Historic, archived document Do not assume content reflects current scientific knowledge, policies, or practices. - UT iagrd, ) hs Bd, eae : es i ; ivi orthrtment of Agricultu e 1 fade est Forest and Range Experiment, Station DA Forest Service RESEARCH NOTE ie Fe Lee 4 = haat eee c cere ONT Ri LER Oa a rane : ncT1¢ war August 1970 (ws ee ee : ates Effect of Zectran on Microbial Activity in a Forest Soil by W. B. Bollen, Principal Microbiologist Use Pastieides Se FOLLOW THE LABEL K. C. Lu, Principal Microbiologist U.S. DEPARTMENT OF AGRICULTURE and R. F. Tarrant, Principal Soi! Scientist Tite publication reports research twolving pesticides. Te does not contain recommendations for their use, nor does tt mpl that the uses diocussed here have been registered. All neo of peocicides must be see by appropriate State and/ or Federal agencies before they can be recommended. CAUTION : Pesticides can be injurious to humane, domestic animals, decrable plante, and fio or other wildlife --1F they oe ea api ed properly. Use all pesticldes selectively and carefully. Follow recommended practices for the disposal of surplus pestictdes and peoticide containers. ABSTRACT When Zeetran or No. 2 fuel otl was applied to soil in concentrations far greater than those that could result from current operational application rates, netther material adversely affected sotl microbial activity. We conelude from these data that low-volume applications of Zectran as an insecticide for forest use poses no hazard to soil microbes. The insecticide Zectranl/ applied aerially at a nominal rate of 0.15 pound per acre was as effective as malathion or DDT applied at 1.0 pound per acre to control western spruce budworm (Chortstoneura ocetdentalis Freeman) in Montana field tests (Buffam, Meyer, and Miskus 1967). On pon- derosa pine foliage in the laboratory, Zectran proved more toxic than DDT against larvae of a pine tussock moth ( Dasychtra sp.) (Lyon, Brown, and Richmond 1967). The chemical nature of Zectran suggests it may be much less persist- ent in the forest ecosystem than DDT (Hammer 1962). Zectran levels in plant foliage decrease rapidly after application (Pieper and Miskus 1967). Thus, it is a promising chemical for use against forest tree defoliators previously con- trolled by aerial applications of DDT (Anonymous 1967). In determining the safety of chemicals proposed for use in the forest environment, we are interested in effects of the chemicals on soil microbes which influence soil fertility. Soil respiration, as indicated by carbon dioxide (CO5) evolution, is a good general index of the activity of microflora involved in organic matter decomposition (Casida 1968). This index was used in our laboratory experiments to measure effects of Zectran on the microflora found in a coastal Oregon forest soil. (See details in table 1 and ''Materials and Methods. "') Zectran applied to soil without tree litter or oil carrier had no apparent effect on CO, evolution (table-1). Tree litter applied to soil without Zectran or oil carrier also had no apparent effect (table 2). Further, there appeared to be no interaction between Zectran and tree litter (table 3). V/ The Dow Chemical Company trademark for 4-dimethylamino-3, 5- xylyl methylcarbamate. Mention of companies or products does not constitute an endorsement by the Forest Service or the U.S. Department of Agriculture. Table 1.--Effect of Zectran on evolution of CO for 70 days from Astoria stlty clay loam Total CO as carbon Wy), Soil treatment comparisons Difference— --------- Mi Lltgrams--------- 1. No treatment Natit vs. Zectran at 1 p.p.m. 107 4 2. No treatment Watal vs. Zectran at 10 p.p.m. 104 7 Be Ze Ctra alceslenp)s pills 107 vs. Zectran at 10 p.p.m. 104 3 1/ First two comparisons not included in tests of significance; difference in third comparison is not signi- ficant at the 95-percent level of probability. Table 2.--Effect of tree litter on evolution of CO. for 70 days from Astorta stlty clay loam 1/ Woah GOs Difference— as carbon Soil treatment comparisons 1. No treatment lil vs. alder litter 105 6 2. No treatment ital vs. conifer litter 122 iil 3. All treatments with alder litter 153 vs. all treatments with conifer litter 159 6 4. All treatments without litter 148 vs. all treatments with litter 156 8 il) — First cwo comparisons not included in tests of signifi- cance; differences in third and fourth comparisons are not significant at the 95-percent level of probability. Table 3.