Functional Response of Gonatocerus Ashmeadi and the “New Association” Parasitoid G

Functional Response of Gonatocerus ashmeadi and the “New Association” Parasitoid G. tuberculifemur Attacking Eggs of Homalodisca vitripennis Author(s): Nicola Irvin, Javier Suarez-Espinoza, and Mark Hoddle Source: Environmental Entomology, 38(6):1634-1641. 2009. Published By: Entomological Society of America DOI: http://dx.doi.org/10.1603/022.038.0616 URL: http://www.bioone.org/doi/full/10.1603/022.038.0616

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BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit publishers, academic institutions, research libraries, and research funders in the common goal of maximizing access to critical research. BIOLOGICAL CONTROLÑPARASITOIDS AND PREDATORS Functional Response of Gonatocerus ashmeadi and the “New Association” Parasitoid G. tuberculifemur Attacking Eggs of Homalodisca vitripennis

1,2 3 1,4 NICOLA IRVIN, JAVIER SUAREZ-ESPINOZA, AND MARK HODDLE

Environ. Entomol. 38(6): 1634Ð1641 (2009) ABSTRACT The functional response of two egg parasitoids, Gonatocerus tuberculifemur (Ogloblin) and G. ashmeadi Girault (Hymenoptera: Mymaridae), to varying densities of Homalodisca vitripennis (Germar) (Hemiptera: Cicadellidae) eggs was studied in the laboratory. G. tuberculifemur showed a type II functional response where parasitism increased at a decreasing rate as host egg density increased from 5 to 40, reaching an asymptote of an upper limit of 9.3 host eggs within the 1-h exposure period. The instantaneous attack rate and handling time as estimated by the random predator equation were 2.28/h and 0.10 h, respectively. In contrast, G. ashmeadi showed a type I functional response where the number of hosts attacked followed a constant linear increase regardless of host density to an upper limit of 11.5 host eggs within the 1-h exposure period. Female G. ashmeadi parasitized on average 4.3 and 3.5 more eggs per hour when presented with 10 and 20 H. vitripennis eggs, respectively, compared with G. tuberculifemur.When5or40H. vitripennis eggs were offered, there was no signiÞcant difference in parasitism rates between parasitoid species. Percentage of female offspring was signiÞcantly higher (5%) for G. ashmeadi compared with G. tuberculifemur. Under the experimental conditions used in these evaluations, results suggest that G. tuberculifemur is an inferior parasitoid to G. ashmeadi.

KEY WORDS biological control, functional response, Gonatocerus ashmeadi, Gonatocerus tuberculifemur, Homalodisca vitripennis

The glassy-winged sharpshooter, Homalodisca vitrip- its northward advance out of southern and central ennis (Germar) (Hemiptera: Cicadellidae: Proconiini California. [formally H. coagulata]), is native to the southeastern In 2001, the California Department of Food and United States and northeastern Mexico. This invasive Agriculture (CDFA) implemented a classical biolog- pest established in California in the late 1980s, where ical control program against H. vitripennis that has it has caused signiÞcant economic problems for pro- involved importing, mass-producing, releasing and es- ducers of agricultural and ornamental commodities. H. tablishing, and evaluating biological control agents for vitripennis is a vector of a xylem-limited bacterium, the suppression of H. vitripennis. To date, a total of six Xylella fastidiosa Wells et al., which causes disease in species of mymarid egg parasitoids (Hymenoptera) a variety of economically important plants including have been released in 13 Californian counties in an PierceÕs disease in grapes, almond leaf scorch, alfalfa attempt to provide effective classical biological con- dwarf, phony peach disease, and oleander leaf scorch trol of H. vitripennis (CDFA 2006). One parasitoid (Blua et al. 1999, UCOP 2000, Varela et al. 2001). species being considered for release and establish- PierceÕs disease is endemic in 28 Californian counties, ment in California is the “new association” (Hokkanen and increased vectoring risk posed by H. vitripennis and Pimentel 1989) biological control agent Gonato- currently threatens grape production that is worth cerus tuberculifemur (Ogloblin) (Mymaridae), a para- $4.14 billion annually (CDFA 2006). The distribution sitoid from Argentina that attacks proconiine sharp- of this pest increased in 2006 and new infestations shooters (the same tribe as H. vitripennis) in South were found in Fresno and Santa Clara Counties America (Leo´n et al. 2008). This parasitoid was im- (CDFA 2006), suggesting H. vitripennis is continuing ported into quarantine in California in 2002 and has been successfully maintained on H. vitripennis egg 1 Department of Entomology, University of California, Riverside, masses with regular infusions of new genetic material CA 92521. from South America. 2 Corresponding author, e-mail: [email protected]. It is argued that new association biological control 3 Department of Statistics, University of California, Riverside, CA agents that have no evolutionary history with the tar- 92521. 4 Center for Invasive Species Research, University of California, get pest should be more effective natural enemies Riverside, CA 92521. because the pest will not be able to adequately defend

