The effects of alternative host plant species and plant quality on Dicyphus hesperus populations

Biological Control 100 (2016) 94–100

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The effects of alternative host plant species and plant quality on Dicyphus hesperus populations Lida Nguyen-Dang ⇑, Meghan Vankosky, Sherah VanLaerhoven Department of Biological Sciences, University of Windsor, Windsor, Ontario N9B 3P4, Canada

h i g h l i g h t s  Dicyphus hesperus populations may be affected by plant species and nitrogen content.  Three alternative host plants were paired nitrogen-manipulated tomato plants.  Greater omnivore emergence and population growth occurred in mullein arenas.  Tomato plant nitrogen content only affected D. hesperus in mullein arenas.  Mullein is a superior banker plant, followed by eggplant and finally, pepper.

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Article history: Received 24 February 2016 Revised 27 May 2016 Accepted 29 May 2016 Available online 30 May 2016 Keywords: Banker plant Mullein Biological control Population Omnivory Nitrogen

a b s t r a c t Biological control can be used to defend important crops against insect pests, including those that are insecticide resistant. Dicyphus hesperus Knight (Hemiptera: Miridae) is a generalist zoophytophagous predator and biological control agent of Trialeurodes vaporariorum (Westwood) (Hemiptera: Aleyrodidae), the greenhouse whitefly. Because D. hesperus is an omnivore, the structure of the plant community and the nutritional value of the plants in the release habitat are likely to affect its establishment and population growth. Fifteen adult D. hesperus (ten females, five males) were placed into arenas that contained a tomato plant (Solanum lycopersicum L., Solanales: Solanaceae) of either high or low leaf nitrogen content and one of three alternative host plants: mullein (Verbascum thapsus L., Lamiales: Scrophulariaceae), pepper (Capiscum annuum L., Solanales: Solanaceae), or eggplant (Solanum melongena L., Solanales: Solanaceae). Adults remained in the enclosures for seven days; following their removal, F1 generation nymphs, and subsequently, F1 adults were counted as they emerged and the percent change in population size between generations was calculated. Nymph emergence was affected by the alternative host plant, such that emergence was greatest in arenas with mullein. Tomato nitrogen content only affected nymph emergence in arenas with mullein; more nymphs emerged when low nitrogen tomato was present. The presence of mullein also resulted in larger F1 adult numbers and a greater change in population size between generations compared to eggplant and pepper. Our results indicate that growing tomato and mullein together, regardless of tomato plant nitrogen content, is beneficial to D. hesperus populations. Ó 2016 Elsevier Inc. All rights reserved.

1. Introduction Relative to prey, plant tissue is abundant in agricultural habitats. Although the dietary value of plant foods tends to be less than that of prey foods, many predators include plant foods in their diet (Cohen, 1996). Predators that consume both plant and prey foods during a single lifestage are defined as omnivores (Coll and

⇑ Corresponding author. E-mail addresses: [email protected] (L. Nguyen-Dang), meghan.vankosky@ gmail.com (M. Vankosky), [email protected] (S. VanLaerhoven). http://dx.doi.org/10.1016/j.biocontrol.2016.05.016 1049-9644/Ó 2016 Elsevier Inc. All rights reserved.

Guershon, 2002). Zoophytophagous omnivores are characterized as having a largely prey based diet that is augmented with nutrients (and water) obtained from plant feeding (Coll and Guershon, 2002; Castañé et al., 2011). Plant feeding by zoophytophagous omnivores facilitates their survival and persistence when prey is scarce (Bugg et al., 1987; Eubanks and Denno, 1999; Gillespie et al., 2012). Thus, plant feeding gives zoophytophagous omnivores certain advantages over predators in biological control programs, including early establishment in annual field crops, which is necessary to prevent exponential growth of pest populations (Wiedenmann and Smith, 1997).

