The other pesticides tested caused only low rates of acute mortality in H. convergens and were considered harmless. However, in addition to direct effects on mortality, it is also important to test for potential sublethal effects of pesticide exposure, such as effects on development time, sex ratio, fecundity, or fertility, before concluding whether a particular pesticide is selective and compatible with a natural enemy species . Therefore, while our laboratory bio-assays based on acute mortality have been effective in identifying potentially disruptive materials for H. convergens, they have not been sufficient to identify truly selective and compatible products from among the seven pesticides tested. Further research on potential sublethal effects of these pesticides on H. convergens is necessary to expand our understanding of how they might impact this important coccinellid predator under field conditions. There is increasing interest in the effects of pesticides on beneficial insects . Natural enemies have been a major focus of this research due to their contribution to pest management. Classic methods for establishing the toxicity of pesticides on beneficial insects were to determine the median lethal dose and/or lethal concentration . While acute toxicity assays may be a good starting point for understanding the effects of pesticides, they are limited in that they only measure survivor ship and cannot account for sublethal effects.Pesticides are also known to influence physiology,dutch buckets for sale with consequent effects on development, longevity, fecundity, fertility and sex ratio .
Natural enemies exposed to pesticides may also experience changes in behavior, including mobility, feeding, and mating. Therefore, to more accurately assess pesticide toxicity, sublethal effects must be considered. It is well known that organophosphate pesticides are highly toxic to natural enemies.From 1990 to 2007, the use of organophosphates decreased from 70% of total insecticide use to 36% . Due to the passage of the Food Quality Protection Act in the United States in 1996, this class of conventional pesticides became increasingly restricted, which initiated the use alternative pesticides for the control of pest populations. One such alterative has been the development of reduced-risk pesticides, promoted by the Environmental Protection Agency Office of Pesticide Programs’ Reduced-Risk Pesticide Initiative. In 1992, via the Federal Register, the EPA-OPP called for incentives to promote the development and use of reduced-risk pesticides, and, the following year, announced the Reduced-Risk Pesticide Initiative. The EPA created incentives to promote registration of these pesticides under new reduced-risk criteria that included reducing human health risks and reducing risks to non-target organisms such as birds, beneficial insects, and aquatic organisms . Recently, reduced-risk pesticides have been found to show sublethal effects on several different species of natural enemies. For instance, chlorantraniliprole is an anthranilic diamide designed to target lepidopteran pests, yet when tested for behavioral effects on the generalist predator Macrolophus pygmaeus , exposure to the chemical reduced feeding . Similarly, the parasitoid Diadegma insulare Cresson was able to distinguish between host larvae fed on plants treated with the pyrethroid, lambda-cyhalothrin, versus non-treated plants, indicating that pesticide exposure can alter host preference .
Even if a pesticide demonstrates no behavioral effects, it may still induce substantial effects on life history traits. For example, cyantraniliprole, another anthranilic diamide that targets lepidopteran pests, caused a reduction in prey consumption and fecundity for the predator mite, Galendromus occidentalis Nesbitt . In addition, when G. occidentalis females were exposed to copper+mancozeb, there was no effect on egg hatch, but, once hatched, the larvae experienced higher mortality . Similarly, while survivor ship of female Chrysoperla carneaand Chrysoperla johnsoni Henry, Wells and Pupedis treated with novaluron was unaffected, their subsequent egg viability was reduced to almost zero . When considering sublethal effects, demographic toxicology provides a useful method of assessment for integrating the effects of a toxicant on individual life history parameters into a combined effect on the population growth rate of an organism . For example, using a demographic approach based on life table data , were able to calculate the critical extinction thresholds for four economically important parasitoids: Diachasmimorpha longicaudata , Psyttalia fletcheri , Fopius arisanus , and Diaeretiella rapae.Our approach in this study was to examine the lethal and sublethal effects of seven pesticides on the convergent ladybird beetle, Hippodamia convergensand their influence on its potential for population growth. The pesticides tested are commonly used in high-value tree crops in western North America, where H. convergens is an important natural enemy . The pesticides included five reduced-risk insecticides that are used for management of codling moth, Cydia pomonella , and two fungicides used for the management of bacterial and fungal diseases. The objectives of the study were 1) to assess the lethal effects of these pesticides on egg, larval, pupal, and adult life stages of H. convergens, 2) to determine the sublethal effects of exposure of first instar larvae and adults to these pesticides, and 3) to combine the lethal and sublethal effects to determine the impact of each pesticide on the potential population growth rate of H. convergens.