--Effect of Zectran tn combination with tree litter on evolution of CO, for 70 days from Astoria silty clay loam Total CO DEES ene ee! as carbon Soil treatment comparisons ----------- Mi Lltgrams----------- 1. Zectran at 1 p.p.m. + alder litter 110 vs. Zectran at 10 p.p.m. + alder litter 96 14 2. Zectran at 1 p.p.m. + conifer litter 100 vs. Zectran at 10 p.-p.m. + conifer litter 113 13 4/ Differences were not significant at the 95-percent level of probability. Comparisons based on means of all treatments combined (table 2) indicate that mean CO, evolution for all treatments combined is substantially higher than means of treatments which included only Zectran or tree litter. Thus, the oil carrier is indicated as having some role in increasing microbial respiration. The comparisons shown in table 4 demonstrate that the carrier indeed is responsible for a substantial increase in microbial respiration. There was no Significant difference in CO9 production between the rates of carrier used, but means of all treatments containing the carrier were signifi- cantly greater than means of all treatments that did not include the carrier. We next looked for interactions between Zectran and the carrier. The CO, index was not significantly different between Zectran in carrier at 1 part per million (p.p.m.) and at 10 p.p.m. (table 5). We do see, however, that when Zectran in carrier is compared with the carrier only, presence of Zectran apparently causes a significant decrease in COs evolution. While this decrease is significant relative to levels obtained with carrier only, the absolute value for CO, production is still substantially higher than values measured in the absence of any carrier. In our study of interactions between the carrier and tree litter (table 6), there was no significant difference between carrier rates of 1 and 10 p.p.m. in combination with conifer litter. However, when alder litter and carrier were together, the 10-p.p.m. carrier rate led to a highly significant reduction Table 4.--Effeet of acetone-fuel oil carrier on evolution of CO. for 70 days from Astoria stlty clay loam Total CO 1/ Soil treatment comparisons Difference— as carbon --------- Mi lligrams--------- 1. No treatment 2/ 111 vs. carrier at l-p.p.m. rate— 183 We 2. No treatment Wail vs. carrier at 10-p.p.m. rate 192 81 3. Carrier at l-p.p.m. rate 183 vs. carrier at l10-p.p.m. rate 192 9 4. All treatments without carrier 106 vs. all treatments with carrier 194 88x* iW — First two comparisons not included in tests of significance; difference marked ** is significant at the 99-percent level of probability. = Carrier rates of 1 and 10 p.p.m. are equal to those used when Zectran at 1 and 10 p.p.m. was dissolved in carrier. Table 5.--Effeet of Zectran in combination wtth acetone-fuel oil earrter on evolution of CO, for 70 days from Astoria stlty clay loam Total CO Ditrerencee Soil treatment comparisons as carbon --------- Mi lligrams---------- l. Zectran at 1 and 10 Dopel. + Carrier 169 vs. carrier only at l1- and 10-p.p.m. rates2/ 188 19* 2a LeCELAnwat al apap. Ms.) it CabrLer 167 vs. Zectran at 10) p.p.m. -- carrier 170 3 iy) Difference marked * is significant at the 95-percent level of probability. 2 Carrier rates of 1 and 10 p.p.m. are equal to those used when Zectran at 1 and 10 p.p.m. was dissolved in carrier. Table 6.--Effect of acetone-fuel otl carrier in combination with tree lttter on evolution of CO. for 70 days from Astorta stlty clay loam Total CO as carbon 1/ Soil treatment comparisons Difference— ---------- Milligrams---------- 1. Carrier at 1l-p.p.m. rate + conifer litter 187 vs. carrier at 10-p.p.m. rate + conifer litter NS)7/ 10 ‘ 2/ 2. Carrier at l-p.p.m. rate + alder litter 201 vs. carrier at 10-p.p.m. rate + alder litter 170 31%** 1/ — Difference marked ** is significant at the 99-percent level of probability. 2/ — Carrier rates of 1 and 10 p.p.m. are equal to those used when Zectran at 1 and 10 p.p.m. was dissolved in carrier. in CO, evolution. Red alder litter has a high content of phenolic compounds, some of which are inhibitory to micro-organisms (Li et al. 1969). We hypoth- esize that the higher rate of acetone-fuel oil carrier released phenolic com- pounds from the alder litter and that some inhibition of microbial activity thus resulted. Again, the reduction we noted as a result of this interaction appears not to be substantial in comparison with COs evolution rates from all treatments that did not contain the carrier. When we looked for interactions between all three treatments used in this study--Zectran, litter, and carrier--we found no significant difference between Zectran in carrier at 1 p.p.m. versus 10 p.p.m. in the presence either of alder or conifer litter (table 7). Finally, in determining the relative effect of acetone and of fuel oil in the carrier mixture, we found no significant differences between untreated controls and fuel oil at either the 1- or 10-p.p.m. rate; nor was there any difference between fuel oil alone at 1- versus 10-p.p.m. rates (table 8). In contrast, presence of acetone at the 1-p.p.m. rate significantly increased CO. evolution over that measured from untreated soil. Acetone at the 10-p.p.m. rate also significantly increased CO» evolution, but the absolute amount of increase was substantially less than that noted with the 1-p.p.m. Table 7.