Nov 9, 2004

Research Projects

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Transgenic fructan potatoes: Altered susceptibility to plant viruses?

(2001 – 2004) University of Hannover, Institute of Plant Diseases and Plant Protection

Topic

The aim of this research was to find out if fructan potatoes differ from non-GM potatoes in their susceptibility to important potato viruses.

It is quite common for plants to be infected with one or more different viruses. Tests in laboratories on plants coinfected with more than one virus have observed recombination of genetic material between different viruses. These recombinations can lead to the formation of new viruses. Although these events are considered very rare and are still only partially explained, they play a central role in viral evolutionary theory.

This project sought to:

• Get a better understanding of the formation of new potato viruses, their population dynamics , and their lineages in both conventional and transgenic potato varieties.
• Analyse possible recombination events in virus populations and to evaluate their consequences.

Summary

Potato virus Y (PVY), which comes in two natural forms: PVY-NTN and PVY-O, was the most commonly found virus throughout the course of the study. Molecular techniques were used to determine that most of the infected samples possessed the strain PVY-NTN. Dual infections with both types were observed but were rare.

The research showed that the transgenic fructan potatoes were no more susceptible to PVY than conventional varieties. Therefore, no increased risk of viral infection is expected.

The sequences of PVY isolated in the experiment were compared with data from the ‘GenBank’ sequence databank. No recombinations were identified.

Experiment description

The experiment encompassed studies on field-grown plants and experiments in the laboratory and in greenhouses.

Analysis of potato virus populations

The research focused particularly on Potato virus Y (PVY), Potato leaf roll virus (PLRV), Potato virus M (PVM), and Potato virus X (PVX). Their natural rate of occurrence among the transgenic potatoes was already known for the field trial site (Dahnsdorf). This data was compared with data on the susceptibility of conventional potatoes to these viruses. Extensive resources on potato virus infection rates were already available from official tests on propagation material. These data were based on visual inspections (phenotype evaluation). Random samples (leaf samples and tubers) were also taken back to the lab for ELISA testing to determine the degree of infection.

Identifying recombination events in transgenic and conventional plants (Laboratory tests)

To determine the extent of potential recombination events of PVY in transgenic and non-transgenic potatoes under field conditions, infected plants grown from tubers collected from the field trials were examined in the laboratory. Molecular techniques included sequences analysis were used to characterise virus isolates and identify recombinant virus particles.

Assessing the fitness of modified viruses (Laboratory test)

No recombinant viruses were found. If they had been found, their fitness would have been compared to the original viral strain. That would have involved determining the relative replication rates of each strains in instances of infection and of coinfection with both strains.

Results

Analysis of potato virus populations

All of the field trials performed to date have confirmed that PVY is the most commonly occurring viral infection. For this reason, PVY played a central in this research project.

Of the 1,200 total conventional and transgenic potato tubers planted in 2001, half became infected with viruses. 90 percent of the infected plants were infected with Potato virus Y (PVY), some of which were also infected with other viruses. The other viruses included in the tests occurred much less frequently.

Potato virus A (PVA) was not detected in any of the plants.

Tests on tubers from the 2002 field trials only detected PVY. A comprehensive test on 1,200 conventional and transgenic potato tubers was again performed in 2003. 37% of the samples tested positive for viral infection. Of these, all but four tubers were infected with PVY (one tuber was infected with PVY, and three were infected with PVS).

Determination of recombination events in non-transgenic and transgenic plants

So far possible recombination events have been investigated using the protein of various PVY types. From leaf tests and harvested tubers from the different field trials the RNA of the PVY virus was isolated. The sequence for the gene of the virus was multiplied and using molecular biological analysis the various isolates could be arranged by PVY type.

Of the samples tested so far around 70 percent can definitely be assigned to the PVY virus and around 22 percent to the PVY family. Just two percent of the samples tested showed a dual infection with PVY and PVY. In six percent the samples did not correspond exactly with PVY or PVY. These samples are currently undergoing further tests.

The sequence data from the various PVY types in these leaf tests were compared with known sequences from the databank “GenBank”. No recombinations have been found in the transgenic potatoes or in non-transgenic tests. Establishing the fitness of changed viruses

No altered viruses have yet been found in the field trials. In a laboratory experiment the protein sequence was exchanged in an infectious Potato virus X (PVX)-clone for the corresponding sequence from the Potato virus M (PVM). The result was repeatedly checked for its infectiveness. In none of the tests was infection achieved. Apparently a change in its protein leads to a loss of infectiveness in the Potato virus X (PVX).