Feeding Behavior of Russian Wheat Aphid (Hemiptera: Aphididae) Biotype 2 in Response to Wheat Genotypes Exhibiting Antibiosis and Tolerance Resistance

PLANT RESISTANCE Feeding Behavior of Russian Wheat Aphid (Hemiptera: Aphididae) Biotype 2 in Response to Wheat Genotypes Exhibiting Antibiosis and Tolerance Resistance

SONIA LAZZARI,1 SHARON STARKEY,2 JOHN REESE,2 ANDREA RAY-CHANDLER,3 4 2,5 RAYMOND MCCUBREY, AND C. MICHAEL SMITH Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021

J. Econ. Entomol. 102(3): 1291Ð1300 (2009) ABSTRACT In this study, wheat, Triticum aestivum L. (em Thell), genotypes containing the Dnx, Dn7, Dn6, and Dn4 genes for resistance to the Russian wheat aphid, Diuraphis noxia (Kurdjumov) (Hemiptera: Aphididae), along with Dn0, a susceptible control, were assessed to determine the categories of D. noxia biotype 2 (RWA2) resistance in each genotype and RWA2 feeding behaviors on Dnx and Dn0 plants by using the electronic penetration graph technique. At 14 d postinfestation, Dn0 plants exhibited intense chlorosis and leaf rolling, and all test genotypes expressed some degree of chlorosis and leaf rolling, except Dn7, which was not damaged. Both Dn7 and Dnx expressed antibiosis effects, signiÞcantly reducing the numbers of aphids on plants and the intrinsic rate of aphid increase. Dn6 plants seemed to contain tolerance, exhibiting tolerance index measurements for leaf and root dry weight and plant height that were signiÞcantly lower than those of the susceptible Dn0 plants. Principal component analyses indicated that antibiosis and leaf rolling data explained 80% of the variance among genotypes. Electronic penetration graph analysis demonstrated contrasting results between RWA1 and RWA2 phloem sieve element phase feeding events, but results indicated that Dnx resistance factors are present in the sieve element cells or phloem sap. Plants containing Dnx exhibit antibiosis resistance to D. noxia RWA2 similar to that in plants containing the Secale cereale L. (rye)-based Dn7 gene without the negative baking quality traits associated with Dn7.

KEY WORDS antibiosis, electronic penetration graph, plant resistance, tolerance index

Diversity for virulence to resistant cereals within the genetic variation for virulence and understand the Russian wheat aphid, Diuraphis noxia (Kurdjumov) genetic and ecological bases for biotype development (Hemiptera: Aphididae), was not known in North and Þtness. America until a virulent isolate was detected in Col- Nevertheless, research has continued since Haley et orado in 2003 (Haley et al. 2004). This isolate, desig- al. (2004) found the single dominant gene Dn7 de- nated as D. noxia biotype 2 (RWA2), was reported to ployed in the wheat genotype 94M370 to be the only be virulent to eight of the nine D. noxia resistance RWA2-resistant gene. 94M370 was developed in South genes (Burd et al. 2006). The discovery of RWA2 is Africa (Marais et al. 1998) and carries Dn7 on a negatively impacting production of wheat, Triticum 1BL.1RS translocation arm transferred from ÔTurkey aestivum L. (em Thell), for which RWA1-resistant 77Ј rye (Secale cereale L.). An additional Dn7-based cultivars have provided good D. noxia management. source of RWA2 resistance, 94M81, was identiÞed by Virulent D. noxia populations also exist in Hungary Porter et al. (2005). Although Dn7 confers a high level (Basky 2003), Iran (Dolatti et al. 2004), Chile (Smith of RWA2 resistance, rye genes related to secalin pro- et al. 2004), and South Africa (Tolmay et al. 2007). duction reduce yield and baking protein quality in Given these developments, Burd et al. (2006) has bread wheat genotypes containing Dn7 (Graybosch et indicated that for D. noxia resistance programs to al. 1990). All other RWA1-resistant Dn genes used in succeed, it will be necessary to assess the amount of North American wheat breeding originate in hexaploid wheat (Liu et al. 2001). Useful RWA2 resistance, if identiÞed there, is preferable, to avoid 1 Department of Zoology, Universidade Federal do Parana, Cu- rye-based baking quality problems. ritiba, PR 81531-980 Brazil. 2 Department of Entomology, Kansas State University, Manhattan, Collins et al. (2005) identiÞed resistance to RWA2 KS 66506-4004. in the wheat cereal introduction (CItr) 2401 as well as 3 Department of Horticulture, Johnson County Community Col- several other wheat genotypes, and Voothuluru et al. lege, Overland Park, KS 66210-1299. (2006) demonstrated that CItr 2401 resistance is the 4 Department of Biostatistics, University of Alabama at Birming- ham, Birmingham, AL 35294. result of a strong antibiosis effect that signiÞcantly 5 Corresponding author, e-mail: [email protected]. reduces the intrinsic rate of increase of RWA2. CItr

