DOI: 10.1111/eea.12325 Feeding location affects demographic performance of on winter canola Ximena Cibils-Stewart1, Brett K. Sandercock2 & Brian P.McCornack1* 1Department of Entomology, Kansas State University, 123 W. Waters Hall, Manhattan, KS 66506, USA, and 2Division of Biology, Kansas State University, 116 Ackert Hall, Manhattan, KS 66506, USA Accepted: 21 April 2015

Key words: finite rate of population change k, life-table response experiment, plant architecture, population growth, prospective demographic analysis, retrospective demographic analysis, brassicae, , , napus, , stage-structured population model

Abstract The cabbage , Brevicoryne brassicae L. (Hemiptera: Aphididae), is a perennial pest that special- izes on plants of the Brassicaceae , attacking winter canola (Brassica napus L.) mainly during and after flowering. Under field conditions, cabbage aphid colonizes the upper flowering canopy. Population dynamics of aphids in the flowering canopy could be regulated by differences in either plant quality (bottom-up) or predatory (top-down) forces. The goal of our study was to determine the effect of feeding location on cabbage aphid demography. A stage-structured matrix population model was constructed for aphids restricted to reproductive or vegetative plant tissues of canola. We found that feeding location had a large impact on demography of cabbage aphid; the finite rate of increase (k SEM) was higher when aphids were restricted to reproductive tissues, compared to aphids feeding on vegetative tissues: 1.25 0.01 vs. 1.17 0.01 (leaves). Aphids confined to repro- ductive tissues with higher k exhibited shorter generation times (T = 14.2 0.2 days) and 53–75%

higher net reproductive rates (R0 = 23.3 1.7) than aphids feeding on vegetative tissues. Prospec- tive analyses showed that there was a nymph-skewed stable stage distribution, and elasticity values revealed that k is most sensitive to changes in stasis of adults staying in the adult stage and to adult survival. Retrospective analyses indicated that variation in adult fecundity (value of 0.05) had the largest effect on population dynamics but collectively, growth of nymphal stage 2–3, 3–4, and 4 to adult accounted for most of the difference in k between the treatments. Monitoring programs should target adults and penultimate instars colonizing reproductive tissues of canola plants in the field as aphids on these plant structures contribute most to population growth.

green peach aphid, Myzus persicae (Sulzer), and cabbage Introduction aphid, Brevicoryne brassicae (L.) (all Hemiptera: Aphidi- Winter canola, Brassica napus L. (Brassicaceae), is a profit- dae) (Franke et al., 2009; Boyles et al., 2012). When able biofuel crop that has increased in acreage in South aphids form dense colonies on developing flowers, yield Central USA since the introduction of cold-tolerant varie- losses of up to 70% have been reported if infestations are ties. New varieties have allowed growers to rotate canola left untreated (Boyles et al., 2012). After adoption of seed- with winter wheat, Triticum aestivum L. (Poaceae), the applied insecticides for managing early-season turnip most abundant crop of this region (Franke et al., 2009; aphids, cabbage aphid has become the most damaging Ash, 2012). Since its introduction as a new crop, winter aphid species colonizing winter canola under mixed canola has been attacked by a complex of aphid species, aphid-species infestations (Boyles et al., 2012). including turnip aphid, (Kaltenbach), Cabbage aphid is a herbivorous perennial pest restricted to members of the Brassicaceae and attacks canola mainly during early flowering and pod development (Boyles et al., *Correspondence: Brian P. McCornack, Department of Entomology, 2012). Cabbage aphid develops through four nymphal in- Kansas State University, 123 W. Water Hall, Manhattan, KS 66506, stars before reaching physiological maturity and starting USA. E-mail: [email protected] parthenogenetic reproduction (Hughes, 1963). In canola,

