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Funded Project

Funding Program: IPM Partnership Grants

Project Title: Trial of a Minimum-risk Botanical Compound to Control the Vector Tick of Lyme Disease

Project Director (PD): Robert P. Smith [1]

Lead State: ME Lead Organization: Maine Medical Center

Undesignated Funding: $41,000

Start Date: May-01-2009 End Date: Apr-30-2011

Pests Involved: Ixodes scapularis Dermacentor variabilis, ticks

Area of Emphasis: botanical control, biological control, biocontrol, public health, community

Summary: Lyme disease is a bacterial infection transmitted in the northeastern United States by the deer tick Ixodes scapularis. Over 20,000 cases are reported to US Centers for Disease Control annually. In northern New England, three deer tick-borne diseases now infect people, dogs and farm animals. The use of synthetic pesticides to control these medically important arthropods, while effective, remains controversial due to non-target impact, health risk, and persistence. Recently, certain minimal risk, botanically-derived compounds have gained interest for inclusion in IPM strategies to control deer ticks. Based on positive results from a preliminary project, we propose to test the efficacy of a rosemary oil-based insecticide, Eco-Exempt IC2 (IC2), to control all stages of the deer tick in southern Maine. Working with licensed applicators, we will record the abundance of nymphal and adult I. scapularis ticks before and following applications of IC2, bifenthrin (a widely used synthetic pyrethroid), and water during the peaks of their seasonal abundance. Multiple study grids will be established within triplicate 70m X 70m grids in forested, infested habitat. In addition, we will examine the compound's effect on all of the tick's life stages in their environment by exposing them to treated leaf litter within enclosures. The effects of IC2 on non-target organisms, including pollinators will also be examined by plot count surveys and pitfall traps. Ancillary studies will examine the persistence of IC2 when sprayed at the beginning of the deer tick nymphal season, and its effectiveness against Dermacentor variabilis, the American dog tick. Objectives: * To assess the ability of an environmentally safe, rosemary oil-based pesticide, EcoEXEMPT IC2 (IC2), to control Ixodes scapularis, the vector tick of Lyme disease. * To assess, in the field and in tick enclosures, the toxicity of IC2 on the individual life stages of the tick. * To assess the impact of IC2 on non-target arthropods * To assess, in tick enclosures and in the field, the acaricidal efficacy of IC2 to D. variabilis, the dog tick. Proposal