0046-225X/09/1634Ð1641$04.00/0 ᭧ 2009 Entomological Society of America December 2009 IRVIN ET AL.: FUNCTIONAL RESPONSE OF Gonatocerus SPP. 1635 itself from novel attacks. However, this strategy has on whether or not to release G. tuberculifemur from been considered risky because natural enemy species quarantine for establishment in California. The results that are most likely to produce effective new associ- of functional response experiments for G. ashmeadi ation biological control are those preadapted to using and G. tuberculifemur are reported here. new hosts and therefore could pose high risk to nontarget species because of this polyphagy (Roderick Materials and Methods 1992). A potentially broad host range for G. tuberculifemur raises concerns about nontarget impacts on Insect Colonies and Laboratory Conditions. Labo- native U.S. proconiine sharpshooters. Environmental ratory colonies of H. vitripennis and G. ashmeadi were concerns could be even greater if G. tuberculifemur maintained at the University of California, at Riverside successfully left California and inÞltrated the home (UCR). Colonies of G. ashmeadi were held at 26 Ϯ 2ЊC range of H. vitripennis, which would put this upper and 30Ð40% RH under a L 14:10 D photoperiod and trophic level organism in sympatry with a variety of reared in cages (50 by 40 by 40 cm) on H. vitripennis other nontarget native proconiine sharpshooter spe- eggs laid on ÔEurekaÕ lemon leaves (Citrus limon L.), cies that G. tuberculifemur could likely use as hosts a preferred lemon variety for H. vitripennis oviposition (e.g., Oncometopia sp.). and parasitoid foraging (Irvin and Hoddle 2004). Cit- The H. vitripennis egg parasitoid guild in California rus limon cultivar Eureka trees, ϳ2 yr old and grafted comprises eight species (CDFA 2006) and is reported to Marcophylla sp. rootstock, were obtained from C & to inßict an average year-round parasitism of 15.5% M Nurseries (Nipomo, CA). Trees were pruned to 60 (Pilkington et al. 2005). Low parasitism rates may be cm in height, potted into 4-liter containers, and fer- because of interference among members of the exist- tilized every 2 wk with Miracle-Gro (20 ml/3.5 liters ing parasitoid guild, which is reducing the effective- of water; Scotts Miracle-Gro Products, Marysville, ness of biological control (Myers et al. 1989, Denoth OH). Female G. tuberculifemur were obtained from G. et al. 2002). Alternatively, low average parasitism may tuberculifemur colonies (clade 1 as described in Leo´n be indicative that an aggressive and efÞcacious natural et al. 2008) maintained in the Insectary and Quaran- enemy that can dominate the guild of H. vitripennis tine Facility at UCR. These colonies had completed egg parasitoids has not been established in California Ϸ75 generations since arriving at UCR in September and is needed for successful biological control. It has 2002 and were augmented periodically with new ma- been proposed that the new association agent, G. terial collected from Argentina. G. tuberculifemur col- tuberculifemur, could be a dominating efÞcacious onies were maintained in ventilated plastic cages (9 by parasitoid species that will provide control of H. vit- 9 by 16 cm) and held at 24 Ϯ 2ЊC and 40Ð50% RH, ripennis in California. However, it is unknown under a L 14:10 D photoperiod. Females were held whether G. tuberculifemur can outperform G. ash- with 50% honey-water for 2Ð3 d before exposure to H. meadi, the dominant H. vitripennis parasitoid in Cal- vitripennis eggs laid on euonymus leaves (Euonymus ifornia, to provide the desired level of control. japonica L.; Celastraceae) or Eureka lemon leaves To measure the efÞcacy of G. tuberculifemur for depending on the source of H. vitripennis eggs. Ap- processing H. vitripennis eggs for comparison to G. proximately 85% of G. tuberculifemur used in this func- ashmeadi, the functional response of G. tuberculifemur tional response study had emerged from egg masses and G. ashmeadi when exposed to different densities laid on Eureka lemon leaves compared with 100% of G. of