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In 1999, work by McGregor et al. demonstrated that the zoophytophagous omnivore Dicyphus hesperus Knight (Hemiptera: Miridae) was an important predator of the greenhouse whitefly (Trialeurodes vaporariorum (Westwood) Diptera: Aleyrodidae) and the twospotted spider mite (Tetranychus urticae Koch, Acari: Tetranychidae). Dicyphus hesperus has been subsequently investigated as a biological control agent in tomato (Solanum lycopersicum L., Solanales: Solanaceae) greenhouses. It is a generalist that consumes a variety of prey species (greenhouse whitefly, twospotted spider mite, green peach aphid [Myzus persicae (Sulzer), Hemiptera: Aphididae)]; McGregor et al., 1999; VanLaerhoven et al., 2006) and plant species, including mullein, Verbascum thapsus L. (Lamiales: Scrophulariaceae), and several species of Solanaceae (Sanchez et al., 2004; VanLaerhoven et al., 2006). Plant feeding serves to provide D. hesperus with water for extraoral digestion, and minerals and nutrients obtained from plant feeding contribute to D. hesperus development, reducing the development time of nymphs by two days (Gillespie and McGregor, 2000). Plant feeding by D. hesperus is generally limited to the stems and leaves of the plant, unless only fruits are available (McGregor et al., 2000), at which time D. hesperus may become a threat to yields of marketable tomato fruits. Female D. hesperus utilize the foliage and stems for oviposition (Gillespie et al., 2004). In the absence of prey, immature D. hesperus may develop to the adult stage, however, reproduction of D. hesperus requires prey feeding unless mullein serves as the host plant (Sanchez et al., 2003, 2004). Not all plant species (or structures) provide a suitable alternative to prey food for D. hesperus. Therefore, host plant species and quality should be considered when designing biological control programs that utilize D. hesperus, in order to maximize pest mortality and minimize damage to tomato fruits. The identity of the crop species and potential alternative host plants is especially important to consider. For example, certain plant species might be more attractive to the predator than the crop plants that require protection (leading to no or reduced prey mortality on the crop plant; Cortesero et al., 2000), or the host plants might be ‘dead end’ hosts that do not contribute to growth of the zoophytophagous omnivore’s population (Eubanks and Denno, 1999). Conversely, some alternative host plants might help to maximize prey consumption by the zoophytophagous omnivore and contribute to its population growth, as mullein does for D. hesperus, even when its prey is absent (Sanchez et al., 2003; Bennett et al., 2009). These ‘banker plants’ that favour the development and survival of biological control agents (Stacey, 1977; Bennison, 1992), are important to include in biological control programs. Although mullein is an acceptable banker plant for D. hesperus in tomato greenhouses (Sanchez et al., 2003; Gabarra et al., 2004), it would be helpful to producers, who tend to be limited by space, if potential banker plants provided a harvestable product in addition to aiding biocontrol programs. Other cultivated Solanaceae may also serve as D. hesperus banker plants. Therefore, we were interested in comparing the potential of eggplant (Solanum melongena L., Solanales: Solanaceae), pepper (Capiscum annuum L., Solanales: Solanaceae), and mullein as banker plants when grown simultaneously with tomato. The size and growth of D. hesperus populations were evaluated to determine the effect of each potential as banker plant in tomato greenhouses. In Canada, 146,792 metric tonnes of tomato, 111,387 metric tonnes of pepper, and 1560 metric tonnes of eggplant were produced in 2014 (Gauthier, 2015). All three are important and valuable food crops characterized in part by alkaloid secondary metabolites including capsaicin (pepper) and solanine (eggplant) (Cowles et al., 1989; Hori et al., 2011). These allelochemicals have phytotoxic (Sun and Wang, 2015) and insecticidal (Cowles et al., 1989; Williams and Mansingh, 1993; Hori et al., 2011) properties and act as deterrents to oviposition and herbivory. Similarly, mullein

