Final Report for ONE07-067
Resistance to neonicotinoid insecticides is widespread, though they still provides significant protection. One of the cornerstones of soil, disease, and insect management is the rotation of crops. Crop rotation decreases the size of colonizing populations of many insects, including Colorado potato beetle, Leptinotarsa decemlineata (Say). However, rotation may also reduce the level of insecticide resistance if resistant individuals are unable or less likely to migrate long distances to colonize new crops. Over two years we tested the prediction that rotation reduces the resistance of colonizing CPB by collecting emerging walking adults in the spring via drift fences, flying adults using harp traps, and colonizing adults using untreated trap plants at increasing distances from the overwintering site at distances of up to 500 meters. We also tested the resistance of colonizing adults on rested and un-rested fields, with the prediction that rotated fields colonized from longer distances will have less resistant colonists. In the first season we experienced difficulties with a larval filter paper assay, and we found data suggesting higher resistance in walking than flying emergers, but no difference between rotated and rested fields. In the second season we used adult assays that surprisingly and significantly showed higher resistance in flying beetles than in walking emergers, and higher resistance in each of three pairs of rotated vs. non-rotated fields from three different farms. Though these results contrast with other measured costs of resistance, they might result from higher condition in more resistant beetles in populations where all beetles are exposed to neonicotinoid insecticides. Even if there is a movement cost of resistance, it may be masked if all individuals are exposed. This project is continuing with laboratory assays of movement in more and less resistant beetles, and studies of resistance and flight in emergers from untreated and treated fields.
Previously we have found that resistance to imidacloprid entails significant costs, including reduced fecundity, overwinter survival, walking speed, and speed of development. The presense of these resistance costs led us to hypothesize that reduced long distance (flying) movement might also be a cost of resistance, which, if present, would lead to reduced resistance and reduced need for insecticide applications in rotated fields. To test this hypothesis we compared the resistance of flying emergers with that of walking emergers from a highly concentrated, central overwintering site surrounded by potato fields in Riverhead NY. We also compared the resistance of colonizing beetles in continuously planted fields, with those from rotated fields, where beetles could only colonize through longer distance movements.
Objective I: Set up drift fences with pitfall traps to collect walking emergers, and harp traps to collect flying emergers, to see if flying CPB in the spring are less resistant than walking emergers.
Objective II: Collect Colonizing CPB 100, 200, and 400 meters from their overwintering sites will be compared in resistance to see if colonizing distance is negatively correlated with resistance.
Objective III: Measure Resistance of CPB colonizing fields rested from, or planted in, potato the previous year, and correlated with the distance to the closest field the previous season.
Materials and Methods:
Objective I: In 2007 we set up drift fences around a small woodlot surrounded by about 325 acres of potato field in Riverhead NY. The woodlot attracts diapausing adults each fall. We collected early and late walking and flying beetles using drift fences made of aluminum roof flashing and pitfall traps along with harp traps to catch flying beetles. Over the course of the emergence season we used tennis racquets to collect flying beetles shortly after take-off, and that was the primary method of capture in 2008. In 2007 separate colonies were established with walking or flying adults captured at emergence from the diapause site. Clutches were collected and both 1st instar filter paper assays, where hatchlings were allowed to feed for six hours and then placed on filter paper which had from 1.6 to 49.5 µg/cm^2 imidacloprid applied in 0.1 ml acetone which was evaporated off prior to introducing the larvae. For adult assay each adult was treated with a single drop of 2 µl of imidacloprid dissolved in acetone at a concentration of 1E-3 g/ml. Adults were held in 60 mm petris with fresh potato foliage replaced daily and were scored at five and seven days following application. Assays were carried out on second-instar larvae by subjecting them to a 1 µl drop of a range of concentrations of imidacloprid dissolved in acetone from 1E-6 g/ml to 1E-4 g/ml plus control. Mortality was assayed after 24 hours. In 2008 adult assays alone were used, with the addition that six doses plus controls were used to determine an LD50 for flying and walking emergers. 153 flying adults were assayed, and LD50 was compared with that of 426 walking emergers assayed. Dose-response were analyzed using Polo-PC (LeOra Software 1987)
Objective II: In 2007 Adults were collected from trap plants at distances 50, 100, 200, 400, 500, and 640 meters from the overwintering site, established in small butterfly cages with potted potato plants, and clutches were collected for 1st instar assays. 10-60 adults were collected from each location, and 2188 hatchlings were assayed. In 2008 adults were collected at 100, 200, and 400 meters from the central overwintering site, and assayed at a single discriminating dose of 2 µl of imidacloprid dissolved in acetone at a concentration of 1E-3 g/ml.