H. vitripennis eggs was studied. The functional re- ashmeadi. Petioles of leaves with H. vitripennis egg sponse deÞnes the change in the number of hosts masses were inserted into 1-cm slits cut into a piece of parasitized per parasitoid, per unit time, in relation to 0.5-cm-thick polystyrene foam so that leaves had a host density (Royama 1971) and has been used ex- vertical aspect. The foam was cut to Þt tightly into the tensively in population ecology and foraging theory to bottom of the G. tuberculifemur colony cage. The study the potential for natural enemies to regulate host bottom of the parasitoid colony cage was perforated populations (Murdoch and Oaten 1975). The func- with holes and placed in a metal tray (20 by 20 by 45 tional response type (i.e., type I, type II, type III) is cm) containing 2 cm of tap water, which watered the characterized by the shape of the curve that describes foam pad holding leaves. G. ashmeadi and G. tuber- attack rates (which are inßuenced by handling times culifemur colonies were provisioned with honey-wa- and search efÞciencies) of natural enemies over in- ter solution (3:1 natural uncooked honey; Wild Moun- creasing host densities within a Þxed exposure time tain Brand, Oakland, CA) and checked daily for (Juliano 1993). QuantiÞcation of parameters deÞning parasitoid emergence. Newly emerged parasitoids functional response curves for G. tuberculifemur and were collected and prepared for functional response G. ashmeadi may provide insight into which natural experiments. enemy is likely to be more efÞcacious at processing Experimental Procedure for Functional Response varying densities of H. vitripennis eggs, thereby pro- Experiments. Newly emerged (Ϸ24 h) naõ¨ve female viding better biological control of this pest. Conse- and male G. ashmeadi were aspirated into 130-ml plas- quently, functional response parameters could be tic vials (40 dram Plastic Vial; Thornton Plastics, Salt used as metrics to provide relative measures of com- Lake City, UT) and 50% honey-water (natural un- petitiveness, which may help to determine which cooked honey; Wild Mountain Brand) was supplied in parasitoid species is likely to be the superior natural droplets on the lid. This procedure was repeated for G. enemy in the Þeld and to scientiÞcally guide decisions tuberculifemur. Parasitoids were held in the laboratory 1636 ENVIRONMENTAL ENTOMOLOGY Vol. 38, no. 6 for24hat26Ϯ 2ЊC and 30Ð40% RH under a L 14:10 vented successful insect emergence. Therefore, un- D photoperiod before use in functional response ex- emerged eggs were dissected and the numbers of periments. Host densities presented to parasitoids unemerged male and female parasitoid pupae were were 5, 10, 20, and 40 H. vitripennis eggs per female, also recorded. Presence or death of parasitoid eggs and and each experimental egg density was replicated larvae were not determined. Female replicates that 13Ð18 times for each parasitoid species. Host density resulted in zero parasitism (G. ashmeadi ϭ 11% of range and exposure time were selected based on pre- replicates; G. tuberculifemur ϭ 14%) within the ex- vious studies with G. ashmeadi and G. tuberculifemur. perimental hour were labeled “incompetent” and re- The upper host density was chosen because female G. moved from analyses. Unemerged eggs that did not ashmeadi are synovigenic, emerging with Ϸ30 mature contain identiÞable parasitoid pupae were recorded as eggs in their ovaries (Irvin and Hoddle 2008). Pilk- “unemerged H. vitripennis nymphs” and contributed ington and Hoddle (2006) provided individual G. ash- to “nymph mortality.” A set of 12Ð16 controls were set meadi with 40 host eggs per day in their life table up for each experiment host egg density and held at studies, which was surplus to what female parasitoids 26 Ϯ 2ЊC and 30Ð40% RH under L 14:10 D. These vials could use in the 24-h exposure period. Exposure time did not contain a parasitoid and were used to deter- in this study was limited to1htoensure that an upper mine naturally occurring mortality of H. vitripennis limit to the functional response was detected, because eggs under the experimental conditions and exposure studies by N.I. and M.H. (unpublished data) had time. Control mortality was calculated as the propor- shown that, on average, female G. ashmeadi parasitize tion of unemerged H. vitripennis nymphs from the eight eggs within 20 min of detecting a patch. Fur- total number of experimental eggs. thermore, a 1-h exposure time is likely to be more Statistical Analysis. Functional response data for realistic in a Þeld situation compared with a 24-h each species was analyzed in two phases using SAS