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contains iridoid glycosides for defense (Alba et al., 2014). In addition to secondary metabolites, mullein, eggplant, pepper, and tomato possess trichomes (glandular and non-glandular) that serve to protect them against herbivory (Abdula-Roberts et al., 2014; Alba et al., 2014). Pepper plants tend to be less ‘hairy’ than eggplant and tomato; Abdula-Roberts et al. (2014) noted that non-glandular trichomes were concentrated on the leaf veins on the underside of the leaf in the majority of pepper varieties studied. The quality of the plant material available in an agroecosystem may also impact populations of zoophytophagous omnivores (Eubanks and Denno, 2000; Coll, 1996). Plant quality can refer to the nutritional value of the plant, the presence or absence of defensive metabolites in consumed plant structures, and the toughness or palatability of a plant (Awmack and Leather, 2002). Nitrogen is especially important, as it is often limited in the diets of insects that consume plant tissues (Mattson, 1980). Whitefly populations, for example, grow more rapidly when feeding on plants with excess nitrogen compared to plants with a nitrogen deficit (Jauset et al., 2000). Nitrogen content can also impact insect survival. For example, nymphs of the big-eyed bug, Geocoris punctipes (Say) (Hemiptera: Geocoridae), that were reared on high quality bean pods survived until the third instar, whereas nymphs reared on low quality foliage only survived until the second instar (Naranjo and Stimac, 1985). Therefore, in addition to manipulating the alternative host plant available for D. hesperus, we also manipulated the nitrogen content of the tomato plants to assess the impact of plant nitrogen content on D. hesperus nymph and adult populations, as nitrogen fertilizers are often applied to increase tomato yield in greenhouse agroecosystems. In the current experiment, we used emerging nymph and adult populations to assess the impacts of alternative host plants and tomato nitrogen content on D. hesperus. We predicted that larger nymph and adult populations would be observed when tomato plants had high levels of nitrogen in their leaves, similar to the results observed by Vankosky and VanLaerhoven (2016). We also expected that high nitrogen tomato plants would contribute to overall growth of D. hesperus populations over a single generation, as host plants with high nitrogen content are known to contribute to increased population growth of a number of Hemipterans, including the greenhouse whitefly (Jauset et al., 2000). Based on results published by Sanchez et al. (2004), we predicted that larger nymph and adult populations would be observed when mullein served as the alternative host plant, and that mullein would contribute to a greater rate of population growth than eggplant and pepper. Given the physical appearance of eggplant and similarity in leaf hairiness between eggplant and mullein, we also predicted that eggplant would be a better alternative host plant than pepper.

2. Materials and methods 2.1. Colony maintenance and rearing The D. hesperus insect colony originated in California, U.S.A. where insects were collected from white stem hedgenettle, Stachys albens A. Gray (Lamiales: Lamiaceae) (McGregor and Gillespie, 2004; Sparkes, 2012). Dicyphus hesperus were maintained in a colony at the University of Windsor where insects were held in cages in a controlled rearing room (20 ± 5 °C, with a 16:8 h light:dark diurnal cycle, and 50 ± 10% humidity) (Sparkes, 2012). All four D. hesperus instars were reared on Nicotiana tabacum L. (Solanales: Solanaceae) and given Ephestia kuehniella Zeller (Lepidoptera: Pyralidae) eggs as a source of protein (Sparkes, 2012). Dicyphus hesperus adults used in this study were collected from the adult cage of the colony, which housed sexually mature, gravid females that ranged in age from 7 to 10 days.