Interim Report: Nov-30-2010

Outcomes Nymphal season application (70m2 grids): Pre-spray IC2, bifenthrin, and reference nymph counts were greater than post-spray counts (Figure 1, included in full report). On all post-spray sampling dates out to 1yr, nymph counts on IC2 and bifenthrin plots were equivalent, and lower than on reference plots (Figure 1, included in full report). IC2 evidently killed ticks on contact, since IC2 breaks down within days and has no lasting residual effect. In reference plots, fewer post-spray nymphs reflect the tapering off of the nymphal season in 2009 and the dry summer in 2010. There were fewer (and equivalent) post-spray larvae in both years on IC2 and bifenthrin plots (Figure 2, included in full report), suggesting IC2 had toxic effects on some tick eggs developing at the time of the nymphal spray. There were fewer (and equivalent) adults in both years on IC2 and bifenthrin plots (Figure 3, included in full report), suggesting IC2 reduced the number of nymphs available to molt to adulthood; this effect was still evident at 1.25yr post-spray. Adult season application (70m2 grids): Pre-spray, adult counts among treatment and reference plots did not differ (Figure 4, included in full report). Post-spray adult counts on IC2 and bifenthrin plots were equivalent and lower than on reference plots through the following spring's adult peak. One year later, adult counts on IC2 plots were lower than on reference plots but higher than on bifenthrin plots. This suggested IC2 was as efficacious as bifenthrin for at least 0.5yr, but not for 1yr. This may reflect bifenthrin's residual toxic effects. Pitfall trap arthropods (70m2 grids): The orders Coleoptera, Collembola, Diptera, and Hymenoptera were selected for analysis because these orders are ubiquitous and abundant in the region and over the duration the sample period. There was a significant treatment × date interaction for Coleoptera and Hymenoptera. One week after spray, Coleopterans were knocked back by both IC2 and bifenthrin, but at weeks 3 and 7 post-spray, only bifenthrin demonstrated a negative impact (Figure 5, included in full report), probably due to bifenthrin's residual toxicity. Most Coleopterans were distributed among the families Carabidae, Staphylinidae, Nitidulidae, and Scarbaeidae. The same pattern was apparent for Hymenopterans, comprising mostly Formicidae (Figure 6, included in full report). For Dipterans there was a significant treatment × date interaction, but with July 12th showing high numbers following the spray, it was not meaningful in terms of acaricidal treatment (Figure 7, included in full report). For Collembolans there was no significant treatment × date interaction (Figure 8, included in full report); dropping this term, only date was significant, meaning neither IC2 nor bifenthrin had a negative impact on Collembolans. Pollinator and pollinator nest count/survival (70m2 grids): In terms of bee and wasp nest production on the nymphal spray grids, there were no significant differences among the grids for the number of nests produced (n = 3 nests per treatment, Kruskal-Wallis P = 0.16). At the end of summer 2009 nest boxes were removed from the spray grids and stored indoors. Four insects emerged from three tunnels on May 25th and June 29th, 2010; it may take up to two years for pollinators to emerge from nests, so final nest survival may be assessed by the end of summer 2011. Bee/non-bee pollinator/total insect abundances on 70m2 spray grids were difficult to interpret given weather-driven sporadic sampling. Pollinator counts on 2m2 flowering plant plots experiment (2m2 plots): There were no significant treatment × date interactions for bee pollinators, non-bee pollinators, and total insects (all P e 0.36, Figures 9-11, included in full report). Upon removing the interaction term, only date was significant in all three models, meaning