We measured the impact of sublethal pesticide exposure on cold tolerance of two common tick species. The proportion of ticks that survived exposure to LT50 (lethal temperature at which 50% mortality is expected) following a sublethal exposure to pesticides. Unfed ticks of both Rhipicephalus sanguineus and Dermacentor variabilis were obtained as eggs, engorged larvae, and nymphs. Chlorpyrifos methyl and propoxur were dissolved in 100% acetone to obtain stock solutions. The experimental solutions were then prepared by serial dilution with ultrapure deionized water at 2.5, 5.0, 10.0, 15.0, and 20.0 PPM. To determine the impact of pesticide exposure on cold tolerance, survival to cold exposure was determined using a 2-hour exposure to specific temperatures including -16°C for 2 hours, which is the published LT50 estimate for D. variabilis, -20°C, -22°C, -25°C, and -28°C, yielding survival estimates that ranged from ~50% at -16°C to nearly zero at -28°C. To measure the impact of pesticide exposure on overwintering survival, we compared two geographic locations that are characteristic of northern (Crooks, SD) and southern (Cincinnati, OH) overwintering conditions within the geographic range for D. variabilis. 400 adult D. variabilis were placed in exclosures in Crooks, SD, USA (North) and Cincinnati, OH, USA (South) from September through April. Adult D. variabilis (even numbers of males and females) were dosed with 2 ppm of chlorpyrifos, propoxur, or solvent and placed in conical tubes with holes drilled in the lid and sides. Ticks were buried at a depth of 10cm and retrieved in April and survival was assessed. The extensive use of pesticides may drive complex interactions between species and their environments, leading to altered thermal tolerance traits and establishment in new habitats. Sublethal doses of pesticides may impact tick cold tolerance and lead to greater winter survival particularly at northern range limits.

Ticks and Pesticides:

The unfed ticks for each life stage of both Rhipicephalus sanguineus and Dermacentor variabilis were obtained from Ecto Services Inc. (Henderson NC, USA) as eggs, engorged larvae, and nymphs. Chlorpyrifos methyl and propoxur were obtained from Chem Services Inc. with 99.7% and 99.9% purity, respectively. Both were dissolved in 100% acetone to obtain stock solutions of 1.0 mg mL−1 and kept at −18 °C. The experimental solutions were then prepared by serial dilution with ultrapure deionized water at 2.5, 5.0, 10.0, 15.0, and 20.0 PPM based on previous studies. Both pesticides were applied using either the larval packet testor submerging adults in 1.5 mL microcentrifuge tubes with pesticide solutions for 20 seconds. 

Thermal Tolerance:

To determine the impact of pesticide exposure on cold tolerance, survival to cold exposure was determined using a 2-hour exposure to specific temperatures including -16°C for 2 hours, which is the published LT50 estimate for D. variabilis, -20°C, -22°C, -25°C, and -28°C, yielding survival estimates that ranged from ~50% at -16°C to nearly zero at -28°C (Rosendale et al., 2016). Ticks were exposed to either propoxur, chlorypyrifos, or the solvent as described above, and left in rearing conditions (93% relative humidity (RH), 22 ± 1°C, and 12:12 h, light: dark (L: D)) for either 24-hours, 5-, 28-, or 55-days post-pesticide exposure. 

Overwintering:

To measure the impact of pesticide exposure on overwintering survival, we compared two geographic locations that are characteristic of northern (Crooks, SD) and southern (Cincinnati, OH) overwintering conditions within the geographic range for D. variabilis. 400 adult D. variabilis were placed in exclosures in Crooks, SD, USA (North) and Cincinnati, OH, USA (South) from September 2020 through April 2021.