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2.2. Plant propagation The plants used in this study were grown at the University of Windsor greenhouse in 2013 and 2014. Temperatures and photoperiod in the greenhouse were not controlled. ‘Patio Hybrid’ tomato seeds, ‘PS09941819’ pepper seeds, and ‘Night Shadow’ eggplant seeds (Stokes Seeds Canada, Thorold, Ontario, Canada) and mullein seeds were sown in small black pots filled with soil consisting of 78% peat and 22% perlite (Berger, Saint-Modeste, Quebec, Canada). Seedlings with true leaves were transplanted into clean brown pots filled with the same potting medium after the first true leaves were unfurled. Transplanted tomato plants were fertilized with 100 mL of either low nitrogen (N) or high nitrogen solutions (39 mg L 1 N and 311 mg L 1 N, respectively) daily for seven days and subsequently, on a schedule of alternating days (Jauset et al., 1998; Stout et al., 1998). Tomato plants grown following this protocol in the same greenhouse had different aboveground biomass and different leaf nitrogen content (Vankosky and VanLaerhoven, 2015). The alternative host plants (pepper, eggplant, and mullein) were provided with 100 mL of 15 g L 1 all-purpose plant food every two days for one week (Scotts Miracle-Gro, Marysville, Ohio, USA) following transplantation. All plants were watered daily or as needed to prevent wilting. 2.3. Experimental arenas Experimental arenas were established in enclosures that consisted of a 36  30  14 cm white plastic dishpan as a base to support the plant pots. White landscape fabric was glued to the rim of the dishpan using hot glue so that the fabric could be gathered and fastened above the height of the plants. Wooden dowels (approximately 60 cm long) were placed in the plant pots inside each enclosure to keep the fabric off of the plants. The enclosures were closed using paperclips to prevent D. hesperus from escaping and to prevent whitefly infestation. Enclosures could be easily opened to facilitate insect observation. Plants placed inside the arenas were pruned to achieve approximately equal biomass (between and within species), ranged in height from 10 to 30 cm, and were not flowering nor fruiting (after VanLaerhoven et al., 2006). The eggplant, tomato, and pepper plants were seeded at the same time and were the same age when experimental arenas were set up. Mullein plants were approximately eight to 12 months older as this species is slow growing and requires multiple seasons to reach maturation. Each arena held one tomato plant of either high or low quality, and one alternative host plant (eggplant, pepper, or mullein). As such, there were three levels of alternative host plant (mullein, eggplant, and pepper), and two levels of tomato plant quality, resulting in a total of six treatment groups. Each treatment was replicated four times; dates on which replicates were conducted are given in Table 1. Insofar as it was possible, trials with each alternative host plant and nitrogen content treatment were run simultaneously. However, due to high mortality of mullein seedlings, and of young mullein plants, all replicates with mullein were run simultaneously when mullein plants were available. Ten female and five male D. hesperus, all 7–10 days old, were haphazardly collected using an aspirator and placed inside each arena (15 insects per arena; F0 generation) by placing the aspirator vial equidistant between the tomato plant and alternative host plant and removing the lid. Arenas were sealed after the vials were opened. Dicyphus hesperus adults that died before exiting the vial were replaced on the second day to help reduce variation in adult densities between arenas caused by insect handling or failure to exit the vials. Adults were allowed to mate and oviposit for seven days before being removed from the arenas. The location of the

Table 1 Dates on which replicates of the experiment were completed (H = high nitrogen; L = low nitrogen; E = eggplant; M = mullein; P = pepper); all mullein trials were completed between March and April 2014 when mullein plants were of sufficient size as the development time of mullein was prolonged compared to eggplant and pepper and the mortality of small mullein plants required for these experiments was high. Date

Trials

October 22, 2013–Dec 26, 2013 6 March 2014–20 April 2014 19 April 2014–24 June 2014 15 July 2014–29 Aug 2014

HE1 HE2 LE1 LE2 HP1 HP2 LP1 LP2 HM1 HM2 HM3 HM4 LM1 LM2 LM3 LM4 HP3 HP4 LP3 LP4 HE3 HE4 LE3 LE4

Table 2 Mean (±SE) number of D. hesperus adults observed on the tomato plant, the alternative host plant, or off-plant during the seven day oviposition period and the total number of D. hesperus adults retrieved after seven days. Trial

Location of F0 D. hesperus Alternative host

Tomato

Off-plant

Retrieved on day 7

High Mullein 1 High Mullein 2 High Mullein 3 High Mullein 4 Low Mullein 1 Low Mullein 2 Low Mullein 3 Low Mullein 4

9.0 ± 0.4 8.5 ± 0.4 7.7 ± 0.6 8.2 ± 0.4 8.8 ± 0.4 10.7 ± 0.5 8.3 ± 0.4 8.8 ± 0.6

2.5 ± 0.3 2.8 ± 0.7 3.3 ± 0.4 2.7 ± 0.6 2.2 ± 0.5 2.2 ± 0.4 1.8 ± 0.4 1.5 ± 0.6

0.3 ± 0.3 0.5 ± 0.3 – 0.2 ± 0.2 – 0.3 ± 0.2 0.5 ± 0.2 0.3 ± 0.3

12 11 11 12 11 14 10 11

High Eggplant 1 High Eggplant 2 High Eggplant 3 High Eggplant 4 Low Eggplant 1 Low Eggplant 2 Low Eggplant 3 Low Eggplant 4