neither IC2 nor bifenthrin had a negative impact on these non-target arthropods. Date differences were driven by greater abundances in August; summer 2010 was drier and sunnier than summer of 2009. IC2 sprayed during the nymphal season was as efficacious as bifenthrin in keeping deer tick larva, nymph, and adult numbers down through 1yr post-spray, which was longer than expected (5wk). Also unexpected was that pollinators (and all insects) on 2m2 flower plots were not affected by either acaracide, whereas expected impacts were seen in the pitfall trap experiment in some arthropod groups. Reasons for this will be explored, such as different time of spray, truck versus backpack method of spray, and different insect niches. A third unexpected result was that IC2 exhibited phytotoxic properties against some understory and ground species. While this was immediately noticeable on the leafy surfaces of the plants, there was no evidence of long-term damage to vegetation. This should be considered, however, if IC2 is used in an area where management or conservation of sensitive vegetation (e.g. ferns, wildflowers) is of concern. Table 2 (included in full report) lists plants affected by the IC2 treatment in summer 2009. In the preliminary 2010 laboratory trials exposing eggs to 10-60µl of IC2 (field concentrations) on filter paper in mesh-covered vials, eggs survived and larvae emerged at lower concentrations; eggs did not survive at higher concentrations. Preliminary trials with adult ticks were similarly successful.

Impacts IC2, a food-grade, commercially available, minimal risk pesticide whose principal ingredient is rosemary oil, is an effective control against Ixodes scapularis when compared to a classic synthetic pyrethroid over the course of one year and when applied via high pressure sprayer. A year following summer application, IC2 had an effect comparable to bifenthrin on nymphal, larval, and adult deer ticks. A year following fall application, IC2 had an effect comparable to bifenthrin the following spring, and was still effective the following fall, but not as effective as bifenthrin. Based on preliminary analysis, IC2 may negatively impact some non-target insects of the type caught in pitfall traps, but not pollinators found on flowering plants or pollinator nests. The preliminary results of this study indicate IC2 offers an alternative, environmentally friendly approach to controlling ticks--on pastures where animals can be allowed to graze shortly after application, for organic farmers, and on the properties of homeowners concerned about the potential environmental damage and toxicity of synthetic acaricides to children and pets. The availability of an effective, minimum risk acaricide will greatly enhance IPM programs to reduce the risk of Lyme disease, anaplasmosis, and babesiosis. We hope to apply our findings to island communities of the Maine coast where IPM is greatly needed to reduce deer tick numbers. Preliminary results of the Minigrant leading to this study, and this study, were presented at the June 2010 at the EcoSmart Scientific Advisory Panel Meeting in Boston. The Minigrant results were published in 2010: Trial of a Minimal-Risk Botanical Compound to Control the Vector Tick of Lyme Disease. Peter W. Rand, Eleanor H. Lacombe, Susan P. Elias, Charles B. Lubelczyk, Theodore St. Amand, and Robert P. Smith, Jr., Journal of Medical Entomology 47(4): 695-698. This two-year project enjoys collaboration with biologists from both the University of Maine (Dr. Constance Stubbs) and the University of Southern Maine (Dr. Joseph Staples). This study will likely lead to a similar study on dog ticks (Dermacentor variabilis) and would include examination of the potential run-off of botanicals into tidal flats and impact, if any, on juvenile lobsters. This would entail collaboration with lobster experts in the Maine research community.