© 2015 The Authors. Entomologia Experimentalis et Applicata is published by John Wiley & Sons Ltd on behalf of Netherlands Entomological Society. Entomologia Experimentalis et Applicata 156: 149–159, 2015 149 This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made. 150 Cibils-Stewart et al. established populations are mainly composed of apterous Materials and methods females, but sexual and alatoid individuals are produced when populations reach high densities. Under asexual and Plant and aphid materials sexual reproduction scenarios, the fitness of newly depos- Canola plants (variety Riley; Kansas Foundation Seed ited apterous nymphs is dependent on deposition sites Service, Manhattan, KS, USA) were seeded in a special selected by their fundatrix mothers on acceptable host soil mixture that contained all required minerals and plants. In the field, cabbage aphid is predominantly nutrients and maintained in the greenhouse with a L16: observed colonizing the top flowering canopy of the D8 photoperiod for ca. 6 weeks (Johnson-Flanagan & canola plant (Hopkins et al., 2009). Occurrence of arthro- Spencer, 1994). Soil and photoperiod provided adequate pod herbivores in different plant structures is a common growth for canola plants to survive the vernalization phenomenon in plant-herbivore interactions. Little is process (M Stamm, canola breeder, Kansas State Univer- known about the consequence of reproductive cabbage sity, pers. comm.). The soil mixture consisted of peat aphid adults depositing nymphs on suboptimal plant moss, perlite, gypsum, slow-release fertilizer, and other structures. Reproductive structures of the canola plant micronutrients (proprietary blend, M Stamm). Canola may provide optimal resources for cabbage aphid growth plants were then artificially vernalized in a growth cham- and development and therefore influence aphid demo- ber for ca. 2 months at L12:D12 photoperiod and con- graphic performance (Smallegange et al., 2007; Malik stant 4 °C to induce reproductive maturity (Murphy & et al., 2010). Scarth, 1994). Vernalized canola plants were watered Feeding location has been reported to influence both fit- daily and used to maintain aphid colonies. Cabbage ness as well as natural-enemy recruitment for herbivorous aphids used in all experiments were obtained from field in other cultivated plant systems including collections but maintained in laboratory-reared colonies chili (Idris & Roff, 2002), cotton (Griogolli et al., 2013), at Kansas State University (KSU) in the Department of alfalfa (Berberet et al., 2009), tobacco (Athanassiou et al., Entomology (Manhattan, KS, USA); aphid species iden- 2005; Kavallieratos et al., 2005), sunflowers (Pekar, 2005), tification was confirmed using alates collected from and other brassica crops, including Brassica nigra (L.) founding populations (Blackman & Eastop, 2000). In W.D.J. Koch (Smallegange et al., 2007). Conversely, feed- April 2011, adult cabbage aphids were collected from ing habits of natural enemies can also have disproportion- winter canola fields (37°1.35396N, 98°22.01688W and ate effects on aphid population growth at locations where 36°59.79996N, 98°29.03508W) in Barber County, Kan- natural enemies are feeding, which has been documented sas, transported to the laboratory in coolers, and trans- for soybean aphid in soybean (Costamagna et al., 2013). ferred to vernalized canola plants. Transfer of aphids to In canola, key top-down and bottom-up forces governing new plants consisted of excising an aphid-infested leaf cabbage aphid population dynamics during early flower- and placing it atop of new leaves from vernalized canola ing and other critical plant developmental stages where plants to allow independent aphid movement to fresh management is most effective (early flowering) are not plant tissue. Using procedures described by Kos et al. well-understood. (2011a), colonies were maintained on vernalized canola The objectives of this study were to use a prospective plants at 22 2 °C, 60–70% r.h., and L16:D8 photope- demographic analysis and a retrospective life table riod. To maintain colony vigor, apterous aphids were response experiment (LTRE) to determine the extent to transferred to non-infested, vernalized canola plants which feeding location affects aphid vital rates (survival, every other week. Voucher specimens (B. brassicae growth, reproduction), and population dynamics. Demo- nymphs and adults) were deposited in the KSU Museum graphic models based on prospective sensitivity analyses of Entomological and Prairie Research (vou- were used to quantify the potential effects of feeding loca- cher number 228). tion on aphid vital rates (reproduction, growth, and sur- vival) and future contributions to population growth rate Aphid population dynamics or k (Caswell, 1996, 2001). We then used a retrospective An exclusion experiment using mesh enclosure cages LTRE to decompose the contributions of different vital was replicated in field and greenhouse trials to assess the rates to variation in k among the experimental treatments. impact of feeding location on aphid population growth We hypothesized that reproductive tissues of the canola rates in the absence of predation. One trial was setup in plant would support higher population sizes of cabbage production canola fields at Ashland Bottoms Research aphid than vegetative tissues. Demographic population Farm near Manhattan, KS, from 27 April to 18 May analyses are useful to predict pest outbreaks and dynamics 2011, and then repeated under controlled greenhouse of colonization in crops (Kasap & Alten, 2006). conditions (22 2 °C, 60–70% r.h., and L16:D8 Feeding location affects aphid demography 151 photoperiod) at KSU (Manhattan, KS) from 25 October the mesh from resting on the flowers or the leaves or to 15 November 2011. Apterous cabbage aphids were disrupting growing aphid populations. In companion restricted to either the upper flowering canopy or a sin- experiments, Cibils-Stewart (2013) showed a lack of cage gle leaf in the lower canola canopy using enclosure cages effect under field and greenhouse conditions across mul- (23 cm diameter and 71 cm long; Figure 1A) adapted tiple years using the same cage design. In particular, from Soper et al. (2013). Cages were placed on the same cumulative degree-days were the same for aphids plant to enable direct comparison of plant location on enclosed on reproductive and vegetative plant structures. aphid population growth, and to control for potential A total of 18 (n = 36) and 30 (n = 60) cages per treat- effects among individual plants in size or nutritional ment were deployed in the field and greenhouse trials, composition. Each cage consisted of white, no-see-um respectively. mesh (Quest Outfitters, Sarasota, FL, USA) with zip- Populations in each enclosure cage were started with pered tops. Zippers provided access to either the flower- five newly reproductive, apterous adult cabbage aphids ing raceme or the vegetative leaf after a cage was secured that were transported to field and greenhouse trials using to a plant structure. The base of each enclosure cage was 2-ml Eppendorf vials (Fisher Scientific, Waltham, MA, secured to the canola plant using 15-cm plastic cable-ties USA). Aphids were transferred to the experimental plants (Gardner Bender, Butler, WI, USA), which were located with fine, camel hair paintbrushes and placed directly on below the last flower of the flowering raceme, or at the the canola flowers or leaves. Aphid populations remained node between the leaf and the plant stem. To allow free on caged sections of the plant for 3 weeks. Plants within a movement of aphids within each cage, we added cylin- trial were arranged in a completely randomized design. At drical supports made of 14-gauge, galvanized steel wire the end of each trial, the base of each canola plant was (Impex Systems Group, Miami, FL, USA). Supports kept excised and all plant material and attached cages were placed in a 7.6-l (2 gallons) plastic bag, which was then immediately stored in a freezer at 20 °C. Freezing stopped aphid development and nymphal deposition and AB allowed for effective counting of cabbage aphid popula- tions at the end of each trial. Plant structures within enclo- sure cages were removed and all aphids within each cage were counted in the laboratory using a magnifying glass. For the greenhouse trial, aphids were categorized into three morphs based on absence or presence of wing pads and wings (Blackman & Eastop, 2000). The three morphs included apterous nymphs and adults, alatoid nymphs, and alates. C The finite rate of population change was calculated for

