Can North American rose hybridizers safely use pollen from PNRSV infected roses

Can North American rose hybridizers safely use pollen from PNRSV infected roses?

By Henry Kuska

(An earlier version of this article was published in the Rose Hybridizers Association Newsletter, volume 34, no. 1, pages 7-10, (Spring 2003).)

This page gives the information that I have collected from my own literature searches and from others posting on the internet. Please let me know if you feel anything is not clear or is not addressed at all as I am continually updating/modifying it as I get feedback.

Bold print in quotes does not mean that the bold print appeared in the original; the bold print was added by me (H. Kuska) for emphasis.

In the 1990s I ran a (free) e-mail rose breeding scientific literature "course". One of the questions that I received was from a prominent North American rose hybridizer (of All American Rose caliber from a northern state). He stated that, from what he had read, rose mosaic virus could not be spread by pollen. However, he had occasionally observed virused roses among his seedlings. He asked if I knew of any scientific rose literature on the subject.

He is not the first hybridizer to report that some of his seedlings were virused. Harvey Davidson reported in 1988 that he had heard, in a recent lecture, Dr. Dennison Morey state that rose mosaic can be transferred through pollen. See footnote (1). Dr. Morey was at one time the head of the breeding program for Jackson and Perkins.

First, I will define my terms. There are a number of viruses that historically have been included under the broad term "rose mosaic virus". In this discussion, I will limit myself to what, at present, appears to be the most prevalent rose "mosaic" virus in North America, prunus necrotic ringspot virus (PNRSV). A more accurate scientific name is prunus necrotic ringspot ilarvirus, but I will use the more common prunus necrotic ringspot virus.

In the "Plant Viruses Online" database, the (general) modes of transmission for PNRSV are given as: "Transmitted by means not involving a vector. Virus transmitted by mechanical inoculation; transmitted by grafting; not transmitted by contact between plants; transmitted by seed (to over 80% in Prunus pennsylvanica but much less in peach); transmitted by pollen to the seed and transmitted by pollen to the pollinated plant."

The above is a general statement. It is possible (but not probable) that PNRSV could be transmitted in pollen of other infected plant species (to some non-zero degree) but not transmitted at all (zero) in roses. To see whether this remote possibility of zero transfer is actually the case, specific scientific studies of roses and PNRSV will be examined.

The earliest pertinent studies that I could find were four that were published in the early 1980s.

One was by D. J. Barbara who reported that in rose, high levels of PNRSV virus were present in petals and stamens but only very low levels of virus were present in the sepals. Note, for the non scientific reader, pollen is released from the stamens.

The second study appeared in a paper published by J. B Sweet. He reported that PNRSV was detected in the pollen of Peace and Queen Elizabeth roses, and in 1 % of two batches of seedling Rosa multiflora rootstocks. As is typical of scientific caution (or at least should be), he points out that finding the virus in the seedlings is not a definitive proof of transmission through the rose seed. However, his statement should not be interpreted that he did not find non grafted seedlings with virus; it means that he cannot be sure that the infected seedlings came from infected seeds (the virus may have infected the seedlings after they were planted (infected shears, infection from thrips, etc.)). What it does show is that there was transmission of PNRSV to non grafted seedlings in 2 different batches. Since multiflora seedlings are grown together in a field, I think that one can exclude the possibility that the seedlings were infected by "root grafts to an infected neighbor plant". If one attempted to use this reasoning, one would be going in a circle logically i.e. one would have to explain how the infected seedling that supplied the root graft got infected.

In a similar (but not a duplicate) paper in the Journal of Horticultural Science Sweet states (this is the exact quote from Sweet's paper (except for parts referring to other plants)): "PNRSV was sap-inoculated directly from the infected ..... Rosa spp. and from .... to herbaceous hosts inducing typical symptoms". Note, the use of the word "directly". This is an important finding because this removes the possibility that the virus is "too fragile" to be transferred by anything but a graft (living phloem cells of the plant). The "too fragile" theory seems to be one of the key assumptions of the "no transmission but by grafting" school of thought.

The fourth 1980s study was published by B. J. Thomas. He found that both the gel immunodiffusion test and the latex test were unable to detect PNRSV in infected roses but that both ELISA (enzyme-linked immunosorbent assay) and SSEM (serologically specific electron microscopy) methods were able to (in some cases ELISA also failed). SSEM was twice as sensitive as ELISA. Of particular importance here is that he was also able to detect the virus in the rose stamens.

The ability to transfer, mechanically, the virus through pollen to indicator plants was reported in 1994 by H. Baumgartnerova.

Unfortunately, nature often is not as simple to understand as we would like. This appears to be one of those times as there are 3 papers that looked for PNRSV transfer to seedlings but did not find it.