Report Appendices Progress Report 2010 [PDF]

Final Report:

Outcomes Nymphal season (summer) application (70m2 grids): * Nymphs: Pre-spray IC2, bifenthrin, and reference nymph counts were greater than post-spray counts (Figure 1, Full Report). On all post-spray sampling dates out to 1yr, nymph counts on IC2 and bifenthrin plots were equivalent, and lower than on reference plots (Figure 1, Full Report). IC2 evidently killed ticks on contact, since IC2 breaks down within days and has no lasting residual effect. In reference plots, fewer post-spray nymphs reflected the tapering off of the nymphal season in 2009 and the dry summer in 2010. Larvae: There were fewer post-spray larvae in both years on sprayed plots (reference > IC2 = bifenthrin) on sampling dates during the larval seasons of 2009 and 2010 (Figure 2, Full Report). This suggested IC2 sprayed during the nymphal season in July had toxic effects on some tick eggs developing at the time of the nymphal spray. Adults: There were fewer post-spray adults in both years on sprayed plots (reference > IC2 = bifenthrin) on sampling dates during the adult tick seasons of 2009 and 2010 (Figure3, Full Report). This suggested IC2 reduced the number of nymphs available to molt to adulthood; this effect was still evident at 1.25yr post-spray. Adult season (fall) application (70m2 grids): * Adults: Relative to reference plots sprayed with water, ticks were substantially reduced on plots treated with IC2 and bifenthrin (Figure 4, Full Report). On pre-spray plots, adult counts among treated and reference plots did not differ. One week post-spray adult counts on IC2 and bifenthrin plots were zero and remained zero or close to zero through the spring of 2010. One year later, adult counts on IC2 plots were lower than on reference plots but higher than on bifenthrin plots. This suggested IC2 was nearly as efficacious as bifenthrin for at least 0.5yr, but not for 1yr. This reflected bifenthrin's residual toxic effects. After the adult spray application on October 21, 2009, we also sampled for nymphs (June 1 and July 15, 2010) and larvae (August 27, 2010) in the adult spray plots. For both dates mean nymphs/m2 in IC2 and bifenthrin plots were zero and significantly less than in reference plots. On August 27th mean larvae/m2 in IC2 and bifenthrin plots were equal, and significantly less than in reference plots. Toxicity and short-term residual effect of IC2 on ticks in the laboratory: * The field-applied concentration of 4oz/gal of IC2 (3.1% solution) applied to a 4cm2 piece of filter paper in the laboratory killed 100% of adult ticks with one week's exposure (Figure 5, Full Report). LC50 and LC90 with 95% fiduciary limits were 1.8% (1.3-2.3%) and 3.7% (2.9-5.4%), respectively. At the end of one week, the field-strength IC2 solution (3.1%) killed 100% of larvae (Figure 6, Full Report). LC50 and LC90 1.0% (0-2.3%) and 2.0% (1.3-9.9%), respectively, but model fit was questionable due to a reversal in percent mortality at the 0.5 and 1.0% concentrations. Concentration-mortality curves for deer tick eggs did not fit the logistic survival curve (Figure 7, Full Report) so rather than use probit models we used chi-square tests for differences in % mortality. For both undeveloped eggs and those with visible embryos mortality was substantially greater (e96%) at the field-strength IC2 concentration (4oz/gal, 3.1%) than at each of the lower concentrations. Also, mortality was greater for eggs without embryos than eggs with embryos for the 0-1.9% concentrations but equivalent at 3.1%. The reason for this difference is not clear. Mortality was 100% when adult deer ticks were exposed to field-strength IC2 (4oz/gal, 3.1% concentration) at the time of application, and 93% when exposed to IC2 two hours after application (Figure 8, Full Report). LT50 and LT90 with 95% fiduciary limits were 8.6hr (6.9-11.0) and 3.0hr (0-4.9), respectively. Mortality at 12, 16, and 20hr was comparable to the control, meaning IC2 has rapidly decreasing efficacy after 8hr. We note that the variability in mortality within doses and across trials could be in part due to the fact that the adult ticks (including females that laid eggs) used were wild-caught rather than laboratory-reared. We also note that ticks that were temporarily immobile yet survived the laboratory experiments might have died in field conditions where temperature and humidity conditions would not be controlled. Pitfall trap arthropods (70m2 grids): * The orders Coleoptera, Hymenoptera, and Collembola were selected for analysis because these orders are abundant in the study area and represented 12%, 5%, 42% respectively, of all arthropods collected in this study. Other orders comprised insects that for the most part would not be expected to be on the ground at the time of the spray. There was a significant treatment × date interaction for Coleoptera (Figure 9, Full Report). Pre-spray, there were no differences among treatments. One week post-spray, a decline in abundance on all plots is evidence of a regional temporal decline, but in both IC2- and bifenthrin-treated plots, relatively greater declines indicated both acaricides caused a significant reduction in abundance Coleoptera (treatment × date interaction July 5th vs. 12th). After July 12th, only bifenthrin demonstrated a significant reduction in abundance. In September and October abundances across all treatments declined, likely due seasonal factors rather than treatments, and no significant differences were observed among treatments. Most Coleopterans were distributed among the families Carabidae (42%), Staphylinidae (20%), Nitidulidae (12%), and Scarbaeidae (10%).* A similar treatment × date interaction was observed for the Hymenoptera as seen in the Coleoptera; pre-spray, there were no differences among treatments (Figure 10, Full Report) but post-spray treatment effects were evident. One week post-spray, IC2 and bifenthrin both demonstrated fewer Hymenoptera (treatment × date interaction July 5th vs. 12th). At week 3 (July 26th), both IC2 and bifenthrin resulted in a significant reduction in total abundance of Hymenoptera relative to the reference treatment, but numbers recovered in IC2 plots by the end of August. However bifenthrin plots demonstrated fewer Hymenoptera throughout the season until October abundance across all treatments declined to end-of-season levels. The two most abundant familes for this order were the Formicidae (66%) and Platygastridae (13%). For Collembola there was a significant treatment × date interaction attributable to the acaricidal treatments (Figure 11, Full Report) one-week post-spray, but beyond this week neither IC2 nor bifenthrin had a negative impact on abundance in this order. Pollinator and pollinator nest count/survival (70m2 grids): * In terms of bee and wasp nest production on the nymphal spray grids, there were no significant differences among the grids for the number of nests produced). At the end of summer 2009 nest boxes were removed from the spray grids and stored indoors. Four insects emerged from three tunnels on May 25th and June 29th, 2010; it may take up to two years for pollinators to emerge from nests, so final nest survival may be assessed by the end of summer 2011. Bee/non-bee pollinator/total insect abundances on 70m2 spray grids were difficult to interpret given weather-driven sporadic sampling. * Pollinator counts on 2m2 flowering plant plots experiment (2m2 plots): * There were no significant treatment × date interactions for bee pollinators, non-bee pollinators, and total insects (Figures 12-14, Full Report). Only date was significant in all three models, meaning neither IC2 nor bifenthrin had a negative impact on these non-target arthropods. Date differences were driven by greater abundances in August 2010; summer 2010 was drier and sunnier than summer of 2009. Unexpected/noteworthy events: * IC2 sprayed during the nymphal season was as efficacious as bifenthrin in keeping deer tick larva, nymph, and adult numbers down through 1yr post-spray, which was longer than expected (5wk). Also unexpected was that pollinators on 2m2 flower plots were not affected by either acaracide, whereas expected impacts were seen in the pitfall trap experiment in some arthropod groups. Reasons for this will be explored, such as different time of spray, high-pressure versus backpack method of spray, and different insect niches. A third unexpected result was that IC2 exhibited phytotoxic properties against some understory and ground species (Cloyd et al. 2009). While this was immediately noticeable on the leafy surfaces of the plants, there was no evidence of long-term damage to vegetation. This should be considered, however, if IC2 is used in an area where management or conservation of sensitive vegetation (e.g. ferns, wildflowers) is of concern. Table 2 lists plants affected by the IC2 treatment in summer 2009. Outputs: Results of the USDA Minigrant leading to this study, and preliminary results of this study were presented at the EcoSmart Scientific Advisory Panel Meeting in Boston, June 2010. The Minigrant results were published in 2010: Trial of a Minimal-Risk Botanical Compound to Control the Vector Tick of Lyme Disease. Peter W. Rand, Eleanor H. Lacombe, Susan P. Elias, Charles B. Lubelczyk, Theodore St. Amand, and Robert P. Smith, Jr., Journal of Medical Entomology 47(4): 695-698. Results of this study will be prepared for publication in 2012. This two-year project was enhanced by collaboration with biologists from the University of Maine, Dr. Constance Stubbs, and the University of Southern Maine, Dr. Joseph Staples, as well as with Theodore St. Amand, president, and the staff of Atlantic Pest Solutions, a registered Integrated Pest Management business.