the entire 21-day trial (k21)asaratioofchangeinaphid densities at the start (N0)andend(N21) of the field or greenhouse trials, where k21 = N21/N0. To compare results between populations and individual aphid experiments,

we then calculated the daily rate of change (kd), where 1/21 kd = (N21/N0) . After calculating the finite rate of pop- ulation growth for each cage location, Student’s paired t-test (Proc TTEST; SAS 2009; SAS Institute, Cary, NC, USA) was used to compare aphid populations restricted to either the top (reproductive) or bottom (vegetative) plant structures in both trials (greenhouse and field). Lastly, aphid morphs (apterous nymphs and adults, alatoid nymphs, and alates) from the greenhouse trial were com- pared between locations using a Student’s paired t-test a = Figure 1 Pictures depicting enclosure cages deployed to (A) (Proc TTEST; SAS 2009) at 0.05. Population structure restrict aphid populations to either reproductive (R) or vegetative of aphids restricted to different plant structures was com- 2 (V) structures, and restrict individual aphids to either (B) pared using a Pearson’s v test for homogeneity between vegetative or (C) reproductive plant tissues. feeding locations. 152 Cibils-Stewart et al.

Individual aphid demographics Following methods of Chaplin-Kramer et al. (2011), we To model effects of bottom-up forces on individual cab- kept nymphs together until they reached their penulti- bage aphid, small enclosure cages were used to measure mate stage to reduce the likelihood of nymphs escaping vital rates (growth, survival, and reproduction) of aphids individual cages. Only Gen2 nymphs were used to esti- feeding on apical reproductive tissues or basal vegetative mate demographic attributes for individual aphids in tissues of a canola plant. Trials were conducted under this experiment. Nymphal development (instar changes), greenhouse conditions (22 2 °C, 60–70% r.h., and L16: adult fecundity, and survival of each Gen2 aphid were D8 photoperiod) at KSU from 23 September to 28 Octo- recorded daily for the entire lifespan of each aphid. ber 2012. In this experiment, individual aphids were ran- Nymphal changes were determined by counting the domly assigned to one of three fixed locations on canola aphid exuvia (exoskeleton molts), which were removed plants: (1) flowering structure (reproductive tissue), or (2) and recorded daily; the number of exuvia was correlated top or (3) bottom surfaces of a single leaf (vegetative with nymphal body size to estimate the proportion of tissues) from the mid-canopy of a canola plant. Enclosure nymphs at a given instar or age. Once Gen2 nymphs cages (Converters, Huntingdon Valley, PA, USA) followed reached the penultimate instar, they were transferred to the design of Nagaraj et al. (2005) and consisted of a individual cages and responses were individually tracked 0.5-cm thick foam rectangle (outside rectangle dimen- (n = 50 aphids per treatment location) for the remain- sions: 6.2 9 3.6 cm; inside rectangle dimensions: der of the experiment (n = 150 total cages). For repro- 5.1 9 2.5 cm) with manufacturer-applied adhesive on ductive Gen2 adults, the number of third generation cage tops and bottoms. For cages placed on leaves, pre- (Gen3) nymphs produced by each reproductive adult applied adhesive was then used to secure no-see-um mesh was recorded and newly deposited Gen3 nymphs were on one side of the cage. Mesh kept aphids from escaping, removed daily to determine fecundity (total number of allowed for adequate ventilation, and facilitated repeated nymphs produced per adult) and reproductive rates counting of aphid populations. The remaining adhesive (mean nymphs produced per day per adult). side was secured to the leave on the mid-canopy leaf loca- The life cycle of individual cabbage aphids was catego- tions to enclose the aphids (Figure 1B). For aphids rized into six stage-classes (Figure 2A): four nymphal restricted to flowers, two cages were secured together with stages (collectively pre-reproductive stages) followed by a single flower stem between them; no-see-um mesh was two adult stages (reproductive and post-reproductive). used on the outside of both cages to restrict aphids to The reproductive stage included the number of days that reproductive structures (Figure 1C). Aphids were not each female deposited nymphs, whereas the post-repro- exposed to sticky surfaces to reduce incidental mortality. ductive stage included the number of days that adult Each tissue type was artificially infested with two females were alive but no longer depositing nymphs. Total apterous adult cabbage aphids (first generation or Gen1) stage duration (number of days) for each nymphal stage as resulting in 10 cages per treatment (n = 30). On the 1st well as total duration of the pre-reproductive, reproduc- day nymphs were produced, ca. 48 h after infestation, tive, and post-reproductive periods (number of days) were Gen1 adults were removed, and a cohort of five nymphs compared between tissue types using a two-way ANOVA (second generation or Gen2) remained in each cage until (Proc MIXED; SAS 2009). In this study, treatment location nymphs reached the penultimate stage (fourth instar). (flowers, leaf top, and leaf bottom) was a fixed effect, and

A