6.7 ± 1.8 7.2 ± 0.8 7.2 ± 1.0 9.0 ± 1.3 8.3 ± 1.0 8.3 ± 0.8 6.7 ± 1.0 8.7 ± 0.8

2.8 ± 0.8 2.7 ± 0.8 4.8 ± 1.5 2.5 ± 0.5 0.8 ± 0.8 1.0 ± 0.9 4.5 ± 0.8 2.7 ± 1.9

0.7 ± 0.8 0.5 ± 0.8 0.2 ± 0.4 0.5 ± 0.8 0.5 ± 0.8 – 0.2 ± 0.4 –

10 11 12 13 9 10 11 10

High Pepper 1 High Pepper 2 High Pepper 3 High Pepper 4 Low Pepper 1 Low Pepper 2 Low Pepper 3 Low Pepper 4

1.0 ± 0.3 0.7 ± 0.5 0.5 ± 0.3 0.5 ± 0.2 0.3 ± 0.2 1.3 ± 0.3 1.0 ± 0.5 1.5 ± 0.4

8.5 ± 0.6 8.2 ± 0.5 8.7 ± 0.5 8.8 ± 0.5 10.0 ± 0.5 9.0 ± 0.5 7.8 ± 0.5 8.3 ± 0.3

0.7 ± 0.3 0.8 ± 0.3 1.5 ± 0.7 0.7 ± 0.3 0.8 ± 0.3 – 0.7 ± 0.3 0.8 ± 0.3

10 9 12 10 11 11 9 10

adult D. hesperus in each arena was recorded every day during the 7 d oviposition period (Table 2), and the number of surviving adults at the end of the oviposition period was recorded (Table 2). All observations were made by the same observer to reduce experimental error. 2.4. Data collection Every second day, the F1 nymphs and adults were counted as they emerged. The date that nymphs of the F1 generation hatched was noted as well as the date of adult emergence. The total population of nymphs was defined as the maximum number of nymphs observed on a single day during the observation period. The adult F1 population was similarly defined based on emergence of adults in the enclosure. From this data, the change in population between the F0 and F1 populations was calculated using: D = (F1 F0)/F0, where positive D-values represent population growth and negative values represent population decline over the generation studied herein. A data-logger (Smartbutton, ACR Systems Inc., Surrey, British Columbia, Canada) was kept in the greenhouse to keep track of the temperature throughout the duration of the experiment. Mean, maximum, and minimum temperatures recorded in the greenhouse in the months when trials were run are provided in Table 3.

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October 2013 November 2013 December 2013 March 2014 April 2014 May 2014 June 2014 July 2014 August 2014

Temperature (°C) Minimum

Maximum

Mean

11.0 15.5 17.0 12.5 7.5 8.5 12.0 15.0 18.0

36.0 34.5 26.0 54.5 57.0 60.0 53.0 36.0 37.5

22.84 24.34 21.90 26.69 26.48 26.76 26.98 24.66 24.55

2.5. Statistical analysis A two-way analysis of variance (ANOVA) was performed using the GLM procedure (SAS Institute, 2009) to determine the effect of tomato plant quality (two levels: high N and low N), alternative host plant (three levels: mullein, pepper, and eggplant), and the interaction between the two factors on nymph and adult populations. Adult emergence data was transformed using the natural log transformation (LN); raw data is reported. Significant interaction effects were further analyzed using the SLICE command in the GLM procedure (SAS Institute, 2009) to ask: what is the effect of alternative host plant in arenas with low quality tomato and in arenas with high quality tomato? Where no interaction effects were observed, significant effects of the main factors (plant quality, alternative host plant) were assessed using the PDIFF function (SAS Institute, 2009). For all tests, a = 0.05; to prevent Type I Errors, a was adjusted using the Bonferroni correction when performing multiple comparisons. To assess effects of plant quality and alternative hosts on the change in population size (D) observed in the experiment, a nonparametric Kruskal-Wallis test was performed using the NPAR1WAY procedure (SAS Institute, 2009), as this data set could not be transformed to meet the assumptions of parametric tests. Separate Kruskal-Wallis tests were used to determine a) the effect of tomato plant quality, and b) the effect of alternative host plant on population change between generations; for both a = 0.025. Significant Kruskal-Wallis test results were followed up with pairedsample Mann-Whitney U tests to determine differences between alternative host plants or tomato plant quality treatments, with a adjusted using the Bonferroni correction (SAS Institute, 2009).