enclosure with one host patch. Functional response (SAS Institute 1990). First, the shape of the functional experiments were conducted between 1000 and 1300 response curve was determined by logistic regression hours on days when parasitoids and host eggs were of the proportion of H. vitripennis eggs parasitized as available. H. vitripennis eggs laid on Eureka lemon a function of initial density (Trexler et al. 1988). Sec- leaves were Ͻ48 h old, an age shown to be successfully ond, the random predator equation was Þtted to data used by both Gonatocerus spp. (Irvin and Hoddle 2005; after the functional response type was determined N.I. and M.H., unpublished data). N.I. and M.H. (un- (Juliano 1993). The random predator equation ac- published data) found that brochosomes cover 64% of counts for host depletion without replenishment over H. vitripennis egg masses; therefore, in this study, all the course of the experimental period, which was the egg masses were gently wiped with a damp paper design in these experiments. This equation was also towel before experiments to remove any brochosomes used in G. ashmeadi functional response studies con- that could interfere with parasitism attempts (Velema ducted by Chen et al. (2006). et al. 2005), thereby standardizing egg mass quality The polynomial function from Juliano (1993) was within and across treatments. For each replicate, the used to Þt data on the proportion of H. vitripennis eggs petioles of leaves containing egg masses were placed parasitized: through holes drilled through the lid of a 130-ml plastic N exp ͑P ϩ P N ϩ P N 2 ϩ P N 3͒ vial that held water. Leaf number was standardized to e ϭ 0 1 0 2 0 3 0 ϩ ͑ ϩ ϩ 2 ϩ 3͒ [1] four leaves per vial by including lemon leaves without N0 1 exp P0 P1N0 P2N0 P3N0 H. vitripennis egg masses. A second 130-ml plastic vial with ventilation (three 2-cm holes [one on the bot- where Ne/N0 is the probability of being parasitized, Ne tom, and one on each of two sides] covered with mesh is the number of host eggs parasitized, and N0 is the netting [80 ␮m; Jelliff, Southport, CT]) was inverted initial host density. Maximum likelihood estimates of and attached to the lid of the vial holding the water parameters P0, P1, P2, and P3 were obtained by applying and lemon leaves. One mated and honey-water fed logistic regression (Proc Logistic) in SAS (SAS Insti- female parasitoid (Ͻ48 h old) was introduced into tute 1990) to a dichotomous variable that equaled one each vial and left to forage for 1 h, after which leaves for the egg being parasitized or zero if the host egg was containing egg masses exposed to parasitoids were not parasitized. The positive or negative signs of the placed into petri dishes (3.5 by 1 cm; Becton Dickin- linear (P1), quadratic (P2), and cubic (P3) coefÞcients son Labware; Becton Dickinson and Co., Franklin from equation 1 can be used to distinguish the shape Lakes, NJ) lined with moist Þlter paper (4.25 cm; of the functional response curve from experimental Whatman International, Maidstone, United King- data (Juliano 1993). Linear terms not signiÞcantly dom). Petri dishes were labeled with replicate num- different from 0 indicate a type I functional response, ber, density, and species and held at 26 Ϯ 2ЊC and a signiÞcant negative linear coefÞcient indicates a type 30Ð40% RH under a L 14:10 D photoperiod for 3 wk II response, and a signiÞcant positive linear term in- to allow parasitoids and H. vitripennis nymphs to dicates a type III response (Juliano 1993). To deter- emerge. mine the signiÞcance of the linear term from polyno- Experiments were conducted in the laboratory at mial equation 1, the log likelihood-ratio test was used 26 Ϯ 2ЊC and 30Ð40% RH under a L 14:10 D photo- to determine whether equation coefÞcients differed period with ßuorescent lighting. Premature drying of signiÞcantly from 0 (Trexler and Travis 1993). In this leaves sometimes occurred, which occasionally pre- test, the difference in log-likelihoods is a ␹2 with 1 df. December 2009 IRVIN ET AL.: FUNCTIONAL RESPONSE OF Gonatocerus SPP. 1637

ai 16 ii 14 1 12 0.8 10 8 0.6 6 0.4 4 2 0.2

Number of hosts parasitized Number of parasitized hosts 0 0 0 102030400 10203040 Proportion of hosts parasitizedProportion Host Density Host density

bi 16 ii 1 14 12 0.8 10 0.6 8 6 0.4 4 0.2 2 0 0 Number of hosts parasitized parasitized Number of hosts 0102030400 10203040 Proportion of hosts of parasitized Proportion Host density Host density

Fig. 1. Functional responses of (a) G. ashmeadi and (b) G. tuberculifemur at 26ЊC. Mean number (ϮSE) of H. vitripennis eggs parasitized (i) and the mean (ϮSE) proportion of host eggs parasitized (ii).