Figure 2 (A) The life cycle diagram and (B) corresponding stage-classified transition matrix for female cabbage aphids. Stages 1–4 = instars 1–4, B adult = reproductive females, and post = post-reproductive adult females. Arrows indicate transitions, where G = growth (aphid survives from one stage to the next), S = stasis (aphid survives and remains in the same stage), and F5 = fecundity (number of nymphs per reproductive female at stage 5). Feeding location affects aphid demography 153 individual canola plants were a random factor in a mixed et al., 1999). The LTRE method was used to decom- model. The LS MEANS statement was used to make pair- pose the effects of each stage-specific vital rate and wise comparisons between tissue types; means were then their contributions to k. A retrospective analysis was separated for multiple comparisons using an adjusted Tu- used to test how observed changes in vital rates (i.e., key method at a = 0.05. Mean daily nymph production reproduction, growth, and survival) influenced the was compared among treatments using a two-way ANO- finiterateofpopulationgrowth(k) of the cabbage VA (Proc MIXED; SAS 2009). Post-hoc comparisons of aphid in each treatment location (reproductive vs. veg- means were conducted with the adjusted Tukey method at etative). Treatments used for the retrospective analysis a = 0.05. included aphids restricted to the flowering canopy and In addition to differences in life stage duration, we com- a mean matrix for aphids restricted to upper or lower pared transition matrix parameter estimates for aphids at parts of the leaves. Leaf treatments were combined all feeding locations. The cabbage aphid life cycle was gen- because responses and rates were similar among aphids eralized into a 6 9 6 stage-classified transition matrix with restricted to the bottom and top leaves of the same a daily time-step (Figure 2B). Apterous aphid adults pro- canola plant (see Results). The formula for calculating duced only female clones through parthenogenesis, and demographic contributions of lower-level demographic fecundity rates (F) were calculated as the mean number of rates in a fixed effect LTRE analysis is given by: female clones produced by each adult female per day. X ok Growth rates (G) were calculated as the product of daily Dk ¼km kr ð m r Þ ; xij xij o survival (a) and the probability of transitioning to the next ; xij i j Aþ stage (g). Stasis (S) was also a product of daily survival (a) m r þ ðA þ A Þ where A ¼ but was multiplied by the probability of remaining in that 2 stage class (s). We parameterized a 6 9 6 stage-classified matrix for each tissue type (aphids restricted to flowers, The difference in the finite rate of population change leaf tops, and leaf bottoms) using methods outlined by (Dk) between a treatment (m) and a reference matrix (r) is Caswell (1996). approximated as a summation for each lower-level demo- graphic rate (x), where the difference is calculated for each Prospective analysis demographic rate between the manipulated and reference m r We used a prospective analysis to identify demographic matrices (xij xij), and multiplied by the lower-level parameters that would be predicted to have a large effect sensitivity value of the same demographic parameter + on future changes in k (Caswell, 1996, 2001). Matrix ele- (ok=oxij) in a mean matrix (A ). ments were calculated for each treatment in separate All prospective and retrospective analyses were per- matrices using a deterministic matrix population model formed in R (version 2.13.0) using the popbio package approach (Caswell, 2001). For each matrix, we calculated (Stubben & Milligan, 2007). We used parametric boot- the finite population growth rate (k), stable age distribu- strapping to obtain confidence intervals for all matrix tion (w), reproductive value of each stage class (v), sensi- properties in R. First, we calculated the mean and stan- tivities (si,j), and elasticities (ei,j). Elasticities measure the dard error for each of 18 demographic parameters in effect of a proportional change in a matrix element on k our model, including stasis, growth, survival, and fecun- and enable direct comparisons of demographic rates dity. Second, we took a random draw for each of the 18 between survival (bounded between 0 and 1) and fecun- demographic parameters. Fecundity was modeled as ran- dity (bounded between 0 and infinity) among the treat- dom draws from a normal distribution using the rnorm ments. Our matrix elements were products of different function. All other demographic parameters were proba- demographic rates; therefore we calculated lower-level bilities and were modeled as draws from a beta distribu- elasticities for all vital rates that comprised the matrix ele- tion using the betaval function of the popbio package. ments in our population model. For each bootstrap replicate, the set of random draws was combined to parameterize the projection matrix, Retrospective analysis and the asymptotic properties were calculated for that An LTRE model based on fixed effects and a single random matrix. Last, we generated bootstrap distribu- classification design was used to calculate contributions tions for each of the matrix properties by repeating the of the lower-level vital rate differences in k between last two steps for 10 000 random matrices. Variances flowers and leaves (Caswell, 1996, 2001). Changes in and 95% confidence intervals (CI) were then taken the vital rates for individual aphids can be used to directly from bootstrap distributions for each matrix identify vulnerabilities in population growth (Mills property. 154 Cibils-Stewart et al.

