Henry Kuska wrote:

Cass, concerning your suggestion to publish an article on the subject in the
American Rose Annual: I assume that the U. Calif. Davis research is nearing
completion. I hope that they would then write such an article as they are
the "horse's mouth".

Henry, I can appreciate your reluctance to survey the literature when
such a report is expected shortly. However, the amount of accurate
information in a form readily accesssible to even well-informed
rosarians is quite limited. I would view a survey as a precursor to the
Davis report, which undoubtedly will include several provisos that
"more research is needed." That is the perpetual state of knowledge.

Additionally, there is an unsatisfying air of "Don't worry, be happy"
surrounding the occurrence of rose mosaic virus. Don't we all wonder if
we would need fewer fungicides and in certain parts of the country
suffer fewer winter losses if there were fewer virused plants foisted
on on the public? I was delighted this spring to see Jackson Perkins
offering guaranteed virus free plants....for close to $20 each.

I "expect" that they are preparing something for the conference covering
virus diseases of ornamental plants which is scheduled for 2004. (Often,
researchers try to present their results at conferences such as this one in
order to assure that the work gets maximum exposure among those working in
the field.)

Looking at this list of mosaic viri (see? kits are available)
http://www.dsmz.de/plvirus/elisa_o.pdf, it is no surprise that roses
suffer from mosaic virus. I've seen common weeds in my yard suddenly
show up with mosaic or variagation of the foliage.

I suspect that the 1 to 4 % contagion rate is low. My experience with
both budded new releases and with old roses is not reassuring, more on
the order of 25% than 1 %. That experience has bolstered my interest in
collecting and propagating own root, healthy old roses from very old
homes. I read your information about mosaic symptoms showing up as
early as the 1860's. Whatever its rate of transmission then, it could
only increase with modern transportation and production techniques.
Apart from different rates of contagion for different virii, it seems
obvious that different modes of transmission would have different rates
of success. Add that to different susceptibility of cultivars, and the
landscape gets complicated quickly. But "Don't worry be happy" seems
just simple minded and an abandonment of efforts to improve.

Transmission by pruning equipment is the most obvious concern of most
home gardeners. Not knowing (a) how easily transmissible the viruses
are and (b) how many roses suffer from them without showing symptoms,
may make all efforts futile. There is no way of assuring the
cleanliness of pruners even after washing, dipping and drying,
especially with little more more than strings of protein involved. That
doesn't mean I will abandon those efforts. Disposable pruners? No
pruning? Not likely.

Just my thoughts after reviewing your materials.

One of our daughters and her family was visiting for the week so I
have been busy being "grandpa", but I did have time during their naps
to search for literature evidence that related to what Tom Liggett
said about rose virus returning to heat treated roses. The first
thing is the following link
http://groups.google.com/groups?hl=e...395300.XAA2410
1%2540ladder01.news.aol.com%26rnum%3D2

That link states that it was stated at a meeting and that others heard
it also.
-------------------------------------------------------------------------------------------
The reference that appears to be the most interesting is:

Title: Recent Issues raised in the Evaluation of Virus Removal and
Inactivation

Authors: White, E.M., and Woodward, R.S.,
Published in Genetic Engineering News, volumn 15, page 6, (1995).
(Note, the page number is ambigious in the source that I found this
in.)

Unfortunately no abstract was given. Can anyone provide look this
reference up?
--------------------------------------------------------------------------------------------------
A number of German papers (by the same labatory plus other laboratory
co-authors) appear to say something which is related to this issue.

I purchased one of the most recent of their papers.

Title: Phytosanitary Improvement of Fruit Tree Species: Diagnostic
Strategies in Virus-Indexing of In Vitro Plants

Authors: A. da Camara Machado, D. MendonCa, M.S. Lopes, E. Knapp, V.
Hanzer, W. Arthofer, H. Katinger, M. Laimer da Camara Machado

Authors affiliation: this was a combined publication from a
laboratory in Portugal and a laboratory in Austria.

Published in: Acta Hort., volumn 472, pages 511-516, (1998).