Mean (±SE) Number of Nymphs

Month

80 70

60

A

50

Pepper

40 B

30

Eggplant

a a

Mullein

20 C

10

b

0 Low N High N Tomato Plant Quality Fig. 1. The interaction of alternative host plant (mullein, eggplant, pepper) and tomato plant quality (low vs. high nitrogen) on the mean number (±SE) of F1 Dicyphus hesperus nymphs. Means with the same capitalized letters or the same lowercase letters are not significantly different (p > 0.05).

arenas with eggplant or mullein (Fig. 2). Tomato plant quality did not affect the size of D. hesperus F1 adult populations (F1,18 = 3.08, p = 0.0963), and the interaction between tomato plant quality and alternative host plant was not significant (F1,18 = 0.50, p = 0.6151). The change in D. hesperus population size between the two generations studied here was not affected by tomato plant quality 20 Number of Adult D. hesperus

Table 3 Monthly minimum, maximum and mean temperatures in the greenhouse during the experiment.

A A

15

10

5 B 0 Mullein

Eggplant

Pepper

Alternative Host Plant Fig. 2. The mean (±SE) number of Dicyphus hesperus that survived to adulthood on the three alternative host plants. Means with the same letters are not significantly different (p > 0.05).

3. Results 20 Percent Population Growth

Tomato plant quality and alternative host plant interacted to affect the size of the F1 nymph population (F2,18 = 5.41, p = 0.0145; Fig. 1). In arenas with mullein as the alternative host, nitrogen content of the tomato plants affected nymph emergence (F1,18 = 19.55, p = 0.0003; Fig. 1); tomato plant quality had no effect in arenas with either pepper (F1,18 = 0.00, p = 0.9698; Fig. 1) or eggplant (F1,18 = 0.78, p = 0.3882; Fig. 1) as the alternative host. In arenas with low nitrogen tomato plants, different numbers of nymphs emerged in arenas when the alternative host plant differed (F = 29.73, df = 2, p < 0.0001; Fig. 1). Alternative host plant also affected nymph emergence in arenas with high nitrogen tomato (F = 5.94, df = 2, p 0.0104; Fig. 1). Alternative host plant affected the size of the F1 adult D. hesperus population (F2,18 = 17.37, p < 0.0001). The largest F1 adult populations were observed in arenas with mullein, followed by eggplant and then pepper; arenas that included pepper as the alternative host plant had significantly smaller populations than

A

AB

B

Mullein

Eggplant

Pepper

0 -20 -40 -60 -80 -100 -120

Alternative Host Plant

Fig. 3. The mean (±SE) change in population size of Dicyphus hesperus reared in arenas with three alternative host plants. Means with the same letters are not significantly different (p > 0.025).

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(v2 = 1.4094, df = 1, p = 0.2352), but was affected by the identity of the alternative host plant (v2 = 12.0068, df = 2, p = 0.0025). In arenas with mullein, the mean D-value was positive, indicating population growth (Fig. 3). In comparison, arenas containing eggplant and pepper had negative D-values, indicating an overall population decline (Fig. 3).