(117 14) alatoid nymphs, and 16% (202 34) alates. Results Aphids restricted to vegetative tissues of the same plant Aphid population dynamics had a similar population structure, where 80% Daily rate (mean SEM) of population growth (k)for (162 23), 5% (9 2), and 15% (28 5) of all aphids cabbage aphids was 8% faster for populations restricted to were apterous nymphs and adults (v2 = 27, d.f. = 59, apical flowers compared to populations restricted to vege- P = 1.0), alatoid nymphs (v2 = 13.75, d.f. = 59, tative leaves under greenhouse conditions (1.25 0.01 vs. P = 0.55), and alates (v2 = 6.66, d.f. = 59, P = 1.0), 1.16 0.01; t = 8.66, d.f. = 59, P<0.001). Daily rate of respectively. population growth (k) for cabbage aphids was 1.17 0.08 for populations restricted to the reproductive Individual aphid demographics structures and 1.11 0.02 for populations restricted to In our trials, 25, 36, and 27 of the 50 aphids survived to vegetative tissue under field conditions (t = 2.41, adulthood on the flower, leaf top, and leaf bottom treat- d.f. = 35, P<0.05). Mean total numbers of aphids recorded ments, respectively. Daily nymph counts from surviving after 21 days were 1 104 88 and 1 090 227 for pop- aphids were used for the fecundity analysis. Development ulations restricted to the reproductive structures, and was 20% faster for cabbage aphid restricted to flowers 185 26 and 348 104 for populations restricted to (10.4 0.8 days) compared to aphids developing on tops vegetative structures in greenhouse and field trials, respec- (12.9 0.7 days) or bottoms (12.3 0.8 days) of leaves = < tively. Flower structures supported 5.99 and 3.19 more (F2,75 19.86, P 0.0001). Significant interactions between aphids than lower leaves of the canola plants after a 3-week plant location and duration of nymphal stage were also = < period in greenhouse and field trials, respectively. observed (F6,300 6.63, P 0.0001) (Figure 3A). Aphids Overall population densities were significantly lower spent 60–70% less time in pre-reproductive when aphids were restricted to vegetative tissue compared (8.6 0.1 days) and post-reproductive (6.2 0.5 days) to reproductive tissues (t = 9.97, d.f. = 59, P<0.001), but stages, respectively, compared to reproductive adults the proportion of apterous nymphs and adults (t = 1.93, (21.0 0.5 days), regardless of feeding location = = d.f. = 58, P = 0.059) as well as alates (t = 0.46, d.f. = 58, (F4,225 2.96, P 0.02) (Figure 3B). Stage duration of P = 0.49) within these populations did not differ signifi- aphids across all plant locations ranged from 7.6 0.1 to cantly between feeding locations in the greenhouse trial. 9.2 0.1 days for the pre-reproductive stage, 19.8 0.1 Proportions of the population that were alatoid nymphs, to 21.7 0.1 days for the reproductive stages, and on the other hand, were significantly higher when aphids 3.9 0.4 to 8.1 0.8 days for the post-reproductive were restricted to reproductive vs. vegetative tissues stage. Aphids restricted to flowers had significantly shorter (t = 3.84, d.f. = 58, P = 0.0003). Population structure of post-reproductive durations than aphids confined to = = cabbage aphid restricted to feeding on the reproductive leaves (F4,225 2.96, P 0.02) (Figure 3B). Hence, mean parts of the canola plant after a 3-week period comprised ( SEM) overall life cycle duration was significantly of 72% (786 74) apterous nymphs and adults, 12% shorter for aphids restricted to flowers (31 3 days) than