In the actual paper they say that one of the points of the
investigation was: "2) Are virus elimination treatments merely
reducing the virus titer, but are they failing to eliminate some
viruses entirely. If so, how long does it take for those viruses to
reaccumulate to detectable levels?"

The above sounds like exactly what we are looking for (but not on
roses). In in their Results section, they say:
"Furthermore, we tried to understand whether, in some cases, virus
elimination treatments merely reduced the virus titer or failed to
eliminate some viruses entirely. If so, how long does it take for
those viruses to reaccumulate to detectable levels? Depending on this
situation, how many tests are necessary to confirm initial negative
test results? It is known that elimination treatments may depress the
pathogen titer below the threshold level of detection (Fridlund,
1989). In this respect the effects of thermotherapy on the titer of
ASGV and ACLSV were investigated in in-vitro-shoots of several apple
cultivars. Irrespective of the duration of thermotherapy treatment
(depending on heat sensitivity of the plant material), the virus titer
of both ASGV and ACLSV initially was decreased dramatically. Shoots
that were multiplied in vitro for over 2 years and used as starting
material. Before undergoing thermotherapy, they tested 100 % positive
for ACLSV, 100 % negative for ApMV and showed values around the
threshold level of 36% for ASGV, as shown in Fig. 1 for the Austrian
cultivar Maschanzker.
After thermotherapy meristems were excised and regenerating shoots
submitted to a multiplication step to increase the number of shoots.
After 7 months, plantlets were tested again and showed mainly negative
values or vahies around the threshold for ApMVand ACLSV, indicating
that the elimination success was satisfactory. On the other hand,
after 7 months 7% of the samples were postive when tested for ASGV,
indicating a different reaction pattern (Fig.
1)......................................
It is a definite fact that plants with a double infection of ASGV and
ACLSV are more difficult to- treat. However, we do not know, so far,
on which mechanism this synergistic effect may rely. Samples of the
different groups were compared (Fig. 2). From the nontreated positive
control plants, only ApMV was not detected after this period in vitro.
Values around the threshold were obtained from the originally negative
plantlets. In other cultivars, from originally negative clones
multiplied in vitro, a high number of samples showed ASGV positive
results after more than one year of in vitro culture.
-------------------------------------
The detection of ASGV by ITP in shoot tips from several potted plants
of Maschanzker grown for 2 years in the greenhouse (data not shown)
was, however, a concern. Therefore, the need for a more reliable
detection system seems evident."

The DISCUSSION section contained the following:

"As it is still common practice in sanitation programs to carry out
in-vitro treatment and ex-vitro re-testing for selection of plant
material. The result is a lack of knowledge of the speed of recovery
of low pathogen levels under in-vitro conditions (IPGRI/FAO,
1994)..........................
Elimination treatments depressed virus titer, as could be shown for a
wide range of cultivars. Thermotherapy, however, alters or destroys
viral proteins, therefore, serodiagnostics are of little value for
reliable early screening. Furthermore, there remain the limitations of
current ELISA-based serological tests which might be not sensitive
enough to detect very low levels of virus. Also, the time required by
the different viruses to recover up to a level of detection from low
levels of infection will again be dependent on the pathogenhost
combination. ACLSV was readily detected after re-accumulation above
the threshold level whereas the well-known problems in reliable
diagnosis of ASGV were further encountered even after several tests of
in-vitro cultures (Fuchs et at, 1988, Gilles and Verhoyen, 1992). We
assume that molecular diagnostics will improve the aforementioned
problems. Other methods which, so far, are not used for routine
diagnostics, like PCR or immuno-capture-PCR, will be introduced into
the sanitation program to improve the system of diagnosis for in-vitro
plants."
-----------------------------------------------------------------------------------------------------------
There were several Japanese papers which also appeared to be related
to our topic. The following is one:

Title: Evaluation of virus-free bulblet production by antiviral and or
heat treatment in in vitro scale cultures of Lilium longiflorum
'Georgia' and L-X 'Casablanca'