Objective III: In 2007, 10-20 potted potato plants were placed in freshly planted potato fields in and near Riverhead, NY and Scuttlehole, NY. A late spring emergence in 2007 delayed appearance of potato beetles until after most field plants had erupted, so adults were collected from field plants, likely after consuming some treated foliage. Adults were housed in butterfly cages as above, and 5791 1st instar larvae were assayed. After collecting clutches for assay, the surviving adults themselves were assayed using a single drop of 2 µl of imidacloprid dissolved in acetone at a concentration of 1E-3. Mortality was assessed after seven days.
In 2008, three pairs of rotated and continuously planted fields were sampled on three different farms. Colonists were collected in May and June, held for several days on untreated plants to recover from exposure to systemic imidacloprid, and tested as above using a range of doses of imidacloprid dissolved in acetone. 115 to 328 adults were assayed from each field, and resistance was compared within each farm between the rotated and the continuously planted field.
In the first season conclusions were difficult because the first-instar assay used on the offspring of most collected adults was not reliable. Intermediate level doses had the highest mortality, suggesting that initial exposure at the highest levels limited uptake of toxin. There were no significant differences in resistance in first instar larvae among distances from the overwintering sites, nor among rotated and non-rotated fields, however, the lack of fit to the probit model in the data make it difficult to conclude anything from those assays.
We did observe some significant results in the second-instar assays. Second-instar larval offspring of walking emigrants from the overwintering site were 4.7 times as resistant as flying emigrants (using the ratio of the maximum likelihood fitted LC50’s), but the difference was marginally insignificant.
From the first season there was the suggestion that more resistant beetles are not able to travel as far to colonize fields in the spring. However, the avoidable difficulties with assays prevent drawing any strong conclusions from the first year, and we decided to use more reliable adult assays, and concentrate on collecting as many adults as possible early in the emergence season.
In 2008 We found flying beetles to be marginally more resistant than walking emergers (Figure 1). Flying emergers were about twice as resistant as walkers, but the relatively small number assayed (153) increased the confidence limits around the estimate, and made the resistance significantly higher at the P=0.1, but not P=0.5 level. The actual difference in resistance of flying and walking emergers may be higher, however, because the pool of walking emergers includes both some that would fly if not captured in the drift fence as well as those that would not fly even if given more time. The comparisons of rotated and continuously planted fields are a stronger test of the role of rotation on resistance management, both because they test the resistance of those that were in fact able to colonize more distant rotated fields. We had three within-farm comparisons of resistance of rotated vs. non-rotated fields, and in each farm, colonists of rotated fields were significantly more resistant than the colonists of fields planted in potato continuously (Figure 2). The results from 2008 appear to contradict the first year’s data, and our findings of significant resistance costs on several life history traits (Baker and Porter 2008, Baker et al. 2007). The 2008 results were based on assays of field-collected adults, all of whom likely exposed to imidacloprid the previous season. In a population where all individuals are exposed to imidacloprid, it is possible that only the most resistant are in sufficient physiological condition to fly and colonize rotated fields. It is also possible that resistant beetles are more motivated to or better able to fly, a result found in beetles exposed to Bt toxins (Alyokhin and Ferro 1999). Regardless of the mechanism, the results of the present study suggest that rotation by itself will not reduce resistance to imidacloprid, and in fact that rotated fields attract the most resistant colonists. They do, however, also attract the smallest number of colonists, depending on distance (Sexson and Wyman 2005), which offers an opportunity to skip neonicotinoid treatment early in the season on rotated fields, and use alternate treatments later in the season, saving neonicotinoids for the second year of planting.
Alyokhin AV, and DN Ferro. 1999. Modifications in flight and oviposition of Bt-resistant and Bt-susceptible Colorado potato beetles as a result of exposure to Bacillus thuringiensis subsp. tenebrionis Cry3A toxin. Entomol Exp Appl 90:93-101.