The first paper is Sweet’s Journal of Horticultural Science report referred to earlier. He germinated seeds from the two infected plants (36 seedlings from one and 24 seedlings from the other. None showed visual symptoms and 5 random samples from each were grafted on P. Persica GF 305 seedlings (virus test plants) and did not test positive. There are several reasons why I do not consider this report definitive: 1) 10 grafting tests is too small a sample size, 2) the use of visual symptoms on 36 samples can now be considered as both too small a sample and to also be deficient in test sensitivity (often infected plants do not have visual symptoms), and 3) if the infected two bushes were randomly positioned in the same field as 198 healthy ones; it is probable that the pollen came from the healthy bushes (this point will be discussed further later).

The second virus-seed transmission test paper was published in 1984, by B. J. Thomas. He reports on three different experiments that could provide some information regarding virus-seed transmission. In the first experiment he crushed 10 seeds from each laboratory infected virused plant with hips, and could not, using ISEM, detect any PNRSV in the seeds (he did not state the number of batches examined). In the second experiment he examined, with ISEM, 1067 seedlings grown from seeds harvested from the infected bushes and found no PNRSV infected plants. A possible explanation for the failure to detect PNRSV in the seeds or resulting seedlings is that the seeds may have also (like Sweet’s samples) been unintentionally produced from non-infected pollen. The roses utilized were all species roses, R. canina, R. canina, var. Brogs, R. corymbifera, R. multiflora, and R. rugosa. Species roses are normally self-sterile (see footnote 2). He should of been examining the seeds from the non virused bushes that were located between the two infected bushes or better yet did hand pollination with infected pollen..

The third paper appeared in 2007 and was published by the U. California Davis. They did not find any pollen/seed transmission, but they did not conduct any experiments where they used virused pollen to produce seedlings. They did examine open pollinated seedlings of virused mothers (page 218 of full paper "Ripe rose hips were harvested from the following seven varieties that tested positive for ApMV and/or PNRSV by ELISA: Rosa hybrida ‘Don Juan’, ‘Fourth of July’, ‘Red Fountain’, ‘Arizona’, ‘Queen Elizabeth’, and ‘Earth Song’.") In addition a hot summer climate is not the ideal place to study any type of above ground transmission as the rose's immune system has been found to be effective against PNRSV at higher temperatures. At high temperatures the virus concentration has been found experimentally mainly in the below ground parts of the plant. See the following link: http://home.roadrunner.com/~kuska/high_temperature_effect_on_pnrsv.htm

Virused (PNRSV) pollen has been found to be at a disadvantage if it has to compete with non virused pollen (infection with PNRSV decreases the germination percentage of pollen grains by more than half and delays the growth of pollen tubes by ≈24 h. ). Plus, an infected plant has some pollen that is not infected ("not all the apricot pollen grains from the same infected flower, or all the apricot flowers from the same infected tree, were infected.")

Thus, even if an infected plant is not self-sterile; the pollen from the nearest neighbor non-infected plant of the same species and non infected pollen from the infected plant itself would have the first opportunity to pollinate the infected bush (assuming that bees were around, which should be a reasonable assumption since all studies so far were done in open fields).

I conclude that the studies indicate that natural field infected pollen transmission in cooler climates will probably be low. But, for a cool climate hybridizer who shields the mother from foreign pollen the existing research cannot be used to exclude the possibility that some infected seedlings will be produced if one uses infected pollen.

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I would like to suggest what I consider to be a definitive experiment regarding PNRSV transfer from infected pollen through rose seed to seedlings: hand pollinate a statistically significant number of fertile non-infected roses (that had their own pollen removed) with diseased pollen from an infected rose (or several infected varieties) that is (are) known to cross with them. The hand pollinated flowers would then be covered to prevent stray pollen from contaminating the chosen flowers (the seedlings raised from the seeds resulting from these crosses would have to be kept in an insect free environment, and sanitary procedures utilized to prevent "after germination" contamination in order for the experiment to be considered truly definitive). The experiment would have to be carried out in a cool climate or in a temperature controlled greenhouse. The seedlings would then have to be tested by one of the sensitive methods as visible symptoms alone is not sufficient.

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Footnote (1) Davidson, H., The American Rose Magazine, volume 29, page 16, (1988).

Footnote (2) In 1986 P. Cole and B. Melton published a paper which investigated the ability of rose pollen to fertilize flowers on the same bush ( .Cole, P., Melton, B., J. Amer. Soc. Hort. Sci., volume 111, pages 122-125, (1986). The diploid species were all highly self-sterile. None of the 23 diploid specimens exhibited over 4 % fertility and 18 of the 23 produced no self-set seed. They also studied the fertility with pollen from another plant of the same species and found that the diploid group was 50 times more cross-compatible than self-compatible. For roses of higher polyploid level 12 of the 16 studied were no more self-compatible than the diploid group.

Related link: When was rose mosaic virus first observed?

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