Where the cubic equation resulted in a nonsigniÞcant parasitized hosts and could therefore engage in self- cubic parameter, in the interest of simplicity, the superparasitism at low host egg densities (Chen et al. model was reduced by eliminating the cubic term and 2006). The signiÞcance of each constant (a and Th) the other parameters were retested (Juliano 1993). was determined using 95% conÞdence intervals Once the functional response type was determined (Juliano 1993). from logistic regression and log likelihood-ratio tests, Gonatocerus ashmeadi showed a type I functional the functional response data were Þtted. For G. tu- response (see Results). To estimate the parameters for berculifemur, the parameters of a type II model were a type I model, piecewise regression (Ryan and Porth estimated using the iterative nonlinear least squares 2007) was used. The upper limit was visually detected regression to Þt the random predator equation 2 (Rog- using Fig. 1ai (see results), and piecewise regression ers 1972): (Ryan and Porth 2007) was performed on the number of eggs parasitized at densities that resulted in para- N ϭ N 1 Ϫ exp[a(T N Ϫ T)] [2] a 0{ h a } sitism below the upper limit (densities Ͻ 13). Linear where Na is the number of parasitized hosts, N0 equals regression was used to conÞrm that the second part of the initial host density, a is the attack constant, Th the type I response (for densities above the upper limit equals the handling time, and T is the total time avail- [densities 13Ð40]) approximated a straight line and able for parasitism. the slope did not differ signiÞcantly from zero. The logistic regression and log likelihood-ratio test If two parasitoid species have a type II functional results suggested that G. ashmeadi demonstrated a response curve, the attack constant (a) and handling type I functional response (see Results). To ensure a time (Th) can be compared using Proc NLIN and type II model was not a more appropriate model for separation of statistically different parameter esti- predicting the functional response of G. ashmeadi, a mates made using 95% conÞdence intervals (Juliano type II functional response was Þtted to the number 1993). However, results derived here suggested that of eggs parasitized at each density using both the G. ashmeadi had a type I response curve, whereas G. random predator equation 2 (Rogers 1972) and the tuberculifemur had a type II response curve (see re- random parasitoid equation 3 (Royama 1971, Kaße et sults). In this case, parameters cannot be compared al. 2005). between species because there is no Th in the type I model, and comparing the attack constant from a type N ϭ N [1 Ϫ exp(ϪaT/1 ϩ aT N )] [3] a 0 h 0 II and slope from a type I has questionable biological It was hypothesized that the random parasitoid equa- signiÞcance (S. A. Juliano, personal communication). tion may be better at detecting a signiÞcant type II Consequently, t-tests were performed in SAS (SAS model, if one existed, because female G. ashmeadi may Institute 1990) to compare the mean number of H. not discriminate between unparasitized and recently vitripennis eggs parasitized by G. ashmeadi and G. 1638 ENVIRONMENTAL ENTOMOLOGY Vol. 38, no. 6

Table 1. Results of logistic regression analyses of the propor- Table 2. Attack rate (a) and handling time (Th) of the functional tion of H. vitripennis eggs parasitized by G. ashmeadi or G. tuber- response of female G. ashmeadi and G. tuberculifemur to densities culifemur compared with the initial host numbers offered in the of H. vitripennis eggs conducted in the laboratory at 26°C laboratory at 26°C Approximate 95% Parameter Estimate Ϯ SE Parameter Estimate Ϯ SE df c2 value P conÞdence intervals G. ashmeadi G. ashmeadi Intercept Ϫ0.235 Ϯ 1.450 1 0.03 0.87 a (hϪ1) 7.217 Ϯ 5.240 Ϫ3.319 17.754 Ϯ Ϯ a Linear 0.474 0.291 1 2.66 0.10 Th (h) 0.078 0.008 0.060 0.095 Quadratic Ϫ0.032 Ϯ 0.016 1 4.26 0.04 G. tuberculifemur Cubic 0.0005 Ϯ 0.0002 1 4.67 0.03 a (hϪ1) 2.280 Ϯ 1.105 0.058 4.500a Ϯ a G. tuberculifemur Th (h) 0.096 0.014 0.069 0.123 Intercept 1.391 Ϯ 0.382 1 13.26 Ͻ0.01 Linear Ϫ0.115 Ϯ 0.036 1 10.37 Ͻ0.01 a Means show signiÞcance at the 0.05 level. Quadratic 0.001 Ϯ 0.001 1 3.40 0.06 Cubic Ñ Ñ Ñ Ñ was signiÞcant for the type I response curve exhibited by G. ashmeadi (t ϭϪ3.33, P Ͻ 0.01). Piecewise regression above the upper host limit of 13 eggs was tuberculifemur for each host egg density to determine conÞrmed to be a straight line (t ϭ 0.25, df ϭ 1, P ϭ whether signiÞcant differences in parasitism between 0.81). These results indicate that the number of H. parasitoid species at each host density existed. One- vitripennis eggs parasitized by G. ashmeadi in1h way analysis of variance (ANOVA) was used to de- increased linearly to a maximum of 11.5 at a density of termine the effect of density on the proportion of 13 hosts and then remained constant (Fig. 1ai). Con- mortality for controls. Finally, percentage of female sequently, it was estimated that, on average, each offspring data was transformed (Box Cox 3) to nor- female G. ashmeadi had the potential to attack a max- malize distribution, and transformed data were imum of 11.5 H. vitripennis eggs over the 1-h period compared