Outcomes Nymphal season (summer) application (70 x 70m grids): * Nymphs: Pre-spray IC2, bifenthrin, and reference nymph counts were greater than post-spray counts (Figure 1, Full Report). On all post-spray sampling dates out to 1yr, nymph counts on IC2 and bifenthrin plots were equivalent, and lower than on reference plots (Figure 1, Full Report). IC2 evidently killed ticks on contact, since IC2 breaks down within days and has no lasting residual effect. In reference plots, fewer post-spray nymphs reflected the tapering off of the nymphal season in 2009 and the dry summer in 2010. Larvae: There were fewer post-spray larvae in both years on sprayed plots (reference > IC2 = bifenthrin) on sampling dates during the larval seasons of 2009 and 2010 (Figure 2, Full Report). This suggested IC2 sprayed during the nymphal season in July had toxic effects on some tick eggs developing at the time of the nymphal spray. Adults: There were fewer post-spray adults in both years on sprayed plots (reference > IC2 = bifenthrin) on sampling dates during the adult tick seasons of 2009 and 2010 (Figure3, Full Report). This suggested IC2 reduced the number of nymphs available to molt to adulthood; this effect was still evident at 1.25yr post-spray. Adult season (fall) application (70 x 70m grids): * Adults: Relative to reference plots sprayed with water, ticks were substantially reduced on plots treated with IC2 and bifenthrin (Figure 4, Full Report). On pre-spray plots, adult counts among treated and reference plots did not differ. One week post-spray adult counts on IC2 and bifenthrin plots were zero and remained zero or close to zero through the spring of 2010. One year later, adult counts on IC2 plots were lower than on reference plots but higher than on bifenthrin plots. This suggested IC2 was nearly as efficacious as bifenthrin for at least 0.5yr, but not for 1yr. This reflected bifenthrin's residual toxic effects. After the adult spray application on October 21, 2009, we also sampled for nymphs (June 1 and July 15, 2010) and larvae (August 27, 2010) in the adult spray plots. For both dates mean nymphs/m2 in IC2 and bifenthrin plots were zero and significantly less than in reference plots. On August 27th mean larvae/m2 in IC2 and bifenthrin plots were equal, and significantly less than in reference plots. Toxicity and short-term residual effect of IC2 on ticks in the laboratory: * The field-applied concentration of 4oz/gal of IC2 (3.1% solution) applied to a 4cm2 piece of filter paper in the laboratory killed 100% of adult ticks with one week's exposure (Figure 5, Full Report). LC50 and LC90 with 95% fiduciary limits were 1.8% (1.3-2.3%) and 3.7% (2.9-5.4%), respectively. At the end of one week, the field-strength IC2 solution (3.1%) killed 100% of larvae (Figure 6, Full Report). LC50 and LC90 1.0% (0-2.3%) and 2.0% (1.3-9.9%), respectively, but model fit was questionable due to a reversal in percent mortality at the 0.5 and 1.0% concentrations. Concentration-mortality curves for deer tick eggs did not fit the logistic survival curve (Figure 7, Full Report) so rather than use probit models we used chi-square tests for differences in % mortality. For both undeveloped eggs and those with visible embryos mortality was substantially greater (e96%) at the field-strength IC2 concentration (4oz/gal, 3.1%) than at each of the lower concentrations. Also, mortality was greater for eggs without embryos than eggs with embryos for the 0-1.9% concentrations but equivalent at 3.1%. The reason for this difference is not clear. Mortality was 100% when adult deer ticks were exposed to field-strength IC2 (4oz/gal, 3.1% concentration) at the time of application, and 93% when exposed to IC2 two hours after application (Figure 8, Full Report). LT50 and LT90 with 95% fiduciary limits were 8.6hr (6.9-11.0) and 3.0hr (0-4.9), respectively. Mortality at 12, 16, and 20hr was comparable to the control, meaning IC2 has rapidly decreasing efficacy after 8hr. We note that the variability in mortality within doses and across trials could be in part due to the fact that the adult ticks (including females that laid eggs) used were wild-caught rather than laboratory-reared. We also note that ticks that were temporarily immobile yet survived the laboratory experiments might have died in field conditions where temperature and humidity conditions would not be controlled. Pitfall trap arthropods (70 x 70m grids): * The orders Coleoptera, Hymenoptera, and Collembola were selected for analysis because these orders are abundant in the study area and represented 12%, 5%, 42% respectively, of all arthropods collected in this study. Other orders comprised insects that for the most part would not be expected to be on the ground at the time of the spray. There was a significant treatment × date interaction for Coleoptera (Figure 9, Full Report). Pre-spray, there