--Effect of Zeetran, tree litter, and acetone-fuel oil carrier on evolution of CO for 70 days from Astoria silty clay loam Total CO» DiPeerencen as carbon Soil treatment comparisons 1. Zectran at 1 p.p.m. + alder litter + carrier 187 vs. Zectran at 10 p.-p.m. + alder litter + carrier 202 15 2. Zectran at 1 p.p.m. + conifer litter + carrier 200 vs. Zectran at 10 p.p.m. + conifer litter + carrier 193 U 1/ — Differences not significant at 95-percent level of probability. Table 8.--Hffect of fuel otl and of acetone on evolution of CO» for 70 days from Astoria silty clay loamb Total C05 Difference?! as carbon 2 - 2 Soil treatment comparisons— --------- Mi Lltgrams--------- 1. Soil alone 81 vs. fuel oil at 1l-p.p.m. rate 78 3 2. Soil alone 81 vs. fuel oil at 10-p.p.m. rate 84 3 3. Fuel oil at l-p.p.m. rate 78 vs. fuel oil at 10-p.p.m. rate 84 6 4. Soil alone 81 vs. acetone at 1l-p.p.m. rate 166 85% 5. Soil alone 91 vs. acetone at 10-p.p.m. rate 121 40% 6. Acetone at 1l-p.p.m. rate 166 vs. acetone at 10-p.p.m. rate 121 45% 1/ — These data are from a separate study. Soil age after col- lection and environmental temperature were different from that in the main study, accounting for the generally lower absolute values for total CO» as carbon. 2/ Rates are equal to those used for acetone-oil carrier in Zectran treatments at 1 and 10 p.p.m.. 3/ Differences marked * are significant at the 95-percent level of probability. rate. We conclude from these comparisons that acetone, not fuel oil, was responsible for increases in CO, evolution noted whenever the acetone-fuel oil carrier was included in treatments. Thus, when Zectran or No. 2 fuel oil was applied to soil in concentra- tions far greater than those that could result from current operational appli- cation rates (Buffam, Meyer, and Miskus 1967), neither material adversely affected soil microbial activity. We conclude from these data that low-volume applications of Zectran as an insecticide for forest use poses no hazard to soil microbes. MATERIALS AND METHODS In general, an Astoria silty clay loam soil was subjected to different treatments which included Zectran, forest tree litter, and an acetone-oil carrier mixture (table 9). Treated soil was incubated for 70 days, and microbial activity was determined from the amount of CO»9 evolved. A bulk sample of the All horizon of Astoria silty clay loam was taken beneath a conifer stand--Douglas-fir (Pseudotsuga menatesit), western hemlock (Tsuga heterophylla), and Sitka spruce (Picea sitchensts)--at the Cascade Head Experimental Forest near Otis, Oreg. A detailed list of chemical and physical properties of this soil is included in Franklin et al. (1968). For the soil used in the present study, organic matter was 28 percent, total carbon by dry combustion was 16 percent, and Kjeldahl nitrogen was 0.41 percent. Fifty-gram portions of sieved (10-mesh) soil, ovendry basis, were transferred into 250-milliliter widemouth Erlenmeyer flasks. Each treatment (table 9) was replicated three times. In treatments which contained forest litter or Zectran powder, these additives were mixed with the soil before it was transferred to the flasks. Distilled water was then added in sufficient amounts to bring moisture content of the treated soil to 50 percent of water- holding capacity (81 percent, dry weight basis). For treatments which included Zectran in an oil carrier, a stock solu- tion was prepared by dissolving 2 grams of the insecticide in 20 milliliters acetone, then adding 80 milliliters No. 2 fuel oil. Subsequent dilutions were made so that 1 milliliter additions of the solution to 50 grams of soil would yield 1 or10p.p.m. actual Zectran. Treatments which included the carrier alone or carrier in combination with litter were in amounts of the carrier equivalent to those in which Zectran had been included. The tree litter used was collected from the Cascade Head Experimental Forest near where the soil was collected for the study. Litter was air dried 8 Table 9.