A B

Figure 3 Mean ( SEM) duration (days) of the (A) nymphal stage for cabbage aphids restricted to specific canola tissues (flower, leaf top, or bottom) and (B) pre-reproductive (Pre-R), reproductive (R), and post-reproductive (Post-R) periods for cabbage aphids restricted to three canola tissue types (flower, leaf top, or bottom) for a greenhouse trial in 2012. Means within a panel capped with different letters are significantly different (adjusted Tukey method: P<0.05). Feeding location affects aphid demography 155 aphids located on upper (39 6 days) or lower leaf sur- Table 2 Growth rates and asymptotic matrix properties for indi- faces (37 7days)(F4,225 = 9.98, P<0.001). vidual cabbage aphids restricted to three specific canola plant Mean daily fecundity values were significantly greater structures (flower, leaf top, and bottom) for a greenhouse experi- for aphids restricted to reproductive canola structures ment in 2012. Results obtained from the prospective matrix k = (3.3 0.1 aphids per female per day) compared to aphids analysis are shown, where finite rate of population change, q = damping ratios, t = days to model convergence, R = net on either top (2.0 0.1) or bottom (2.2 0.1) vegetative 20 0 reproductive rate, and T(d) = generation time in days tissues (F2,75 = 19.86, P<0.001). Vital rates of survival, sta- sis, and mean daily fecundity were pooled across aphids Parameter Feeding location Mean SEM 95% CI limits within the same treatment; and mean values were used for k – the prospective analysis (Table 1). Flower 1.25 0.01 1.23 1.26 Leaf top 1.17 0.01 1.16–1.19 Cabbage aphid restricted to reproductive structures of Leaf bottom 1.17 0.01 1.16–1.18 the plant had higher demographic performance than q Flower 1.69 0.02 1.65–1.73 aphids restricted to vegetative plant tissues; population Leaf top 1.33 0.02 1.30–1.37 k growth rate ( ) was 7% higher for aphids restricted to the Leaf bottom 1.38 0.01 1.36–1.40 k = reproductive parts of canola plant ( 1.25 0.01) t20 Flower 5.7 0.1 5.5–6.0 compared to aphids on vegetative tissues Leaf top 10.4 0.1 9.6–11.4 (k = 1.17 0.01) (Table 2). Damping ratios (q)and Leaf bottom 9.4 0.2 8.9–9.8 – time of convergence (t20), measured in days, showed that R0 Flower 23.3 1.7 20.2 26.8 populations restricted to flowers should converge faster to Leaf top 15.1 3.2 10.1–22.2 – the stable-age distribution (5.7 0.1 days) than aphids Leaf bottom 13.3 1.0 11.5 15.3 T(d) Flower 14.2 0.2 13.9–14.5 restricted to vegetative structures (t20 = 9.4 0.2 and Leaf top 16.8 0.8 15.3–18.5 10.4 0.1 days) (Table 2). Stable age distributions (w) Leaf bottom 16.4 0.2 16.0–16.8 revealed that first instars are the most abundant stable stage class among all populations (w>0.30, Figure 4A), and reproductive values (v, or mean number of offspring in stasis as adults (s5) and also to survival of adults (a5) theoretically produced by post-transients) were consis- (Figure 5A). tently highest for reproductive adults (v>9), followed by Results from the retrospective analysis comparing the the fourth instar stage (v 5) (Figure 4B). Life-history two leaf treatments and a projection matrix of individual patterns were consistent across treatments. Elasticity aphids on reproductive tissue showed that the sum of all values indicated that k would be most sensitive to changes elements in the contribution matrix (∑c) equaled 0.0740; this sum was a good approximation to the expected treat- ment effects (Dk: km kr = 0.0739). Positive contribu- Table 1 Vital rates for individual cabbage aphids (n = 50 per tions of improved adult fecundity (F5: 0.049), growth feeding location) feeding on canola, which includes: growth (G), from nymphal stage 1–2(g1:0.007),2–3(g2:0.038),3–4 stasis (S), and fecundity (F, number of females produced by (g : 0.021), and 4 to adult (g : 0.011), and survival of nym- females per day) rates for nymph (S1–4), adults (S5), and post- 3 4 reproductive (S6) cabbage aphids, restricted to specific tissue phal stage 3 (a3: 0.005) accounted for nearly all of the dif- k types (flower, leaf top, or bottom) for a greenhouse experiment ference in between aphid populations restricted to in 2012 canola flowers vs. a leaf on the same plant. These six demo- graphic rates accounted for ca. 74% of the effects of feed- Vital rate Flower Leaf top Leaf bottom ing location on fecundity, growth, and survival of cabbage aphid (Figure 5B). Additionally, there were negative con- S1 0.32 0.41 0.31 S2 0.30 0.57 0.42 tributions for stasis of early nymphal stages to the variation k S3 0.41 0.43 0.53 in , where only stasis of the second nymphal stage was = – S4 0.49 0.53 0.52 higher for aphids on leaves than flowers (s2 0.025) S5 0.95 0.95 0.95 (Figure 5B). S6 0.74 0.88 0.85 G1 0.60 0.56 0.63 G2 0.57 0.33 0.46 Discussion G3 0.41 0.39 0.29 Feeding location on canola had a large effect on the demo- G4 0.37 0.34 0.33 graphic performance of mixed-age and even-aged popula- G5 0.05 0.05 0.05 tions of aphids. Vital rates and the intrinsic rate of growth F1 3.25 2.04 2.15 (k) were significantly higher for aphids confined to repro- 156 Cibils-Stewart et al.

