RESEARCH REPORT

High temperature sanitation of styrofoam tobacco float trays by George Duncan, Bill Nesmith, Billy Tapp1

Introduction: The use of styrofoam trays for transplant production has become widespread in the tobacco belt. The re-use of un-sanitized tobacco trays is a potential for pathogen carry over and re-infection of the new crop. A good washing helps clean the trays of media and other debris but does not constitute a proven means of adequately disinfecting infested trays, even with additive chemicals. The better procedures for sanitizing trays after washing include steaming, methyl bromide gas and chlorine bleach. The quaternary ammonium chloride salts (Q-salts) alone have not proven as effective as the above methods for trays, but they are improvements over washing alone. (Ky Pest News, Jan. 26, 1998) Steaming has been the most effective treatment in past studies. Steam is used in many commercial applications but is not convenient and suitable for most farmers. The styrofoam trays are easily damaged by steam temperatures if the steam is not mixed with air and cooled to less than 180 deg. F. Methyl bromide is almost as effective as steam against certain pathogens but requires careful and safe use. It is favored by many growers because of its easy use. But the product is being phased out by EPA regulations and is not a long term solution for tray sanitation. Chlorine bleach is used widely for tray sanitation but success on the farm is highly variable because of the wide range of use patterns. Used properly, it has performed almost nearly equal to fumigation or steam in many studies, but it is difficult to use and many steps are required. It is rather effective when clean trays are dipped several times (within a few seconds) in a 10% solution; followed by stacking and enclosing with a clean tarp to keep them wet overnight with the bleaching solution. Afterwards, the bleach solution should be washed from the trays with clean water or a water plus Q-salt mix solution, followed by aeration, to eliminate the chlorine and salts. The need for an alternative and effective means of sanitizing styrofoam tobacco trays at the farm prompted a study to evaluate high temperature conditions for sanitizing used styrofoam tobacco trays. Since steam is not readily available at the farm and the cost of equipment to produce steam and control it seemed too expensive, an alternative concept envisioned stacking wetted trays in an insulated chamber and using a high temperature heat source to provide a temperature for a period of time to successfully sanitize the trays. Pathological studies over the years have generally shown that a temperature at least 160 °F for 30 minutes or longer is needed to sanitize materials reused in the greenhouse.

1 George Duncan, Bill Nesmith and Billy Tapp; Ext. Prof., Biosystems and Agr. Engr. Dept, Ext. Prof., Plant Pathology Dept., and Research Technician, Bio. & Ag. Engr. Dept., respectively, Coll. of Agric., Univ. of Ky., Lexington. Dec., 2003

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It is known that styrofoam cannot withstand high temperatures or it will deform and even melt and curl to a non usable state. Information from two tray fabricators reveal that around 186-190 °F is the beginning of a softening temperature for the styrofoam used in the trays. Objective: The objective was to evaluate the efficacy a high temperature method for sanitation of moistened styrofoam tobacco trays. Methods: The following time-temperature exposure treatments were established for this study based on other known conditions for controlling various biological pathogens: Temp., °F Treatment time, Min 175 (79.4°C 15 30 60 160 (71.1°C) 30 60 120 145 (62.8°C) 60 120 240 Separate treatments with steam and a clorox dip-plastic cover were used for comparisons. The effect of the high temperatures on tray stability and breaking strength was also evaluated. Experimental Procedures: For this study, a 4x4x8-ft insulated chamber with internal heating elements, fan circulation and temperature control was fabricated to hold and treat approximately 90 standard tobacco float trays, each approximately 14 x 27 x 3 inches (Fig. 1). The chamber was constructed of 2x2 wooden members with a one inch thickness of high temperature foil faced fiberboard insulation on the outside and a vapor barrier of six mil clear plastic on the inside. Electric heating elements (two 500 watt resistance strips, one with a manually operated variable voltage solid state controller) provided the heat source. A circulating fan (12 inch dia., 1/6 hp motor) forced air upward and outward past the heating strips, across the ceiling and along the side walls of the chamber. The stack of trays had approximately two inches of clearance from the sidewalls and approximately eight inches from the top fan blade. One-quarter inch thick by three-eights inch wide wooden spacer strips were placed between tray layers to allow air passage. The electric motor was positioned outside the insulated chamber with a long shaft extending through the insulated wall with the fan blade inside to protect the motor from the high heat. Even then, enough heat was conducted through the motor shaft to cause the thermal overload protection of the motor to activate during the latter stages of the higher temperature tests. Another fan blade was placed on the exposed motor shaft (a double shaft motor design) to provide cooling for the motor. Test tray specimens were thoroughly contaminated with some of the most prevalent tobacco field and float bed diseases and pathogens (Rhizoctinia, Pythium, black shank, black root rot and Fusarium) by contaminating the soil during the growing of a crop of transplants with these diseases and pathogens. In addition, two edges of each tray specimen were dipped into a muddy slurry made of field soil from a black shank, black root rot, fusarium wilt nursery to simulate field contamination of trays during

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transplanting operations. These procedures coated major portions of the tray samples with a very potent level of pathogens.

