Figure 1. Typical symptoms caused by a petunia isolate of TMV;
A. Flower color break on blue petunia cultivar used in these trials;
B. Mosaic pattern on petunia foliage.

The introduction of vegetatively propagated petunias into the landscape market in the early 1990s resulted in increased reports of virus infections, including those caused by Tobacco mosaic virus (TMV). Among more than 130 viruses known to infect petunias, TMV is the most commonly detected and costs the floriculture industry millions of dollars in losses each year.

Symptoms of TMV on petunia include mosaic patterns, flower color break, stunting and leaf curling and twisting (Figure 1). TMV has a very wide host range and is transmitted mechanically, often by greenhouse workers. TMV particles are extremely stable and can survive for months in plant debris and on surfaces

Avoiding TMV

During vegetative propagation, disinfection of contaminated tools is critical to prevent the spread of diseases. Most disinfectants for greenhouse use are labeled with a 10-minute contact time for disinfecting tools, which is not practical in most situations. Unfortunately, there are very few replicated trials that compare greenhouse disinfectants against viruses. The initial goal of our research is to determine which treatments are most effective to prevent the spread of TMV to healthy plants. Ideally, a disinfectant would be effective with a short contact time (less than one minute), which would enable growers to soak one tool while another is being used. In addition, an ideal disinfectant would be reliable, safe for workers, tools and plants, be inexpensive and easy to obtain, and affect a broad spectrum of plant pathogens. The disinfectants chosen for this study were based on preliminary studies, recommendations by industry professionals and/or widespread industry usage.

Based on preliminary studies in 2007, several products were omitted from the replicated trials in 2008 and some treatments were modified in an attempt to improve efficacy. In these trials, we used two vegetative petunia cultivars; a total of 1,148 six-week old liners (about 115 plants per disinfectant treatment).

Table 1. Percent infected petunias following disinfectant treatment of
TMV-contaminated razor blades. All treatments used a one-minute contact time
except GreenShield (three minutes). Mock treatment used clean razor blades.
The following treatments were freshly prepared: 20 percent non-fat dry milk (NFDM);
20 percent NFDM plus 0.1 percent Tween-20; 1 percent VirkonS; 1:10 household bleach;
GreenShield 1 tsp/qt for three minutes; GreenShield 2 tsp/qt; 3 percent tri-sodium
phosphate (TSP); 1:100 ZeroTol.

Our approach was to rigorously test each disinfectant in a series of replicated trials. Inoculum was freshly prepared by grinding TMV-infected petunia tissue in water. Razor blades were dipped in the sap for 30 seconds, rinsed briefly in water and then immersed in the disinfectant solution for one minute except GreenShield, which had a three-minute soak time.

Each blade was then used to cut the main stem of a healthy petunia liner. The test plants were grown in liner trays for 10 weeks, until positive control plants (water treatment) showed strong symptoms of virus infection. Each asymptomatic plant was then tested for TMV infection using a serological assay. A subset of the symptomatic plants were arbitrarily chosen and tested to confirm TMV infection. Following the assay, all asymptomatic plants were potted into 3-inch pots and maintained in the greenhouse for at least six weeks, until all TMV-positive plants developed symptoms, at which time plants were rated for TMV symptoms and a subset of plants was retested serologically for TMV infection.

Forty-seven percent of the positive control plants (a dip in infected sap followed by a one-minute water treatment) became infected based on symptoms and/or serological assay, whereas the negative control (mock) had zero percent infection (Table 1). None of the disinfectant treatments tested was completely effective, although all treatments reduced the percentage of TMV-infected plants (Table 1). Non-fat dry milk (NFDM) (0.9 percent infected), NFDM with a surfactant (1.7 percent infected), 10 percent bleach (1.7 percent infected), and Virkon S (1.7 percent infected) had the greatest effect on reducing TMV transmission. These four most effective treatments were chosen for trials that more closely simulated vegetative propagation.

In these trials, each blade was contaminated by cutting the stem of a TMV-infected petunia immediately before the one-minute disinfectant treatment. Each blade was then used to cut a single healthy petunia (100 plants per treatment). Based on these trials, only NFDM and bleach completely eliminated TMV transmission, although the other treatments also greatly reduced the percentage of infected plants compared to the water control

Summary

We have established a two-phase robust screening system that separates effective disinfectants from those with reduced efficacy. This study is the initial phase of a larger program to develop a research-based list of effective treatments (additional products or modified conditions) for greenhouse sanitation. Although our initial efforts are focused on minimizing TMV transmission, our future plans are to test the more effective products against other important greenhouse bacterial and fungal pathogens.