A of food quality; top-down effects of predators or parasi- toids needs further investigation under field conditions. It is well-known that reproductive and vegetative plant tissues of host plants can provide different resources to herbivorous , and these differences in resource allo- cation can differentially affect herbivore growth rates and development. For example, Smallegange et al. (2007) and Malik et al. (2010) reported that flowers contain signifi- cantly higher levels of compared to leaves in other brassica species. In addition, availability and concen- trations of nitrogen can differ across different plant organs and across plant stage (Augustinussen 1987; Malagoli et al., 2005) with direct effects on aphid growth and devel- opment (Walter & DiFonzo, 2007; Winde & Wittstock, 2011; Winder et al., 2012). Plant-herbivore interactions B may be complex in canola and other brassica crops, as al- lelochemicals like glucosinolates can have both synergistic and antagonistic effects on herbivores and natural ene- mies. Previous studies have documented toxic effects of these secondary compounds on natural enemies, yet cab- bage aphid populations can thrive on a diet with glucosin- olates (Pratt et al., 2008; Ahuja et al., 2009; Hopkins et al., 2009; Chaplin-Kramer et al., 2011; Kos et al., 2011a,b). Consequently, cabbage aphid evolved mechanisms to miti- gate exposure to allelochemicals produced by cruciferous plants by sequestering toxins in body tissues (Slemens & Mitchell-Olds, 1996; Ratzka et al., 2002; Pratt et al., 2008). Allelochemical composition varies among different culti- vated and uncultivated brassica species and sometimes varies across plant organs (Smallegange et al., 2007). Inter- actions between herbivores and secondary compounds in Figure 4 Mean ( SEM) (A) stable stage distribution (w) and different feeding sites warrant further study. (B) reproductive values (v) for cabbage aphids restricted to Three other possible mechanisms may explain the specific canola tissues (flower, leaf top, or bottom) for a increased demographic performance of aphids on canola greenhouse trial in 2012. 1st – 4th, nymphal stages; Post, post- flowers compared to leaves. Although glucosinolates are reproductive adult female. feeding stimulants to the specialist cabbage aphid, high concentrations of these compounds can negatively affect demography of other specialist herbivores like Pieris rapae ductive tissues in the apical region of a canola plant. Indi- L. (Agrawal & Kurashige, 2003). In our study, cabbage vidual aphids feeding on canola flowers exhibited shorter aphids had higher fecundity and a shorter generation time generation times (T), higher net reproductive rates (Ro), (T) when restricted to reproductive parts of the canola and higher fecundities than aphids feeding on vegetative plant. Tradeoffs between nitrogen and con- plant tissues. Similar effects of feeding location have been centrations may occur in different canola plant structures reported for soybean aphids (Aphis glycines Matsumura) (Cibils-Stewart, 2013) and could explain demographic dif- restricted to mature or fully expanded leaves (lower can- ferences we observed. Higher isothiocyanate concentra- opy) vs. newly expanding soybean leaves (upper canopy). tions in reproductive tissues may result in higher fecundity Here, differences in growth rates were mainly attributed to and lower longevity of aphids on leaves (Agrawal et al., a combination of top-down and bottom-up factors gov- 1999; Agrawal & Kurashige, 2003; Smallegange et al., erning aphid population growth (McCornack et al., 2008; 2007; Malik et al., 2010; Winde & Wittstock, 2011), but Costamagna et al., 2013). By using exclusion cages in our these differences need to be further investigated for this field experiment, we demonstrated that effects of location system. Last, inherent differences in life histories among on aphid demography are mainly due to bottom-up effects aphid species colonizing canola may select for demo- Feeding location affects aphid demography 157

