya was being grown, and was
widespread throughout the growing region by 1995.
Production steadily dropped from 53 million pounds in
1992 to 26 million pounds in 1998.
Quarantine measures
Restricted distribution
Removal of infected plants (rouging)
– Used protective netting against aphid vectors
because the island was too small to effectively
isolate plantings by moving
• Tolerant or resistant cultivars
Cross Protection
• Phenomenon in which plants systematically
infected with mild strain of the virus are protected
against infection by a more virulent related strain
• Used to control
– Citrus tristeza virus
– Tobacco mosaic virus
– Zucchini yellow mosaic virus
• Early attempts failed, but 2 mild strains were
eventually isolated after a virulent strain PRSV
HA was treated with nitrous acid
– PRSV HA 5-1
– PRSV HA 6-1
Cross Protection
• Results
– Delay in severe effects of the virus
– No complete protection from the virus
• Not Widely Accepted
– Adverse effects of mild strain
– Requires extra cultural management
– Reluctance of farmers to infect their trees
Pathogen Derived
• Concept conceived in 1980s
• First demonstrated by Beachy’s group in
transgenic tobacco resistant to TMV
Coat Protein Mediated
Protection (CPMP)
• Inhibition of disassembly in initially infected cells
– Interferes with release of encapsidated RNA
– Re-encapsidation
– Overcome by inoculation with naked (+) RNA
• Coat protein may play a role in replication and expression
– Requires high level of expression
– Interacts with infection cycle
• Interference with spread of virus from cell to cell
– Blocks movement through vascular tissue
• Examples
– TMV and Alfalfa mosaic virus
RNA Mediated
• Post transcriptional gene silencing
– Homology dependant
– Base pairing between the sense RNA transcript of the
transgene and the negative strand of the viral RNA
– Antisense RNA produced from the transgene could pair
with the viral RNA transcript.
• Duplex RNA is target for degradation
• Base pairing could inhibit translation
• Examples
– PRSV, PVX and PVY, and all potyviruses
Development of
Transgenic Papaya
• Hawaiian papaya industry was in trouble
• Dennis Gonsalves at Cornell in collaboration with
Upjohn, scientists at the University of Hawaii and
John Sanford at Cornell try to develop transgenic
papaya resistant to PRSV
• Target was the coat protein gene of PRSV HA 5-1
– 97.7% identity to PRSVw from Florida
– Gene was engineered a chimeric protein
– 17 amino acids of CMV at the N-terminus
Transformation of
Embryogenic Tissue
• Used gene gun newly invented by John
• Tissue bombarded with
tungsten particles coated
with the engineered
Resistant Lines
• Inoculation tests conducted with transgenic
plants and PRSV HA
• Line 55-1 showed resistance in greenhouse
– female rather than hermaphrodite so progeny
could not be obtained
– Crosses with non-transgenic papaya fallowed
by screening provided R1 plants resistant to
• Experiments showed
– Resistance of transgenic papaya to PRSV was
not correlated protein expression
– R1 plants were highly resistant to Hawaiian
strains of PRSV
– Line 55-1 had variable levels of resistance to
non- Hawaiian strains
– Resistance due to RNA mediated mechanism
rather than by coat protein
Field Trials
• 1991 APHIS issued a permit for field trials of the
new transgenic plants
• First trails designed to asses resistance to
mechanical and aphid inoculations of PRSV
• Large Scale Tests
– Pros
• Industry desperate
• Line 55-1 performed well in previous trials
• Could be done at sufficiently isolated site
– Cons
• Spread of pollen to commercial plants
• Pilferage
• Resistant plants may become weeds
Trial Results
• 50% of the non-transgenic control plants
infected within four months
• 100% within seven months
• Improved performance of transgenic plants
• Assessments of taste, production, color,
size, and packing and shipping qualities
were positive for the new plants
– Heteroencapsidation – might lead to non-vectored virus
to become vector transmissible if co-infection
– Recombination – might lead to novel viruses
– Transgenic proteins considered pesticides
– Nutrition and vitamin content
– Presence of GUS and
– Presence of benzyl thiocynnate genes
• Beachy, R. N., Loesch-Fries, S., Tumer, N. E., 1990. Coat Protein- Mediated
Resistance Against Virus Infection. Annu. Review Phytopathol. 28:451-74
• Tumer, N. E., Kaniewski, W., Haley, L., Gehrke, L., Lodge, J. K., Sanders, P.,
1991. The second amino acid of alfalfa mosaic virus coat protein if critical for
coat protein-mediated protection. Proc. Natl. Acad. Sci. USA. Vol. 88 pp 23312335, March 1991
• Gonsalves, D., 1998. Control of Papay Ringspot Virus in Papaya: A Case
Study. Annu. Review Phytopathol. 36:415-37
• 2001. Chiang, C., Wang, J., Jan, F., Yeh, S., Gonsalves, D., Comparative
reactions of recombinant papaya ringspot viruses with chimeric coat genes and
wild type viruses on CP-transgenic papaya, Journal of general Virology. 82,
• res/ringSpot.htm
• 82/11/2827
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