LeOra Software. 1994. POLO-PC: probit and logit analysis. LeOra Software, Berkeley, CA.
Baker, M. B., and A. H. Porter. 2008. Use of sperm precedence to infer the overwintering cost of resistance in Colorado potato beetle. Agricultural and Forest Entomology 10: 181-187.
Baker, M. B., Alyokhin, A., Porter, A. H., D. N. Ferro, S. R. Dastur, and N. Galal. 2007. Persistence of costs of resistance to imidacloprid in the Colorado potato beetle, Leptinotarsa decemlineata (Say). Journal of Economic Entomology 100: 1871-1879.
Sexson, D. L., and J. A. Wyman. 2005. Effect of crop rotation distance on populations of Colorado potato beetle (Coleoptera: Chrysomelidae): development of areawide Colorado potato beetle pest management strategies. J. Econ. Entomol. 98: 716-724.
This study contributes to the management of resistance to neonicotinoids, and potentially other treatments as well. The more resistant beetles become to neonicotinoids, the higher the application rates used by conventional growers, and the increased reliance on spinosad, which is a less persistent, biologically based insecticide, but which will also select for resistance. The results of this study were so unexpected that we are continuing the project in two ways to pin down the relationship between rotation and resistance management before making recommendations for growers. We are replicating last year’s design, comparing rotated and non-rotated fields within farms, and flying and walking emergers from the original emergence site and one newly identified emergence site. My collaborating graduate student is carrying out laboratory crosses and a flight-mill study to determine whether increased flight in more resistant beetles is due to higher body condition following exposure to insecticide, or is seen also in untreated individuals. Partly because of the preliminary nature of our results we have presented them at scientific meetings but not in outreach to farmers, but we will present this data at the next Long Island Ag forum.
This grant provided materials and expenses for the major thesis project of one PhD student, Kathleen Schnaars, and supported training of four other student researchers and technicians, Lynette Nitschke, Simon Greenbaum, Mary Polsunus and Karyn Collie. We worked with six cooperating farm families on Long Island over the two years; Steele, Zilnicki, Wines, Wesnofske, Kujawski, and Barbuto.
Education & Outreach Activities and Participation Summary
Baker, M. Rotation distance and resistance management in Colorado potato beetle: is movement a cost of resistance? Entomological Society of America meeting, November 2008, Reno, NV
Schnaars-Uvino, K. 2009. Resistance, movement and rotation distance in the Colorado potato beetle, Leptinotarsa decemlineata (Say). Entomological Society of America Eastern Branch, 80th Annual Meeting, March 2009, Harrisburg PA.
Schnaars-Uvino, K., and M Baker 2009. Resistance, movement and rotation distance in the Colorado potato beetle, Leptinotarsa decemlineata (Say). Entomological Society of America Annual Meeting, November 2008, Reno, NV.
Thus far no farmers have decided to leave rotated fields untreated if they are not collaborating with us on a study. Crop rotation is frequently practiced by conventional as well as organic potato growers on Long Island, mostly for plant disease rather than for resistance management. Discussions with farmers suggest that neonicotinoids will be used at planting for the foreseeable future for reducing potato beetle populations and providing perceived insurance against other pests, primarily leafhoppers and aphids, even in the face of increasing resistance to neonicotinoids. There are alternative treatments for potato beetle available, but despite the presence of alternatives a stronger case needs to be made for reducing early season neonicotinoid use to slow resistance evolution and reduce total insecticide application.
Areas needing additional study
The results from this study need replication, and point to a more complicated story than anticipated. This season the comparisons of rotated and non-rotated fields are being replicated, as well as the comparisons of flyers and walkers. Assuming the results are consistent, resistance may be genetically linked with increased movement, or resistance may just allow greater movement following field exposure to neonicotinoids. We are carrying out experiments this year with the untreated offspring of field-caught adults that will undergo diapause in the lab and then be tested for flight propensity on a flight mill and later assayed for resistance. This experiment will clarify which of the two mechanisms are operating. If resistance is genetically linked to greater movement, then there will be an argument for using alternative treatments in rotated fields, since neonicotinoids will be less effective and resistance potential is highest. If the increased movement is due to higher condition in resistant individuals in the face of exposure to neonicotinoids, then that would argue for reducing neonicotinoid exposure late in the summer, prior to crop rotation.