A

Figure 5 (A) Mean ( SEM) elasticity values for lower-level vital rates for three populations of cabbage aphids restricted to specific canola tissues (flower, leaf top, or bottom) for a greenhouse trial in 2012, and (B) contributions of lower-level vital rates B to variation in the finite rate of population change (k)betweencabbageaphid populations restricted to flowers vs. vegetative tissues (mean of leaf top and bottom locations) of canola plants; positive contributions indicate better demographic performance in flowers. s, stasis; g, growth; a, survival; f, fecundity; 1- 4, nymphal stages; 5, adult; 6, post- reproductive aphid. graphic parameters that contribute differently to popula- vs. leaves; reproductive adults had the single greatest posi- tion growth. For instance, a fitness consequence for aphids tive contribution to k.Weobservedthatstasisduringearly feeding in glucosinolate-rich tissues of the plant is reduced nymph development resulted in negative contributions to longevity, but mechanisms that effectively sequester gluco- k, which can be explained by higher mortality rates among sinolates may counteract reductions in longevity by short- early nymphal stages in cabbage aphid. High mortality ening generation time and increasing female fecundity (50–72%) during early nymphal stages observed in our (Agrawal & Kurashige, 2003), resulting in more progeny study may be due to host-specific attributes, which has produced earlier in the life-history and over a shorter been documented in other studies. Specifically, Ulusoy & duration. Shelton (2005) reported that spatial variation in Olmez-Bayhan (2006) reported host-specific mortalities defenses within or among plant tissues could slow the evo- ranging from 16 to 88% when aphids were confined to dif- lution of resistance to herbivores by creating uneven selec- ferent brassica hosts (, cabbage, mustard, cauli- tion pressure on herbivores and their natural enemies. flower, turnip, and rapeseed); the lowest mortality (16%) Quantification of glucosinolates within different plant tis- was observed when cabbage aphids were confined to cab- sues and corresponding effects on canola aphids requires bage. further investigation. Feeding location influenced demography of aphids Our prospective analysis showed a nymph-skewed sta- directly, where different plant structures are acting as ble stage distribution, which is common in growing popu- either sources or sinks to cabbage aphid. Source-sink rela- lations (Taylor, 1979). Unsurprisingly, reproductive values tionships within the plant directly affected aphid demogra- were higher for adults and the penultimate nymphal stage phy at both the individual and population level. Our result as only adult aphids are reproductive. Elasticity values is a novel finding that helps shape our understanding of showed that k is most sensitive to future changes in stasis how canola plant structures differentially affect aphid and survival of adults. Even with a nymph-skewed stable demographic parameters. To our knowledge, this is the stage distribution, nymphal stages have the highest mortal- first study to use a combination of prospective and retro- ity rates within populations; therefore, adults and penulti- spective demographic analyses to identify vital rates that mate instars are more important in shaping cabbage aphid contribute most to differences between aphids reared on population dynamics. Understanding the demographic different plant structures. Our results indicate that special- composition of aphid populations under field conditions ist aphids not only have greater growth rates, but also can guide classical biological control programs. For exam- shorter generation times and higher fecundities when ple, identifying predators or that prefer nym- restricted to reproductive tissues of the canola plant, which phal aphid stages would have the greatest influence on k may be a direct outcome of a specialist aphid adapting to (Latham & Mills, 2012). Retrospective analysis indicated canola secondary compounds. We hypothesize that cab- that contributions from nymphal growth stages 2–3, 3–4, bage aphids developing on flower structures will also con- and 4 to adult (Figure 5B) collectively accounted for most tribute more to aphid migration events and colonization of the variation in k between feeding locations on flowers of new canola plants, as the proportion of alatoid nymphs 158 Cibils-Stewart et al. was significantly higher when aphids were restricted to Ahuja I, Rohloff J & Bones AM (2009) Defense mechanisms of reproductive vs. vegetative tissues. Our findings are unsur- Brassicaceae: implications for -plant interactions and prising because changes in dispersal polymorphisms are potential for integrated pest management. Agronomy for Sus- – directly related to aphid population size and to crowding tainable Development 30: 311 348. (Boyles et al., 2012). Ash M (2012) Canola. USDA Economic Research Service. 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