0022-0493/09/1291Ð1300$04.00/0 ᭧ 2009 Entomological Society of America 1292 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 102, no. 3

2401 also exhibited tolerance to RWA2 chlorosis and ability to reach the phloem and the quantitative and leaf rolling but not antixenosis. Resistance to RWA2 in qualitative properties of the sieve elements. After in- CItr 2401 is conferred by two dominant genes serting their stylets into plant tissues, aphids probe the (Voothuluru et al. 2006) and is equivalent to that in leaf substrate, brießy penetrating cells before reaching 94M370. Ennahli et al. (2009) conÞrmed antibiosis to the phloem. All phases of aphid feeding, including the RWA2 in CItr 2401 and identiÞed antibiosis to RWA2 stylet penetration of tissues, may be detected by in barley genotype IBRWAGP4-7. Neither genotype the electronic penetration graph (EPG) technique expressed tolerance of leaf dry weight loss to RWA2 (McLean and Kinsey 1964; Tjallingii 1985; Backus feeding. Weiland et al. (2008) determined that Dnx, a 1994). Over time, changes in the resistance and elec- hexaploid wheat gene identiÞed as resistant to RWA1 tromotive force of the circuit consisting of plant tis- (Harvey and Martin 1990), expressed resistance to sues and the aphid result in recognizable waveforms RWA1, -6, -7, and -8 in Colorado and intermediate that can be correlated to insect stylet penetration