Adult H. convergens were obtained from a commercial source . In order to maintain insects in a state of hibernation, adults were stored for up to two months in a screened wooden cage at 6 °C. To prevent desiccation, the colony was sprayed with distilled water on a weekly basis and several 8.5 cm petri dishes lined with cotton wool were included in the cage. Adults were removed from cold storage, sexed, and individual mating pairs were placed into 96.1 ml cups , capped with perforated plastic lids for ventilation, and placed into an incubator maintained at 22 °C, 60-70% RH, and a 16:8 h photoperiod. Mating pairs were provided a 1:1 honey-water solution that was replaced every two days and were reared on a diet of Acrythosiphum pisum, which are known to provide a high quality diet for H. convergens . After 3-4 days, males were removed from the cups. Mated females were fed 20 adult aphids per day for a period of 10 days, which allowed time for oviposition to begin. After this period, mated females were then used for bio-assays. A separate group of mated females were used to produce eggs to be used in egg, larval, and pupal bio-assays. These females were placed individually to the same plastic cups, fed the same diet and assigned to the same incubator as described above. When an adult female oviposited an egg clutch on the inner surface of a cup, it was transferred to a new cup. Egg clutches aged 1-2 days were used in the egg bio-assays. For larval bio-assays,hydroponic net pots newly laid egg clutches were allowed to develop and hatch into first instar larvae . They remained in the same cup and were fed a diet of A. pisum, ad libitum, for a period of 48 h. First instar larvae aged 48 h were used in the larval bio-assays. For pupal bio-assays, egg clutches were allowed to develop into first instar larvae, and individual insects were transferred to 22.1 ml PETE cups with perforated lids to allow for ventilation. On a daily basis, each larva was provided with following number of adult A. pisum, first instar larvae were fed 5 aphids, second instar 10 aphids, third instar 15 aphids, and fourth instar 20 aphids. When larvae developed into pupae, they attached themselves to the cup surface. Pupae 1-2 days old were used in the pupal bio-assays.Seven pesticides were tested, including two anthranilic diamides , a pyrethroid , an insect growth regulator and a spinosyn , all of which are alternatives to OPs for management of codling moth; a mixture of copper hydroxide and mancozeb used for management of walnut blight and apple scab; and dry flowable sulfur used for management of mildew and apple scab. All pesticides were tested at the 100% maximum field rate in comparison to distilled water as a control. The pesticides were tested as formulated materials: chlorantraniliprole, cyantraniliprole and a mixture of copper hydroxide and mancozeb from DuPont, Wilmington, DE; lambda-cyhalothrin from Syngenta, Greensboro, NC; novaluron from Chemtura, Middlebury, CT; spinetoram from Dow AgroSciences, Indianapolis, IN; and dry flowable sulfur from Arysta LifeScience, Cary, NC. The 100% field rate concentrations were prepared as 50 ml solutions in distilled water using the following amounts of formulated material: 16.85 mg for chlorantraniliprole; 80 µl for cyantraniliprole; 240 mg for copper hydroxide and 107 mg for mancozeb; 9.9 µl for lambda-cyhalothrin; 195.2 µl for novaluron; 26.2 mg for spinetoram ; and 860 mg for dry flowable sulfur. All bio-assays were conducted at 22 °C, 60-70% RH, and a 16:8 h photoperiod.
Egg clutches and pupae remained in the cups in which they were reared and were coated with 200 µl of a given pesticide dripped from a pipette. Excess pesticide was poured out, and the cups were inverted and allowed to dry in a fume hood for 24 h. Egg hatch was recorded for 6 days after and pupal emergence for 10 days after the exposure event, and stage survivor ship was recorded in each case. For eggs, a minimum of 30 replicates and for pupae a minimum of 29 replicates were used. Mated adult females and first instar larvae were exposed to all pesticides, with the exception of lambda-cyhalothrin. In previous experiments, lambda-cyhalothrin was shown to be acutely toxic to both life stages which prevented the successful estimation of sublethal effects for this particular pesticide. In a field setting, it is possible for life stages of natural enemies that are actively mobile and feeding to receive topical exposure from direct spraying, residual exposure from a treated plant surface, and/or oral exposure from ingestion of a treated food source . Consequently, a maximum exposure scenario was assumed, and all three routes were tested simultaneously. 28-30 replicates were used for larval bio-assays, and 25-30 replicates for adult bio-assays. For residual exposure, single 15 x 45 mm glass vials were treated by adding 4 ml of a pesticide solution and turning the vial 360 degrees to coat the entire inside surface before pouring out the excess solution. Vials were inverted, suspended on a rack, and allowed to dry in a fume hood for a period of 24 h. For oral exposure, A. pisum were treated by being individually dipped in a pesticide solution, and then placed on a paper towel in a fume hood until excess pesticide had dripped off the aphids and they were able to crawl. 10 topically-treated adult A. pisum were placed in each residually-treated glass vial. For topical exposure, H. convergens were placed in an 8.5 cm petri dish with filter paper lining the bottom and sprayed in a Potter spray tower set to 68.9 kPa. For each pesticide solution, the spray volume per application was 1.4 ml, resulting in a spray deposit of 2.50 mg cm-2 . The deposit is similar to that used in other studies on non-target insects and to the recommendations of the IOBC Working Group “Pesticides and Beneficial Organisms” . The treated insects were allowed to dry and were then placed in treated vials containing treated aphids. Cotton wool stoppers were used for the glass vials to allow for ventilation. After the initial day of exposure to treated aphids, larvae were fed a diet of untreated aphids on a daily basis. The daily rate of provisioning of A.pisum adults for individual larvae was 5 for first instars, 10 for second instars, 15 for third instars, and 20 for fourth instars. Larvae were also transferred to untreated vials two days after the exposure event. They were monitored for 10 days after entering the pupal stage, after which they were considered dead. Larval stage survivor ship was recorded, and, for groups with sufficient adult emergence, gender ratio was recorded. Similarly, after the initial day of exposure to the pesticides, the mated females were transferred to untreated 96.1 ml cups and fed approximately 20 untreated adult A. pisum per day for a period of 14 days. Throughout this period, survivor ship and fecundity were recorded daily. Each day that a female oviposited an egg clutch, she was transferred to a new cup and the egg clutch remained attached to the wall of the initial cups. In order to determined fertility, egg clutches were checked daily for 7 days and hatched larvae were counted, removed, and recorded.