between species using t-test in SAS (SAS regardless of whether available host egg densities ex- Institute 1990). Means presented here are back- ceeded 13 hosts. transformed. Functional Response of G. tuberculifemur. Gonato- cerus tuberculifemur exhibited a type II functional response because the linear term of equation 1 was both Results negative and signiÞcantly different from 0, indicating Functional Response Determination for G. ash- that the proportion of H. vitripennis eggs parasitized meadi. Gonatocerus ashmeadi exhibited a type I func- decreased signiÞcantly as egg density increased (Ta- tional response because results from logistic regres- ble 1; Fig. 1bii) (Juliano 1993). The number of H. sion and log likelihood-ratio tests showed that vitripennis eggs attacked and parasitized by G. ash- quadratic and cubic coefÞcients were signiÞcantly dif- meadi increased at a decreasing rate until reaching an ferent from 0, whereas the linear term was not signif- upper plateau with increasing H. vitripennis density icant (Table 1) (Juliano 1993). The proportion of H. (Fig. 1bii). Each female G. tuberculifemur, on average, vitripennis eggs parasitized by G. ashmeadi remained had the potential to attack 9.3 H. vitripennis eggs over constant for host egg densities 5 and 10 and then the experimental 1-h exposure period. The instanta- decreased (Fig. 1aii). This observed trend is consistent neous attack rate (a) and handling time (Th) for G. with a type I functional response (Juliano 1993). The tuberculifemur as estimated by the random predator instantaneous attack rate (a) and handling time (Th) equation was 2.28/h and 0.10 h, respectively, and these for G. ashmeadi as estimated by the random predator parameters were signiÞcantly different from zero (Ta- equation was 7.22/h and 0.08 h, respectively (Table 2). ble 2). Handling time was signiÞcantly different from zero, Comparing Mean Parasitism and Percentage Fe- whereas attack rate (a) was not signiÞcantly different male Offspring Between Parasitoid Species. Compar- from zero. The latter result, a Ϸ 0, conÞrmed that type ing the mean number of H. vitripennis eggs parasitized II is not an appropriate model for describing the func- between species showed that female G. ashmeadi par- tional response of G. ashmeadi under these experi- asitized on average 4.3 and 3.5 more eggs per hour mental conditions. Furthermore, when the random when presented with 10 and 20 eggs, respectively, parasitoid equation (equation 3) was used to estimate compared with G. tuberculifemur (Table 3). When 5 handling time and instantaneous attack rate for G. or 40 H. vitripennis eggs were presented to parasitoids, ashmeadi, the iterative methods failed to converge, there was no signiÞcant difference in mean parasitism and no results were obtained for these parameters rates between species (Table 3). One-way ANOVA with this model, thereby further conÞrming the ap- indicated that there were no signiÞcant differences in propriateness of the type I functional response model. H. vitripennis egg mortality in control vials with vary- Linear regression analyses for host egg densities ing egg densities that lacked exposure to parasitoids Ͻ13 was signiÞcant (y ϭ 0.88x ϩ 0.02, F ϭ 26.90, df ϭ (mean range ϭ 28Ð31%; F ϭ 0.05, df ϭ 3,53, P ϭ 0.99). 1,23, P Ͻ 0.0001, R2 ϭ 0.55), conÞrming that type I is Percentage of female offspring was signiÞcantly 5% the best model for predicting the functional response higher for G. ashmeadi (mean ϭ 84 Ϯ 2%) compared of G. ashmeadi within this range. Piecewise regression with G. tuberculifemur (79 Ϯ 2%; t ϭ 2.14, df ϭ 96, P Ͻ indicated that a breakpoint (upper limit) of 13 hosts 0.05). December 2009 IRVIN ET AL.: FUNCTIONAL RESPONSE OF Gonatocerus SPP. 1639

Table 3. Mean no. ؎ SE of H. vitripennis eggs parasitized by the ability to discriminate between unparasitized and female G. ashmeadi and G. tuberculifemur when offered four dif- hosts previously parasitized by conspeciÞcs. However, ferent host densities in the laboratory at 26°C given that the egg load of newly emerged G. ashmeadi Ϸ Density G. ashmeadi G. tuberculifemur t-valuea df P is 30 eggs (Irvin and Hoddle 2008) and the 10:250 ratio used in Chen et al. (2006) is equivalent to 12.5 5 4.44 Ϯ 0.53 4.07 Ϯ 0.52 0.48 20 0.64 10 8.87 Ϯ 0.57 4.52 Ϯ 0.62 3.44 24 0.01 hosts per female in 24 h, it is conceivable that females 20 11.53 Ϯ 1.01 8.00 Ϯ 1.12 2.33 28 0.03 may have been forced to superparasitize during pro- 40 12.00 Ϯ 1.79 9.29 Ϯ 1.23 1.29 21 0.21 longed exposure to parasitized hosts, even if they possess the ability to discriminate between parasitized a Test statistics reported from t-tests conducted to compare para- and unparasitized hosts. In this study, female G. ash- sitism between species at each density. meadi were caged singly for a short period of time (i.e., 1 h), and it is unknown whether female G. ashmeadi can discriminate between unparasitized hosts and self- Discussion parasitized hosts when given a choice between host Response Curves for Individual Species. Gonato- eggs in either of these two states. cerus tuberculifemur showed a type II functional re- Both G. ashmeadi (type I functional response) and sponse where parasitism increased