--Desertption of treatments to Astoria stlty clay loam and total CO. evolved over 70 days of tneubation Total CO f 1/ Soil treatment— as carbon Milligrams None--control 111 Z1 107 Zl, conifer 100 Z1, alder 110 Z10 104 Z10, conifer 113 Z10, alder 96 Conifer 122 Alder 105 “AILS coplal 167 Zl, oil, conifer 200 Zl, ofl, alder 187 Z10, oil 170 Z10, oil, conifer 193 ZO; ofl, alder 202 0i1(1) 183 0i1(10) 192 0i1(1), conifer 187 0i1(10), conifer 197 Oi1(1), alder 201 0i1(10), alder 170 1/ — 2Z1 or Z10 = Zectran at 1 or 10 p.p.m.; conifer or alder = type of forest litter; oil(1) or oil(10) = oil without Zectran but in same amount as used when Zectran in oil carrier was at 1 or 10 p.p.m. Oil carrier was a 1:4 mixture of ace- tone and No. 2 fuel oil. at room temperature and ground to pass through a 60-mesh sieve. The alder litter contained 51.74 percent carbon and 2. 31 percent nitrogen; the conifer litter, 51.85 percent carbon and 0. 84 percent nitrogen. Each flask containing soil and additives was connected to a COp-free, water-saturated air supply. The exit connection from the flask was inserted into a tube containing 1N NaOH to absorb CO, evolved from the soil by micro- bial activity. The entire respiration setup was incubated at 259 + 1° C. for 70 days. The tubes of NaOH were replaced at intervals of 1, 4, 8, 14, 28, 42, and 70 days. Amount of CO, absorbed by the NaOH was determined by double titration on a Beckman automatic titrator. Details of the apparatus and methods are given in Bollen and Lu (1961) and Bollen and Glennie (1961). Resulting data were subjected to two analyses of variance in which treatment degrees of freedom were partitioned into an orthogonal set of com- parisons (tables 1-8). LITERATURE CITED Anonymous. 1967. Zectran--for control of spruce budworm. Pacific Southwest Forest & Range Exp. Sta. USDA Res. Fact Sheet, 6 pp. Bollen, W. B., and Glennie, D. W. 1961. Sawdust, bark, and other wood wastes for soil conditioning and mulching. Forest Prod. J. 11: 38-46, illus. and Iu; Ke CG: 1961. Microbial decomposition and nitrogen availability of reacted sawdust, bagasse, and coffee grounds. J. Agr. Food Chem. 9: 9-15. Buffam, Paul E., Meyer, Hubert E., and Miskus, Raymond P. 1967. Small scale trials of five insecticides sprayed on spruce budworm in Montana. Pacific Southwest Forest & Range Exp. Sta. USDA Forest Serv. Res. Note PSW-159, 4 pp. Casida,, Was Handi. 1968. Methods for the isolation and estimation of activity of soil bacteria. In The ecology of soil bacteria, T. R. G. Gray and D. Parkinson (eds.), pp. 97-122. Liverpool Univ. Franklin, Jerry F., Dyrness, C. T., Moore, Duane G., and Tarrant, Robert F. 1968. Chemical soil properties under coastal Oregon stands of alder and conifers. In Biology of alder, J. M. Trappe, J. F. Franklin, R. F. Tarrant, and G. M. Hansen (eds.). Northwest Sci. Ass. Fortieth Annu. Meeting, Symp. Proc. 1967, pp. 157-172. Hammer, O. H. 1962. Zectran pesticide--some chemical and physical properties and some results from use on pests of ornamental plants. Down to Earth 17: 9-18. Li, C. Y., Lu, K. C.; Nelson, Eo E., and others. 1969. Effect of phenolic and other compounds on growth of Porta wetrtt in vitro. Microbios 3: 305-311. 10 Lyon, Robert L., Brown, Sylvia J., and Richmond, Charles E. 1967. Insecticides tested on new tussock moth--defoliator found in Montana. Pacific Southwest Forest & Range Exp. Sta. USDA Forest Serv. Res. Note PSW-161, 4 pp. Pieper, G. R., and Miskus, R. P. 1967. Determination of Zectran residues in aerial forest spraying. J. Agr. Food Chem. 5: 915-916. 11 The mission of the PACIFIC NORTHWEST FOREST AND RANGE EXPERIMENT STATION is to provide the knowledge, technology, and alternatives for present and future protection, management, and use of forest, range, and related environments. Within this overall mission, the Station conducts and stimulates research to facilitate and to accelerate progress toward the following goals: 1. Providing safe and efficient technology for inventory, protection, and use of resources. 2. Development and evaluation of alternative methods and levels of resource management. 3. Achievement of optimum sustained resource produc- tivity consistent with maintaining a high quality forest environment. The area of research encompasses Oregon, Washington, Alaska, and, in some cases, California, Hawaii, the Western States, and the Nation. Results of the research will be made available promptly. Project headquarters are at: College, Alaska Portland, Oregon Juneau, Alaska Roseburg, Oregon Bend, Oregon Olympia, Washington Corvallis, Oregon Seattle, Washington La Grande, Oregon Wenatchee, Washington