Fig. 1. Insulated chamber built for high temperature tray treatments.

Fig. 2. Treated specimen trays inserted in middle of stack with wooden strip spacers.

Four one-quarter size treated tray specimens were placed in the middle of the three-wide by three-long by ten-high stack of trays (Fig.2). These four quarter-tray segments represented four replications for each treatment. One-quarter inch thick by three-eights inch wide wooden spacer strips were used between all trays to provide a space for air and heat circulation. Notice that the heated air had to penetrate through the equivalent of one tray width on the sides, one tray length on the ends and four to five tray heights to reach and affect the test tray specimens.

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Thermocouples (28 gauge Cu-Cn) were placed at strategic locations in the chamber to monitor the dry bulb and wet bulb temperatures. The thermocouple locations were: room ambient, approximately three inches along the air flow from a heater element, a bottom corner of the chamber, taped against the bottom of the styrofoam tray specimen and a 0.010 inch thermocouple imbedded about 1/8 inch into the surface of a styrofoam test tray specimen. A 'wet bulb' thermocouple was covered with a snug wick located just above (and supplied by) a small water bottle at the bottom corner of the chamber. After the temperature-time treatment of the one-quarter size trays specimens, each was placed in a sterile bag, later filled with steamed growing media, transplanted with small tobacco seedlings (plug plant size) that had been started in a lab environmental chamber and the plants grown in individual pans filled with sterilized water in the greenhouse under close observation for disease symptoms. Every effort was made to avoid cross contamination of tray specimens because the experimental design included contaminated trays that received no sanitation treatment randomly interspersed with the contaminated trays within the growing environment. Two other comparison treatments included a bleach/fumigation treatment as described above and aerified-steam treatment (172-183 °F for 30 minutes). Results and Discussion: Disease symptoms did show rather quickly in the greenhouse grow-out trays for the lower temperature treatments. No treatment was effective at eliminating all diseases, but several treatments significantly reduced diseases compared to the untreated-contaminated controls. Treatments experiencing the least disease were those with the 175 °F treatments (Table 1). The 175 °F x 15 min. and 175 °F x 60 min. temp.-time exposures had an Infectious Disease Index2 very similar to the steam and bleach/fumigation treatments whereas the 175 °F x 30 min. temp.-time treatment had two replicates that showed detrimental effects of diseases, hence an elevated disease rating. The Infectious Disease Index (IDI) ratings with statistical difference ranges are summarized below. The treatment numbers (#) reference several color photos of the plant grow-out results on later pages. This was a very rigorous test by design. It proved the importance of tray sanitation as a necessary step in re-use of contaminated trays. (Any tray after one season of use and soil-water exposure could be considered 'contaminated' to some extent.) Growers may get by with less rigorous treatment regimes when their trays have less contamination and because they are also using a range of fungicides to assist in the disease control. But this study did re-affirm the value of steaming trays and proper bleach/fumigation as useable treatments for tray sanitation. In addition, it lead to the discovery of an additional approach using a high temperature method that may be more practical for many growers. One disappointment was that we were unable to arrange for a methyl-bromide fumigation of any of these trays for comparison in these studies.

2 Infectious Disease Index = involves weighted variables including: time of first disease, root rot severity, plant chlorosis and plant death. In this study, the worse-case events were scored as 30 and a disease-free event was scored as 0.

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Table 1. Treatments, time of exposure, Infectious Disease Index and significance groupings for the tests.

Treatment, °F x time, Min #1-145 x 30 No Treatment #7-145 x 60 #8-130 x 30 #12-145 x 30, No Spacers #4-145 x 240 #5-160 x 30 #11-145 x 120 #9-160 x 120 #6-175 x 30 #2-160 x 60 Steam Clorox #3-175 x 15 #10-175 x 60

IDI Index 27.8 a 26.0 a 21.5 ab 19.8 a- c 18.8 a- c 17.3 a- d 11.0 b- e 10.8 b- e 10.8 b- e 10.0 b- e 9.8 b- e 8.0 c-e 6.5 de 4.5 e 3.5 e