at a decreasing rate G. tuberculifemur (type II) showed an upper limit to as host density increased, reaching an asymptote that host processing either because of limited numbers of reßected an upper processing limit of 9.3 hosts/h. In eggs for oviposition or they are unable to attack more contrast, G. ashmeadi showed a type I functional re- hosts because of handling time limitations within the sponse where the number of hosts attacked followed 1-h search period. Mymarid egg parasitoids are con- a constant linear increase regardless of host density sidered to be proovigenic where females emerge from below the upper limit of 11.5 hosts processed per hour. hosts with a full complement of mature eggs (Jervis et The main difference separating a type I and type II al. 2001). However, Irvin and Hoddle (2008) recently functional response is the handling time of the natural showed that G. ashmeadi is synovigenic. Dissection of enemy across varying host densities. Handling time female G. ashmeadi immediately on emergence indi- refers to host processing time and includes time spent cates that females have Ϸ30 mature eggs in their ova- searching, antennating, drilling, ovipositing, resting, ries for oviposition (Irvin and Hoddle 2008). This and grooming. For natural enemies showing a type II complement of Ϸ30 mature eggs on birth would sug- response, handling time increases across increasing gest that female G. ashmeadi used in this study only host densities, which reduces the time available for oviposited 60% of their egg load within the experi- searching (Hassell 1978). In contrast, a type I func- mental 1-h time period. Therefore, in this study, fe- tional response implies that, as host densities increase, males may have been time limited, unless self-super- natural enemy handling time is constant and is not parasitism by female G. ashmeadi resulted in the signiÞcantly affected as patch size increases (Hassell oviposition of more eggs into hosts than the number of 1978). In the laboratory at 26ЊC, visual observations of offspring recorded indicated. Chen et al. (2006) re- female G. ashmeadi foraging for H. vitripennis egg ported that superparasitism at low host densities may masses indicated that parasitoids spend Ϸ100Ð300 s cause functional response studies to underestimate ovipositing and preparing for the next oviposition oviposition and attack rates because all of the ovipos- event (N.I. and M.H., unpublished data). The random ited eggs are not accounted for because only one predator equation used in this study estimated han- parasitoid emerges from a superparasitized host egg dling time for female G. ashmeadi as being 281 s, which that contains more than one parasitoid egg. G. ash- is within the observed range for G. ashmeadi under meadi superparasitized 25% of H. vitripennis egg experimental conditions. masses at ratios 10 females:250 H. vitripennis when Mills and Lacan (2004) noted that, for Tri- given a 48-h exposure time (Chen et al. 2006). Al- chogramma sp. (Hymenoptera: Trichogrammatidae) though these ratios are similar to the 1 G. ashmeadi:20 egg parasitoids and the whiteßy parasitoids Eretmo- H. vitripennis eggs and 1:40 ratios used in this study, cerus sp. (Hymenoptera: Aphelinidae) that exhibit superparasitism may be less likely in this study because type I functional responses, the following character- hosts were exposed to parasitoids for only 1 h. istics are shared: (1) these natural enemies search for In contrast to results presented here, Chen et al. hosts by walking, (2) they attack exposed and immo- (2006) described a type II functional response for G. bile hosts, and (3) they can use behavioral mecha- ashmeadi attacking H. vitripennis eggs over a 24-h nisms to minimize superparasitism. Additionally, the period at 22ЊC. However, similar to this study, Chen et parasitoidÕs type I search strategy must be indepen- al. (2006) also reported that the linear coefÞcient dent of host density and therefore unchanged by ex- resulting from Þtting the polynomial function to data perience and physiological status (Mills and Lacan foreggs1dofagewasnotsigniÞcantly different from 2004). G. ashmeadi shares the Þrst two characteristics zero, suggesting that a type II functional response may listed above (N.I., unpublished data). Chen et al. not be the most appropriate model for describing the (2006) showed that superparasitism by G. ashmeadi functional response of G. ashmeadi. Furthermore, occurred at all host densities ranging from ratios of 10 Chen et al. (2006) did not report whether the attack females:10 hosts to 10:250 within a 48-h exposure pe- constants for G. ashmeadi were signiÞcantly different riod and therefore suggested that G. ashmeadi lacked from zero. In light of results obtained in this study, we 1640 ENVIRONMENTAL ENTOMOLOGY Vol. 38, no. 6 suggest that the data presented in Chen et al. (2006) Study of the Þeld host range in the area of origin of G. portrays the linear part of a type I functional response tuberculifemur in Argentina showed that this parasi- and that host egg densities used in experiments were toid successfully parasitized at least Þve species of not high enough to detect the upper processing limit Cicadellini, a