4. Discussion Zoophytophagous omnivores, including D. hesperus, benefit from the availability of host plants and banker plants in agroecosystems, as these plants provide oviposition substrate, water for extraoral digestion, and additional nutrients that contribute to their development and survival (Gillespie and McGregor, 2000; Sanchez et al., 2003, 2004; Maleki et al., 2006; Castañé et al., 2011). Variation between different species of host plant, such as the presence and type of secondary metabolite defenses, morphology (i.e. leaf thickness, presence of trichomes), the timing of fruit production, and abundance of pollen may make certain plant species more beneficial than others to zoophytophagous insects (Coll, 1998). Therefore, we predicted that D. hesperus populations would be influenced by the species of alternative host plant provided as a banker plant in test arenas. Because forms of nitrogen that can be utilized by animals are limited in many habitats (Mattson, 1980; Denno and Roderick, 1990), we also predicted that the nitrogen content of tomato plants would affect populations of D. hesperus in the absence of prey when a primary host plant (tomato) and a banker plant or alternative host (eggplant, mullein, or pepper) were provided. We observed that the nitrogen content of tomato plants did affect nymph populations, but that this effect depended on the identity of the banker plant. We did not observe any effect of tomato plant nitrogen content on adult D. hesperus populations, or on the change in population size between the two generations included in this experiment. When mullein served as the banker plant, we observed that the number of nymphs that emerged in arenas with low nitrogen tomato plants was greater than in arenas with high nitrogen tomato plants. This observation is contrary to that made by Vankosky and VanLaerhoven (2016), who found that tomato plants with high nitrogen content were preferred oviposition hosts of D. hesperus. One possible scenario that might explain this result is that in arenas with low nitrogen tomato, the vast majority of eggs were laid on mullein rather than tomato and thus, egg and nymph mortality was negligible because mullein is known to support egg and nymph development and survival (Sanchez et al., 2003). In arenas with high nitrogen tomato, we expect that approximately equal numbers of eggs were laid on both the mullein and tomato plant, as high nitrogen tomato is preferred relative to low nitrogen tomato (Vankosky and VanLaerhoven, 2016). However, tomato is not known to support egg and nymph survival, so egg and nymph mortality was likely greater in these test arenas than in arenas with low nitrogen tomato, contributing to our results. This would make sense, as slightly higher numbers of adults were found on high nitrogen tomato plants compared to low nitrogen tomato plants, but the vast majority of adults were consistently found on mullein. The fact that plants with excess nitrogen might possess greater concentrations of defensive metabolites, including alkaloids, which could deter females from ovipositing or result in nymph mortality at the egg or early instar stages (Awmack and Leather, 2002) might have also contributed to our observed results. Closer observation of female D. hesperus to evaluate preference between the tomato plants and mullein plants would help us to further understand our results. For example, by evaluating female preference for high and low nitrogen tomato in the presence of mullein, we could assess whether or not females select oviposition

host based on their own needs, or those of their offspring. This disconnect between female choice and offspring performance is referred to as optimal bad motherhood and has been described for a few herbivorous insects (Scheirs et al., 2000; Mayhew, 2001). If this is the case for D. hesperus, our results suggest that nitrogen rich plants may be beneficial for the female, but not for her offspring, perhaps due to high levels of nitrogenous defensive compounds that killed plant-feeding nymphs. We did not observe differences in the number of adult D. hesperus that emerged in communities with low and high nitrogen tomato plants. This was contrary to our initial prediction; however, it has since been observed that tomato nitrogen content does not provide a strong developmental advantage to D. hesperus nymphs (Vankosky and VanLaerhoven, 2016). In light of these findings, our results make more sense. Other considerations to keep in mind are that high nitrogen host plants might only provide an advantage early in development, especially if nymphs are sensitive to plant alkaloid levels and prey scarcity forces continued plant feeding, as in this experiment. Dicyphus hesperus are also opportunistic cannibals (Laycock et al., 2006), so some cannibalism may have occurred in the arenas that our design was unable to account for. Experiments designed to assess the mortality of D. hesperus nymphs at each instar, and observe potential cannibalism events, would be helpful in clarifying our results. Our second prediction was that nymph and adult populations would be larger when mullein was the banker plant, followed by eggplant, and lastly pepper. Our results agreed with our prediction, as plant communities with pepper had the smallest F1 nymph and adult populations. In comparison, mullein had a positive effect on D. hesperus populations in our experiments, hence, mullein is likely to help this zoophytophagous omnivore to quickly establish populations for biological control, as other authors, including Sanchez et al. (2003, 2004) have observed. Communities with eggplant supported intermediate numbers of D. hesperus; its potential as a banker plant is not as great as mullein, but eggplant may provide some benefits to D. hesperus, while also providing producers with yield from a second crop. Experiments on a larger scale (i.e. in greenhouses) using eggplant should be conducted to determine the effect of D. hesperus on whitefly populations and to assess the economic aspects of growing mixed crops of tomato and eggplant in greenhouses. Differences in the physical characteristics between mullein and eggplant (the more suitable hosts) and pepper (the least suitable host) likely contributed to our results. The presence or absence of certain plant structures, particularly trichomes, can play a key role in the formation of insect communities and in determining the nature of interactions between insect species, and between insects and their host plant (Price et al., 1980; Coll, 1998; Peeters, 2002). Both mullein and eggplant possess many trichomes. Trichomes and epicuticular waxes often act as deterrents or defenses against herbivorous insects by making it difficult for insects to manoeuvre (Smith, 2005). Although the effects of trichomes on the locomotor ability of D. hesperus has not been specifically assessed, the closely related Dicyphus errans Wolff (Hemiptera: Miridae) has long slender legs and curved tarsal claws that allow it to navigate and efficiently capture prey on hairy plant surfaces (Voigt et al., 2007). Dicyphus hesperus likely has similar morphological characteristics that make plants with trichomes suitable hosts. The fact that F0 females avoided pepper plants, which are relatively free of trichomes, in the test arenas supports this assumption. Additionally, Laycock et al. (2006) observed that starved female D. hesperus were more likely to cannibalize first instar nymphs on unshaved mullein than on shaved mullein, suggesting that the trichomes on the mullein facilitate either nymph avoidance of conspecific predators or the predation efficiency of adults D. hesperus for heterospecific prey. Overall, based on our results and those of Sanchez et al.