were no differences among treatments. One week post-spray, a decline in abundance on all plots is evidence of a regional temporal decline, but in both IC2- and bifenthrin-treated plots, relatively greater declines indicated both acaricides caused a significant reduction in abundance Coleoptera (treatment × date interaction July 5th vs. 12th). After July 12th, only bifenthrin demonstrated a significant reduction in abundance. In September and October abundances across all treatments declined, likely due to seasonal factors rather than treatments, and no significant differences were observed among treatments. Most Coleopterans were distributed among the families Carabidae (42%), Staphylinidae (20%), Nitidulidae (12%), and Scarbaeidae (10%). A similar treatment × date interaction was observed for the Hymenoptera as seen in the Coleoptera; pre-spray, there were no differences among treatments (Figure 10, Full Report) but post-spray treatment effects were evident. One week post-spray, IC2 and bifenthrin both demonstrated fewer Hymenoptera (treatment × date interaction July 5th vs. 12th). At week 3 (July 26th), both IC2 and bifenthrin resulted in a significant reduction in total abundance of Hymenoptera relative to the reference treatment, but numbers recovered in IC2 plots by the end of August. However bifenthrin plots demonstrated fewer Hymenoptera throughout the season until October abundance across all treatments declined to end-of-season levels. The two most abundant familes for this order were the Formicidae (66%) and Platygastridae (13%). For Collembola there was a significant treatment × date interaction attributable to the acaricidal treatments (Figure 11, Full Report) one-week post-spray, but beyond this week neither IC2 nor bifenthrin had a negative impact on abundance in this order. Pollinator and pollinator nest count/survival (70 x 70m grids): * In terms of bee and wasp nest production on the nymphal spray grids, there were no significant differences among the grids for the number of nests produced). At the end of summer 2009 nest boxes were removed from the spray grids and stored indoors. Four insects emerged from three tunnels on May 25th and June 29th, 2010; it may take up to two years for pollinators to emerge from nests, so final nest survival may be assessed by the end of summer 2011. Bee/non-bee pollinator/total insect abundances on 70 x 70m spray grids were difficult to interpret given weather-driven sporadic sampling. * Pollinator counts on 2 x 2m flowering plant plots experiment (2 x 2m plots): * There were no significant treatment × date interactions for bee pollinators, non-bee pollinators, and total insects (Figures 12-14, Full Report). Only date was significant in all three models, meaning neither IC2 nor bifenthrin had a negative impact on these non-target arthropods. Date differences were driven by greater abundances in August 2010; summer 2010 was drier and sunnier than summer of 2009. Unexpected/noteworthy events: * IC2 sprayed during the nymphal season was as efficacious as bifenthrin in keeping deer tick larva, nymph, and adult numbers down through 1yr post-spray, which was longer than expected (5wk). Also unexpected was that pollinators on 2 x 2m flower plots were not affected by either acaracide, whereas expected impacts were seen in the pitfall trap experiment in some arthropod groups. Reasons for this will be explored, such as different time of spray, high-pressure versus backpack method of spray, and different insect niches. A third unexpected result was that IC2 exhibited phytotoxic properties against some understory and ground species (Cloyd et al. 2009). While this was immediately noticeable on the leafy surfaces of the plants, there was no evidence of long-term damage to vegetation. This should be considered, however, if IC2 is used in an area where management or conservation of sensitive vegetation (e.g. ferns, wildflowers) is of concern. Table 2 lists plants affected by the IC2 treatment in summer 2009. Outputs: Results of the USDA Minigrant leading to this study, and preliminary results of this study were presented at the EcoSmart Scientific Advisory Panel Meeting in Boston, June 2010. The Minigrant results were published in 2010: Trial of a Minimal-Risk Botanical Compound to Control the Vector Tick of Lyme Disease. Peter W. Rand, Eleanor H. Lacombe, Susan P. Elias, Charles B. Lubelczyk, Theodore St. Amand, and Robert P. Smith, Jr., Journal of Medical Entomology 47(4): 695-698. Results of this study will be prepared for publication in 2012. This two-year project was enhanced by collaboration with biologists from the University of Maine, Dr. Constance Stubbs, and the University of Southern Maine, Dr. Joseph Staples, as well as with Theodore St. Amand, president, and the staff of Atlantic Pest Solutions, a registered Integrated Pest Management business.