Authors: Xu PS, Niimi Y

Authors affiliation: Xu PS, Niigata Univ, Fac Agr, Ikarashi 2-8050,
Niigata 9502181, Japan

Published in: JOURNAL OF THE JAPANESE SOCIETY FOR HORTICULTURAL
SCIENCE, volumn 68, pages 640-647, (1999)

Abstract: "This study evaluated the effects of chemotherapy (addition
of ribavirin or 2-thiouracil to a medium) and/or heat treatment at 35
degrees C on the Production of virus-free bulblets in the scales
culture of Lilium longtiflorum 'Georgia' and L. x'Casablanca',
infected with lily symptomless virus (LSV), tulip breaking virus-lily
(TBV-L), and cucumber mosaic virus (CMV).

1. When scales were cultured on a medium with ribavirin at 50 mu M,
ELISA-absorbance values at 405 nm for LSV in the incubated scales
decreased as the incubation Lime lengthened, whereas the TBV-L, values
of the scales decreased at almost the same rate as those of the
control.

2. When scales of L. longiflorum 'Georgia' were cultured on medium
with antiviral chemicals, the number of bulblets formed decreased as
concentrations of antiviral chemicals increased. Viruses were detected
in about 20% of the bulblets at the end of culture when the scales
were cultured on medium with ribavirin and 2thiouracil at 50 mu M;
however, mo than 44% of the bulblets, which were transplanted into
soil and cultivated in the greenhouse for 6 months, showed the
positive reaction in viruses.

3. Scales excised from bulblets heat-treated at 35 degrees C for 4
weeks formed fewer bulblets than those of control, especially the
scales of 'Georgia'.

4. Chemotherapy in combination with thermotherapy was more effective
in decreasing the number of virusinfected bulblets than was the single
treatment. When scales, kept at 35 degrees C for 4 weeks, were
cultured on medium with 5 mu M ribavirin, viruses were detected in 30%
of the bulblets of 'Georgia' and 6% of those in 'Casablanca' at the
end of in vitro culture. However, viruses were detected in 100% of the
bulblets in 'Georgia' and 44% of those in 'Casablanca' which were
transplanted into sail and cultivated in the greenhouse for 6 months."

The Ohio State Research Laboratory at Wooster does subscribe to this
journal. However, the article is written in Japanese so I cannot add
to the abstract.
----------------------------------------------------------------------------------------------
My comment (Henry Kuska): it appears that, at least in some plant
systems, treated material may have a virus level below the detection
limit of ELISA, and that small amount of undetected virus may then
grow to detectable levels.
------------------------------------------------------------------------------------------------
..

The following is a paper which compares ELISA detection of PNRSV to
other methods:

Title: Comparative analysis of ELISA, nonradioactive molecular
hybridization and PCR for the detection of prunus necrotic ringspot
virus in herbaceous and Prunus hosts

Authors: Sanchez-Navarro, Apancio, Rowhani & Pallas

Published in: Plant Pathology, volumn 47, page 780, (1998).

Abstract: "Comparative analysis of ELISA, nonradioactive molecular
hybridization and PCR for the detection of prunus necrotic ringspot
virus in herbaceous and Prunus hosts
Three methods were compared for the detection of prunus necrotic
ringspot virus in herbaceous and woody plants: DAS-ELISA, nonisotopic
dot-blot hybridization and reverse transcriptional polymerase chain
reaction (RT-PCR). When purified virus preparations were used, the
detection limit of the RT-PCR technique was 1.28 pg mL-1 whereas
nonisotopic molecular hybridization and DAS-ELISA allowed detection of
0.8 ng mL-1 and 4 ng mL-1, respectively. Several sample processing
procedures were evaluated for virus detection by the nonisotopic
molecular hybridization technique. When a very short and simple sample
processing method was used, the detection limit of the nonisotopic
molecular hybridization technique was 25 times higher than that of
DAS-ELISA and 625 times lower than that of RT-PCR. A comparison of the
level of virus accumulation in mature fruits and in leaf tissue showed
that, on average, 125 times more virus was found in the fruits."