resistance to RWA2. behavior. IdentiÞcation and quantiÞcation of insect Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021 As indicated by Voothuluru et al. (2006), both the waveforms provide important information about the chromosome location of a resistance gene, as well as physical location of plant resistance factors and the knowledge of the category of resistance present in a time course of insect responses to these factors. Sev- germplasm are important for developing sustainable eral studies have correlated EPG waveforms to spe- resistant cultivars. Resistance may be categorized as ciÞc aphid probing behaviors during feeding. EPG antibiosis, antixenosis, or tolerance (Smith 2005). An- monitoring has been used to identify plant phloem as tixenosis occurs when the plant is a poor host, resulting the site of resistance in several aphid species and other in reduced colonization of a plant by arthropods and, Hemiptera. Girma et al. (1992) used EPG monitoring therefore, losses caused by the pest (Pedigo 1999). to determine that RWA1 salivated more, ingested less, Antibiosis is present when a plant adversely affects and took 4 times longer to reach the phloem when growth and development of a pest arthropod. This may feeding on nonhost sorghum plants than on wheat and be caused by the presence of certain chemicals or the other host plants. On the basis of their results, they absence of or insufÞcient nutrients in a plant, which proposed that feeding behavior detected by the EPG results in reduced infestation of the arthropod and a could be used to understand the resistance mecha- considerable reduction in plant damage (Painter nisms exhibited by resistant genotypes and identify 1951). Tolerance resistance, unlike antibiosis and an- new sources of resistance to D. noxia. tixenosis that involve plantÐarthropod interactions, is This study was conducted to evaluate the wheat the ability of a plant to withstand arthropod damage genotypes containing Dnx, Dn4, Dn6, and Dn7 for and yield signiÞcantly higher dry mass than the sus- antibiosis and tolerance to RWA2 and to evaluate the ceptible plant under similar conditions. Tolerance is feeding behavior of RWA1 and RWA2 on leaves of an attractive trait for incorporation into cultivars for plants containing Dnx, compared with susceptible and durable arthropod resistance because it raises the level resistant genotypes, by using EPG analysis. The Dnx at which the economic injury level occurs and delays gene is of particular interest to our efforts, because it the need for chemical control. Many arthropod-resis- is derived from hexaploid wheat, as opposed to the tant genotypes contain one, two, or all three categories rye-based Dn7 gene. In addition, several genotypes of resistance and each category alone or in combina- containing Dnx have been produced for development tion has been identiÞed in several cereal accessions. as wheat cultivars. Several techniques are in use for assessing tolerance and antibiosis to aphids. However, some fail to sepa- Materials and Methods rate the two components, whereas others do not con- sider the variation in plant biomass among cultivars. Plant and Insect Materials. The plant genotype Reese et al. (1994b) proposed a method for assessing KS041140, which contains the Dnx gene from hexaploid tolerance to the greenbug, Schizaphis graminum (Ron- wheat plant introduction (PI) 220127 originally de- dani), in sorghum, Sorghum bicolor (L.) Moench, scribed by Liu et al. (2001), was evaluated for RWA2 which is independent of antibiosis effects. The resistance. KS041140 was developed by bulking ten method combines measures of proportional tissue highly RWA1-resistant F2:3 plant families originating weight change (DWT) and aphid population to cal- from the cross (PI 220127/ÔSandoÕs SelectionÕ) and culate a tolerance index (TI) originally developed by advancing this population in bulk to the F5 generation Dixon et al. (1990). Genotypes with TI values signif- (C.M.S. et al., unpublished manuscript). The breeding icantly lower than those of the susceptible controls are lines 94M370, containing Dn7; CO960223, containing considered tolerant. Voothuluru et al. (2006) used the Dn6; the variety Yumar, containing Dn4; and the sus- intrinsic rate of increase (rm) formula developed by ceptible control ÔJaggerÕ (Dn0) also were evaluated. Wyatt and White (1977) to assess antibiosis to D. noxia Seed stocks are currently maintained by the Plant biotype 2 in Cltr 2401. Resistance Laboratory, Department of Entomology, Selection of a feeding site by aphids and the time Kansas State University (KSU), Manhattan, KS. required to reach the phloem are determined by an- RWA1 used in experiments originated from a cul- atomical and chemical characteristics of the plant tis- ture established with aphids collected from Hays, KS, sue. According to Klingauf (1987), the acceptance of in 2004, courtesy of Dr. J. P. Michaud (Department of a host plant by an aphid depends upon the aphidÕs Entomology, KSU). RWA2 used in the experiments June 2009 LAZZARI ET AL.: FEEDING BEHAVIOR OF RUSSIAN WHEAT APHID 1293 were from an aphid colony originally collected near (length to the tip of the longest leaf before drying) was Briggsdale, CO, in 2004. Each biotype was cultured also determined. Tolerance for each plant genotype separately on susceptible Jagger plants in the green- was calculated using the percent proportional plant house. The genetic identity of each colony has been root or shoot dry weight (DWT) or the percent pro- veriÞed yearly in diagnostic plant differential assays at portional plant height, expressed as DWT ϭ [(WC Ϫ Manhattan, KS, since 2005. WT)/WC] ϫ 100; where WC ϭ DWT (or height) of Plant Phenotypic Analyses. Experiments comparing noninfested control plants and WT ϭ DWT of infested plants containing Jagger (Dn0), Yumar (Dn4), plants. The tolerance index (TI) of each experimental CO960223 (Dn6), 94M370 Dn7, and KS041149 (Dnx) and control genotype was determined to compensate were carried out to determine whether each gene for the confounding effect of differing numbers of expressed antibiosis or tolerance to RWA2. Seeds of aphids on infested plants. TI was calculated as TI ϭ each genotype were planted in 20 individual, 10-cm- DWT/number of aphids produced on infested plants