Impacts Safeguarding human health and the environment: This project has confirmed and extended the findings of previous research (Rand et al. 2010) that IC2, a commercially available, minimal risk botanical product containing rosemary oil, is as effective for months after application as the synthetic pyrethroid bifenthrin in controlling nymphs and adults of Ixodes scapularis, the vector tick of Lyme disease, anaplasmosis, babesiosis and deer tick virus. In addition, the compound, when applied pre-eclosion, also appears to inhibit larval emergence. While IC2 may negatively impact three orders of non-target insects (Coleoptera, Hymenoptera, Collembola), this effect was less than that of the synthetic pesticide, and populations were greatly restored within one month of application. Neither synthetic nor botanical compounds were shown to harm pollinators visiting flowering plants. Laboratory studies confirm that the lethality of volatile IC2 on adult I. scapularis declines rapidly after 4 hrs of application, its long-term success thus being the result of very effective initial knockdown. One negative impact of IC2 as applied by high-pressure sprayer is a degree of phytotoxicity, which, while evident post-spray, appears not to be permanently harmful to vegetation. To minimize the number of comparisons to be made, no adjuvant for IC2 was used in our study; its addition, as recommended by the manufacturer, might allow reduction in IC2 concentration to a level that should also reduce phytotoxicity without loss of effectiveness. Human health impact: Since infected I. scapularis were first reported in Maine in 1988 (Ginsberg and Ewing 1988), this tick has become abundant and firmly established in the southern half of the state, particularly along the coast. I. scapularis is now found, and is transmitting Lyme disease, in northernmost Aroostook County towns along the Canadian border (Rand et al. 2007, 2011). In a recent national tick survey (Diuk-Wasser et al. 2006), the highest density of nymphal ticks was collected in south coastal Cape Elizabeth. Over the last 2½ decades, cases of human, canine, and equine Lyme disease have increased, gradually during the 1990s, then precipitously in the last decade. In 2008, reported human cases per 100,000 reached 169 in mid-coastal Knox County (Robbins 2008). Close to 1000 cases were reported statewide in 2009, likely representing one-fifth to one-half of actual infections (Meek et al. 1996). In the mid 1990's the pathogen of human anaplasmosis was first found in Maine ticks (Holman et al. 2004, and now increasing cases of that disease are being reported in southern counties (Cahill et al. 2009) while exposure to the pathogen has become widespread in the canine population (Rand et al. 2011). In addition, within the last ten years tick-transmitted babesiosis has appeared in Maine and the number of reported cases of that disease is also rising. With no human vaccines available, prevention of these diseases will rest on an informed, motivated, and responsive public. Part of the message that public health authorities can now convey is that tick control can be achieved at minimal risk to the environment. Environmental impact Public concern, in part inflated by chronic Lyme disease activism, has reached high levels, and with it increasing demand for the use of tick control products both by the homeowner and by professionals. Given the high effectiveness of synthetic acaricides, this demand has undoubtedly resulted in increase in their use. Recognizing the public's concern about the potential damaging effects of synthetic pesticides on non-target species, the environment, and human health, there has been a surge of interest and research in the development of effective, botanically-derived, minimal risk acaricides (i.e. Dolan et al 2009), particularly as a component of a multi-faceted IPM approach involving habitat modifications and management of white tailed deer, the tick's primary reproductive-stage host (Stafford 2004). As even more effective botanically-derived acaricides are discovered, years may take place before they are commercially developed and marketed. The research reported here demonstrates that an effective, minimal risk tick control product is already available. Economic Benefits It appears that the major challenge to the botanical acaricide industry is not so much improvement of effectiveness as it is to reduction of cost. For example, the price of sufficient IC2 to high-pressure spray one acre would be roughly $490, while that of an equivalent amount of bifenthrin would be ~$27, or 18 times less. While this differential would be less important on small peridomestic plots, considering the total costs of application, it rapidly becomes more significant as treated areas increase. A similar issue greets the homeowner considering self-application of products off the hardware store shelf, where a 20 lb bag of a synthetic (Triazicide), recommended to treat 25,000 ft2, can be purchased for $14, while next it will be a 10 lb bag of a botanical (EcoSMART Organic Insect Killer) for $11, which will treat only 5000 ft2. A mitigating factor that has yet to be explored for botanicals, however, is the possibility that, as reported for synthetics (Schulze et al. 2008), a single application to eradicate adults in the fall may sufficiently reduce ticks year-round, particularly if combined with other components of an integrated tick management program. Other benefits of IC2 application affect the balance of environmental protection and costs. Being a food grade compound, there are few concerns about the return of pastured animals to grazing after application. Anecdotally, we first recognized the effectiveness of this product when it was applied to the pasture of a neighbor whose two Belgian draft horses had required massive dosing with doxycycline to treat Lyme borreliosis. This has not been a problem in the ensuing four years. The major financial benefit of acaricides, botanical or synthetic, is the prevention of diseases which, for some, may be very expensive in terms of treatment and lost income, not to mention pain and emotional stress. Implementation of IPM We believe that this field trial and the one that preceded it confirm the acaricidal efficacy of this rosemary-containing compound -- and that this effectiveness invites further studies to determine the most appropriate times (or time) for its application relative to the phenologies of the nymphal and adult stages of I. scapularis. This knowledge will guide just how its use can be most effectively and economically incorporated into an integrated tick management plan for use on residential properties, farms, recreational areas, public properties, schools, and outdoor industrial sites. Implementation, however, will require concerted public education about tick-borne diseases, tick habitats, the role of critical tick hosts (deer), the span of tick control methods, as well as ways to prevent exposure to ticks. We are currently working with an offshore Maine island community, which, due to its centralized structure and limited recruitment of white tailed deer, provides an excellent opportunity to implement all components of a comprehensive tick management program. Our primary educational contribution has been the publication of our preceding IC2 study (Rand et al., 2010), and with completion of our laboratory studies on the effect of IC2 of I. scapularis egg development, we will submit the findings of the present study for publication. On our agenda is the development of an on-line decision analysis tool for the public to select the best approach to tick control based on site-specific habitat, property size, current exposure and transmission risk, family structure, motivation, affordability and other risk data that we have acquired over two decades of work in Maine. Our work is summarized in our website, www.mmcri.org/lyme, which also contains a wide variety of information on tick-borne diseases, their prevention, and tick control.

Report Appendices Final Report 2011 [PDF]

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