tribe to which H. vitripennis does not for this parasitoid when offered a 24-h exposure time. belong (Jones et al. 2005). These laboratory and Þeld In this study, inclusion of a greater range of host results suggest that G. tuberculifemur is polyphagous densities between 0 and the upper limit of 11.5 hosts and may successfully exploit nontarget sharpshooter would have increased the number of data points de- species in the tribes Proconiini and Cicadellini if sus- scribing the attack trajectory as it approached satu- ceptible host species are encountered. ration. Should G. tuberculifemur be established in Califor- Comparing Efﬁciency Between Species. Compar- nia, there is a possibility that G. tuberculifemur could ing the mean number of H. vitripennis eggs parasitized inadvertently spread into areas outside of this state, between parasitoid species showed that G. ashmeadi possibly through interstate commerce of ornamental signiÞcantly outperformed G. tuberculifemur at host plants bearing parasitized H. vitripennis egg masses on densities of 10 and 20 eggs but not at 5 or 40 host eggs, leaves. The signiÞcance of the potential incursion, the even though a higher proportion of eggs were attacked magnitude of foodweb perturbations, and the ecosys- by G. ashmeadi. Female G. ashmeadi also produced a tem impacts posed by G. tuberculifemur to native higher percentage (5%) of female offspring compared sharpshooters and their associated egg parasitoids out- with G. tuberculifemur. Additionally, functional re- side of California is completely unknown but could sponse data indicated that female G. ashmeadi exhib- cause considerable concern for ecologists and taxon- ited an upper oviposition limit of 11.5 host eggs in a 1-h omists studying native ecosystems and documenting search interval, making this species more effective nontarget impacts resulting from biological control than G. tuberculifemur, which showed an upper par- programs. asitism limit of 9.3 host eggs in1h(Ϸ24% fewer eggs In conclusion, this study showed that G. tubercu- attacked compared with G. ashmeadi). Consequently, lifemur failed to outperform the dominant resident H. under the experimental conditions used in these eval- vitripennis parasitoid, G. ashmeadi. Laboratory testing uations, results suggest that G. tuberculifemur is likely studing aspects of host egg age preferences, the com- an inferior parasitoid to G. ashmeadi. Hosts and para- petitive ability of G. tuberculifemur when simulta- sitoids were spatially conÞned in this study, which neously foraging with G. ashmeadi on H. vitripennis removed the parasitoidÕs need to locate the host plant. egg masses in simple and complex environments, and These conditions are highly conducive to parasitism aggressive behavior between these two species when and may not represent the Þeld environment (Legaspi competing for host eggs has been completed and will et al. 1996). The temperature range over which the provide additional information as to whether G. tu- parasitoid must Þnd hosts also affects functional re- berculifemur should be considered a new association sponse (Kaße et al. 2005, Kalyebi et al. 2005, Zamani biological control agent of H. vitripennis. et al. 2006). Furthermore, these functional response studies failed to directly assess patch residency times, egg load, host egg age preference, and aggressive be- Acknowledgments havior, all of which may affect parasitoid efÞcacy and competitiveness. However, completed studies inves- This work was supported in part by the California De- partment of Food and Agriculture (CDFA) PierceÕs Dis- tigating host patch behaviors have shown G. ashmeadi ease-Glassy-Winged Sharpshooter Management Research to be a more aggressive competitor when foraging Program. We thank S. Triapitsyn and V. Berezovskiy, De- concurrently with G. tuberculifemur (Hoddle and partment of Entomology, UCR, for supplying G. tuberculife- Irvin 2007). Aside from these limitations, functional mur for experiments; R. Vega, M. Lewis, H. Kim, and D. Lim response studies may provide a useful Þrst step for for assistance in the Þeld; S. Juliano (Department of Biolog- comparing the efÞciency of different species (Over- ical Science, Illinois State University) for providing valuable holt and Smith 1990). comments on functional response analyses; and the editor of Conclusions on Using G. tuberculifemur as a New Environmental Entomology (J. Ruberson) and the anony- Association Biological Control Agent for H. vitripen- mous reviewers who provided many helpful comments on an earlier draft of this manuscript. nis Control in California. There is substantial uncer- tainty whether G. tuberculifemur could provide signiÞcant additional control of H. vitripennis if References Cited established in California. Furthermore, “new-association” biological control of H. vitripennis with G. tu- Blua, M. J., P. A. Phillips, and R. A. Redak. 1999. A new berculifemur raises concerns about potential un- sharpshooter threatens both crops and ornamentals. wanted impacts on native nontarget species of Calif. 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Molecular characterization of Gonatocerus tuberculife- Received 18 August 2008; accepted 14 August 2009.