diameter plastic pots Þlled with Pro-MixBx (Hummert (Dixon et al. 1990). Genotypes with TI values signif- Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021 International, Topeka, KS) potting mix and grown in icantly lower than those of the susceptible Jagger Dn0 the greenhouse (24ЊC day:20ЊC night, at a photoperiod control were considered to be tolerant to RWA2 feed- of 14:10 [L:D] h). Ten pairs of plants, each containing ing. an aphid-infested and a noninfested plant, were then Electronic Penetration Graph Analyses. For EPG established within each genotype, on the basis of analysis, plants containing Dnx were from F5 seed of height and growth, and pots were arranged in a com- the original 10 F2:3 bulked resistant families from the pletely randomized design. Plants were grown to the cross (PI 220127 (Dnx)/SandoÕs Selection) (Liu et al. Ϸ two-leaf stage ( 6 d after germination), and in each 2001). Similarly, Dn0 plants were from F5 seed of the treatment pair, one plant was infested with Þve apter- original ten bulked susceptible families of the same ous last-stage nymphs or new adults. A noninfested cross. Plants were grown to the two- or three-leaf stage plant in each pair served as a control. Infested and in 3.8-cm-diameter by 21-cm-high poly-cast container noninfested plants were covered with a nylon mesh tubes containing Pro-MixBx (Hummert International) cage, and aphids were allowed to feed and reproduce in the greenhouse at 30ЊC day:22ЊC night and a pho- until the infested Jagger plants showed complete leaf toperiod of 14:10 (L:D) h. Seeds of the RWA2 resistant rolling and 95% chlorosis of the youngest true leaf at wheat cereal germplasm CItr 2401 (Voothuluru et al. Ϸ14 d after infestation. Plants of each genotype were 2006) were planted and grown similarly. Plants were rated on a 0Ð3 scale for D. noxia leaf streaking (chlo- taken to the laboratory, where the upper (adaxial) rosis): 0, no damage; 1, Ͻ50% streaking; 2, Ͼ50% surface of the distal third of the newest leaf of each test streaking; and 3, 100% streaking; and for leaf rolling plant was secured with cellophane tape to a cardboard damage: 0, no damage; 1, Ͻ50% rolling; 2, Ͼ50% roll- stage supported by a bamboo stake to maintain a ßat ing; and 3, 100% rolling (Liu et al. 2001). and immobile surface for the duration of the EPG Antibiosis. Antibiosis was determined by collecting recording. Newly molted apterae RWA1 and RWA2 aphids on each of the infested plants on a sheet of wax adults were individually held in place with a light paper, placing them in 70% alcohol, and counting vacuum and connected at the dorsum to a 3-cm-long them. To further assess antibiosis, the intrinsic rate of by 18-␮m section of gold wire with conductive silver natural increase (rm) of RWA2 on each genotype was paint (Diaz et al. 2006). The other end of the wire was determined. RWA2 rm was calculated by placing one previously connected to an ampliÞer input. After wir- late-instar RWA2 nymph (P1) on a plant leaf and ing and before testing, aphids were placed on an input conÞning it in a ventilated translucent drinking straw holder and withheld from food for2htoensure hun- cage. After producing its Þrst progeny (F1), P1 was ger and minimize aberrant feeding behavior. Individ- removed and placed on a second leaf of the same plant ual aphids were then placed on the upper surface of and in a separate cage. When an F1 produced its Þrst each plant leaf. offspring, the experiment was stopped, and the time EPG recordings were conducted in the laboratory Ϯ Њ required for a F1 to produce its Þrst offspring “d” was at 25 2 C with overhead ßuorescent lighting sup- determined. The total number of offspring produced plementing natural daylight. Infested wheat plants byaP1 during d was determined as Md. The intrinsic were placed inside a Faraday cage during the record- rate of RWA2 increase on each test plant genotype was ing process. A Giga eight DC ampliÞer with eight ϭ 9 then calculated as rm logeMd/d (Wyatt and White channels and 10 -ohm input resistance (Wageningen 1977). The mean RWA2 rm on each plant genotype Agricultural University, Wageningen, The Nether- was determined using 10 replicates per genotype. lands) was used with output signal converted from Tolerance. Shoots from noninfested and infested analog to digital by a DI-720 Data Acquisition System plants were cut at the soil level and placed in pre- and recorded with WinDaq Lite Waveform Recording weighed, aluminum foil pouches. Roots were washed Software (both supplied by Dataq Instruments, Ak- thoroughly to remove attached soil particles and ron, OH) onto a computer hard disk. Four resistant placed in similar preweighed, aluminum foil pouches. and four susceptible plants were randomly assigned to Pouches with the shoots and roots were dried at 70ЊC the Þrst four or last four of eight available recording for 60 h, and root and shoot weights were determined channels. Actual recording time began just before by subtracting the weight of the foil pouch from the lowering the aphids onto the leaf surface, and com- combined pouch and tissue weight. Shoot height plete experimental data collection ran for 10 h. After 1294 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 102, no. 3

(Table 1. Mean ؎ SE aphid number, leaf chlorosis, and leaf rolling for ﬁve wheat genotypes after 14 d of D. noxia biotype 2 (RWA2 infestation under 24°C day:20°C night, and a photoperiod of 14:10 (L:D) h