Selected graphs of the various temperatures for the duration of the tests are shown below (Fig. 3 and 4). For these tests, the lab room temperature (Rm. Amb.) was around 20-23 °C. The other temperature legends and locations are: "Top Htr." was the temperature of the forced circulation heated air exiting the heater strips at the top of the chamber, "Bott. Chmbr" was the temperature in a bottom corner of the chamber, "Dry Bulb" was the temperature adjacent to the wet bulb, "Wet Bulb" was a thermocouple with a snug wick suspended just above a small water bottle, "Tray Bott." was the temperature measured by a thermocouple taped against the bottom of the styrofoam tray specimen, "Probe" was the temperature measured by a 0.010 inch thermocouple imbedded about 1/8 inch into the surface of the styrofoam test tray specimen. The "Test Temp" shows the period the actual temperature level was maintained for the styrofoam test tray specimen in the center of the stack of 90 trays (three wide, three long and 10 deep). The following photos (Fig. 5-17) show results of the four replicated treatments of plant growth and disease manifestations for representative treatment conditions. Notice that a temperature of at least 175 deg. F 1/8 inch deep in the center tray surface for 60 minutes with "moist" trays was needed to give effective treatment of the trays and reduce pathogens comparable to or better than appropriate clorox dip or steam treatments. A 15 minute treatment showed similar effectiveness but may not be consistently acceptable as the 30 minute treatment showed some inadequate pathogen control as did the 160 deg. F treatment. Also realize there was an hour or more of time required for the temperature to build up to these sustained levels as shown by the graphs following. Also realize these trays were stacked with a 1/4 by 1/2 inch wood spacer between all trays to allow all surfaces to be readily exposed to the high temperature moist air. Whether tightly stacked trays will achieve equal effectiveness by high temperature moist treatments in the same time intervals has not been verified. However, one treatment at 130 deg. F for 30 minutes with 'no spacers' (stacked trays) had poor pathogen control.

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Tobacco Styrofoam Tray Treatment #1 - 30 Min @ 62.8C (145 °F)

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Fig. 3. Temperature-time graphs for the environmental conditions of the 62.8°C-30 minute test.

Tobacco Styrofoam Tray Treatment #10 - 60 Min @ 79.4C (175F)

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Fig. 4. Temperature-time graphs for the environmental conditions of the 79°C-60 minute test.

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Fig. 5. Grow-out of test trays on greenhouse bench showing effects of

uncontrolled disease and tray treatments.

Fig. 6. Treatment #2 - 160 °F, 60 min. Fig. 7. Treatment #3 - 175 °F, 15 min.

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Fig. 8. Treatment #4 - 145 °F, 240 min. Fig. 9. Treatment #5 - 160 °F, 30 min. Fig. 10. Treatment #6 - 175 °F, 30 min. Fig. 11. Treatment #7 - 145 °F, 30 min.

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Fig.12. Treatment #8 - 130 °F, 30 min. Fig. 13.Treatment #10 - 175 °F, 60 min. Fig. 14. Treatment #11 - 145 °F, 120 min. Fig. 15. Treatment #13 - Steam

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Fig. 16. Treatment #14 - 10% Clorox Fig. 17. Treatment #16 - Check Heat effect on tray dimensions: The dimensional stability of trays was measured by: 1) the physical length, width and height dimensions of the trays after treatment and 2) placing trays after treatment onto a smooth marble top lab bench and measuring the tray height at the corners and mid points of each side with a micrometer dial indicator. A one gallon plastic container filled with tap water to equal five pounds weight was set on the middle of each tray as a ballast for the height measurements. These data revealed no abnormal styrofoam tray deformations for the treatments of these tests. Heat effect on tray strength: The strength tests (breaking of trays) was accomplished by gently clamping approx. four inches of the end of a tray into a holder and using a lab testing machine to apply a force via a one inch wide bar at the middle of the tray with the top of the tray (open cell surface) in tension until rupture of the cells walls and failure occurred (Fig. 18). The failure occurred in the walls of the first cell space from the clamping position which was the point of greatest tensile stress, thus the point of failure. By positioning trays in this manner, the tray could be tested from each end giving two strength tests per tray. Fig. 18. Tray strength test.

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Data for new trays of different cell numbers and company sources (coded to protect identity) are shown below. The tray treatments exposed the trays to the same temperatures as the disease tests and for the longest time of the respective treatment temperature. Tray Treatment, Cell# & Co. Breaking Strength, lbs, avg. of 4 tests Untreated New 253, Co. A Avg.: 35.6 Std. Dev.: 2.2 253, Co. B 40.2 1.7 200, Co. A 43.5 2.7 200, Co. B 43.7 3.1 175 °F x 1 hr 253, Co. A Avg.: 36.0 Std. Dev.: 1.3 253, Co. B 38.2 2.8 200, Co. A 54.9 3.1 200, Co. B 47.2 1.9 160 °F x 2 hr 253, Co. A Avg.: 37.1 Std. Dev.: 2.0 253, Co. B 41.5 0.9 200, Co. A 48.7 5.4 200, Co. B 41.4 3.1 145 °F x 4 hr 253, Co. A Avg.: 35.5 Std. Dev.: 3.0 253, Co. B 41.2 1.5 200, Co. A 47.8 3.8 200, Co. B 44.4 0.5 These data results generally show the 200 cell trays to be stronger than the 253 cell trays but no noticeable effect of the temperature x time treatments on either size. Acknowledgements: We gratefully acknowledge and appreciate the financial support of the Council for Burley Tobacco and DIMON International for this study; to Ms. Shari Dutton for the valuable assistance and supervision in the plant growing and diagnostic tasks; to Burl Fannin for lab instrumentation assistance and to Mr. James Penman for the chamber construction.