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(2003, 2004), we conclude that the abundance of trichomes on mullein (and to a lesser extent, eggplant) probably facilitates the survival and reproduction of D. hesperus by improving their locomotor ability (Voigt et al., 2007), and subsequently, their predation efficiency. Other physical and chemical characteristics of the alternative host plants may have also contributed to the difference in nymph and adult populations that we observed. For example, eggplant and mullein have larger leaves than pepper plants that might provide additional microclimates for D. hesperus nymphs, or additional area for oviposition. We seldom observed individuals from either the F0 or F1 adult generation on the pepper plants during this study, which may be due to chemical deterrents, such as capsaicin, found in pepper tissues (Cowles et al., 1989; Hori et al., 2011). The deterrent effect of pepper extracts on insect herbivores is well documented (i.e. Cowles et al., 1989; Dekebo et al., 2007; Hori et al., 2011). We suspect that F0 D. hesperus were deterred from ovipositing on pepper leaves due to chemical deterrents possessed by pepper plants, contributing to reduced overall nymph populations in pepper arenas. Sanchez et al. (2004) observed that D. hesperus rarely laid eggs on pepper plants, which supports our conclusions. Based on our results, we do not suggest using pepper as a banker plant in tomato greenhouses. Eggplant and mullein are more likely to benefit D. hesperus populations than pepper plants. 5. Conclusions and future directions In the current study, tomato plant nitrogen content did not impact the emergence of F1 generation adults, as expected. Therefore, we do not believe that tomato plant nitrogen content should be a primary concern of producers when designing biological control programs using D. hesperus. It may be of interest to repeat the experiment with prey provided for developing nymphs and foraging adults in order to determine if tomato plant nutrition contributes to D. hesperus development and survival when prey foods are available for consumption. In agroecosystems where crop plants generally occur in monocultures, the presence of alternative host plants can provide important resources or habitat for biological control agents (Wheeler and Krimmel, 2015). In the current experiment, with no prey present in test arenas, only mullein contributed to the growth of D. hesperus populations in the absence of prey. Therefore, our results support the use of mullein as a banker plant (Sanchez et al., 2004; Lambert et al., 2005). Eggplant might also prove to be an effective banker plant, but this should be investigated in arenas where prey is present and over multiple generations. Unlike mullein, eggplant could provide a second profitable crop in tomato greenhouses, and for this reason, it is important to investigate further. Acknowledgments We would like to thank Vincenzo Pacheco and Krystal Hans for their editorial comments, assistance, and support throughout this project. We acknowledge our sources of funding: the University of Windsor, the Ontario Innovation Trust, the Early Researcher Award from the Ontario Ministry of Research and Innovation and the Canada Foundation for Innovation. References Abdula-Roberts, L., Berny-Mieryteran, J.C., Mooney, K.A., Moguel-Ordonez, Y.B., TutPech, F., 2014. Plant traits mediate effects of predators across pepper (Capsicum annuum) varieties. Ecol. Entomol. 39, 361–370. Alba, C., Bowers, M.D., Blumenthal, D., Hufbauer, R.A., 2014. Chemical and mechanical defenses vary among maternal lines and leaf ages in Verbascum thapsus L. (Scrophulariaceae) and reduce platability to a generalist insect. PLoS One. http://dx.doi.org/10.1371/journal.pone.0104889.

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