Wheat D. noxia resistance Mean Ϯ SE no. aphids Mean Ϯ SE leaf chlorosis Mean Ϯ SE leaf rolling genotype gene Jagger Dn0 (none) 155.1 Ϯ 15.51a 3.0 Ϯ 0.0aa 2.9 Ϯ 0.10aa Yumar Dn4 127.8 Ϯ 12.67a 2.3 Ϯ 0.21b 2.5 Ϯ 0.22ab CO960223 Dn6 135.2 Ϯ 18.27a 1.2 Ϯ 0.20d 2.2 Ϯ 0.25b KS041140 Dnx 83.0 Ϯ 14.22b 1.8 Ϯ 0.13c 1.5 Ϯ 0.17c 94M370 Dn7 3.2 Ϯ 1.35c 0.3 Ϯ 0.15e 0.1 Ϯ 0.10d

Means within a column followed by the same letter are not signiÞcantly different (␣ ϭ 0.05, PROC GLM; LSD). a Damage rating on 0Ð3 scale for D. noxia leaf streaking (chlorosis) damage: 0, no damage; 1, Ͻ50% streaking; 2, Ͼ50% streaking; and 3, 100% streaking; and for leaf rolling damage: 0, no damage; 1, Ͻ50% rolling; 2, Ͼ50% rolling; and 3, 100% rolling. Experiments were conducted in October

and November 2006. Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021 recording, waveforms for each aphid were identiÞed genotypes. Pearson correlation coefÞcients were cal- by type and time-marked, and data were exported into culated for chlorosis, leaf rolling, aphid number, and an electronic spreadsheet for statistical analysis. Data DWT, and these data were also subjected to a log10 collection used for each aphid began 8 h from the time transformation and principal component analysis of the Þrst probe into the plant tissue. The longer (PCA; PAST version 1.73; Hammer et al. 2001) by experimental data collection time allowed for the vari- using the normalized variance-covariance correlation ability in time to Þrst probe for each aphid. Experi- mode. The Jolliffe cutoff value (Jolliffe 1986) was used mental runs in which individual aphids became un- to indicate the signiÞcance of the components. Com- glued from their wire tethers were replaced with runs ponents with eigenvalues smaller than the Jolliffe cut- under the same conditions. The numbers of replica- off were considered insigniÞcant. TI values were not tions were as follows for biotype 1: Dn0, n ϭ 15; Dnx, included because variables that compose them (DWT n ϭ 13; and CItr2401, n ϭ 8; and biotype 2: Dn0, n ϭ and aphid number) were transformed independently. 5; Dnx, n ϭ 13; and CItr2401, n ϭ 11. RWA2 fecundity and respective standard errors were The end of each experiment was determined by calculated by the jackknife method (Tabvida version adding exactly8htothetime of the Þrst probe, thus 1.0), and the means were compared by the unilateral eliminating the highly variable time period required to t-test. initiate probing. Time to Þrst probe was calculated EPG data were found to be non-normally distrib- separately from the actual 8-h experiment. Other pa- uted according to AndersonÐDarling tests of normality rameters included beginning and end time of the Þrst (Stephens 1974) and were then subjected to analysis sieve element phase (SEP) (Prado and Tjallingii of variance (ANOVA) (PROC ANOVA). KruskalÐ 1994), beginning and end times of subsequent SEPs, Wallis tests were conducted with paired mean com- duration of all SEPs, and duration of the available SEP, parisons to determine signiÞcance at ␣ ϭ 0.05. Data for calculated as the difference between the beginning of total numbers of SEP bouts events and for the mean the Þrst SEP and the end of individual aphid data proportion (sum of all SEP bouts/available SEP ac- collection. Thus, the available SEP represented the tivity) were analyzed by using a generalized linear amount of time available to an aphid for feeding, model (PROC GLM). Data for the Þrst probe to the depending upon when it began probing a leaf. These Þrst SEP event were subjected to log10 transformation. data were used to calculate the following parameters of comparison between aphid feeding on different Results and Discussion plant genotypes: mean time until initiation of Þrst probe, mean Þrst probe plus 8 h, mean difference in Plant Phenotypic Analyses. Differences between time from Þrst probe to Þrst SEP, mean available SEP, genotypes were signiÞcant for chlorosis (F ϭ 42.12; mean sum of all bouts of SEP, mean duration of SEP df ϭ 4, 45; P ϭ 0.0001) and leaf rolling (F ϭ 37.75; df ϭ events, mean duration of total pathway phase (the sum 4, 45; P ϭ 0.0001). Shoots of plants containing Jagger duration of all probing activity excluding SEP activ- (Dn0) and Yumar (Dn4) were greatly deteriorated ity), mean total number of SEP bouts, and the mean when they were harvested at 14 d postinfestation with proportion (sum of all SEP bouts/available SEP ac- RWA2. Leaves presented heavy chlorosis resulting tivity). SEP events included the E1 salivation phase, from aphid feeding, and the newly formed leaves were the E2 phloem sap ingestion phase, and continuous trapped inside the convoluted, rolled leaves where salivation. aphids took refuge. Despite sustaining high aphid pop- Statistical Analyses. Data for phenotypic ratings, ulations, plants containing CO960223 (Dn6) showed antibiosis, and tolerance were analyzed by using limited chlorosis or leaf rolling relative to the suscep- ANOVA PROC GLM equal and unequal variance tible Jagger Dn0 control (Table 1). Plants containing models (SAS Institute 1999). The least signiÞcance KS041149 (Dnx) did not begin to exhibit chlorosis difference (least signiÞcant difference [LSD]) pro- until 14 d postinfestation (Table 1). 94M370 Dn7 cedure (␣ ϭ 0.05) was used, when appropriate, to plants exhibited few chlorosis and leaf rolling symp- determine the signiÞcance of mean differences among toms, apparently because very few aphids survived on June 2009 LAZZARI ET AL.: FEEDING BEHAVIOR OF RUSSIAN WHEAT APHID 1295 these plants (Table 1). Leaf trapping was observed in on KS041140 Dnx plants (Table 1). The initial popu- plants of Jagger Dn0, Yumar Dn4, and CO960223 Dn6, lation of Þve aphids on Jagger Dn0 plants increased but 94M370 Dn7 and KS041149 Dnx plants had signif- Ϸ30-fold (mean, 155 aphids) in 14 d postinfestation icantly fewer plants with trapped leaves (data not compared with a mean of 83 aphids (Ϸ50% less) on shown). KS041140 Dnx plants. Infestations on 94M370 Dn7 There were small differences between leaf chlorosis plants resulted in a mean population decrease (mean, and leaf rolling ratings in our experiments and those of 3.2 aphids). Weiland et al. (2008) for plants containing Dn7 and On all genotypes, 50Ð76% of the morphs were Þrst Dnx. When the Weiland et al. (2008) 0Ð12 scale was to third instars when plants were harvested at 14 d adjusted to a 0Ð6 scale, both studies detected similar postinfestation, and few had reached the adult stage combined mean chlorosis and leaf rolling for 94M370 (data not shown). Similar antibiosis effects were ob- Dn7 plants (0.4 of 6.0 in the current study; 1.4 of 6.0 in served by Voothuluru et al. (2006), who noted Ϸ150

the Weiland et al. study). For plants containing Dnx, aphids per plant on susceptible wheat varieties com- Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021 combined leaf damage was 3.3 of 6.0 in the current pared with Ϸ25 aphids per plant on CItr 2401 at 15 d study with Dnx in the KS041149 genotype compared postinfestation. Mean numbers of aphids produced on with 4.3 of 6.0 in the Weiland et al. (2008) study Dn0 and Dn4 plants in our experiments compared with Dnx in the KS94WGRC29 genotype. The favorably to results of Jyoti et al. (2006), who infested KS94WGRC29 genotype pedigree (PI220127/P5// plants with two fourth-instar nymphs and counted a ÕTAM-200Ј/KS87H66) is more complex than that of mean of 124.9 aphids at 20 d postinfestation on Dn0 the KS0411049 (PI220127/ÔSandoÕs SelectionÕ). Nev- plants and a mean of 116.3 aphids on Dn4 plants. ertheless, combined leaf damage scores from both Smith et al. (2005) demonstrated antibiosis to studies indicated that Dnx possesses moderate RWA2 RWA1 in Dn4 and Dn6 plants, as evidenced by aphid resistance. Combined mean leaf damage for suscep- populations that were signiÞcantly reduced compared tible controls in each study were also very similar (5.9 with those on plants of the susceptible cultivar Wich- of 6.0 for Jagger Dn0 plants in the current study and ita. In contrast, our results showed no signiÞcant dif- 5.9 of 6.0 for Yumar in the Weiland et al. study). ferences in the numbers of RWA2 on plants with Dn4, Our results for the performance of plants containing Dn6, or the susceptible Jagger Dn0 control, indicating Dn4 are supported by those of Collins et al. (2005), that RWA2 has overcome the RWA1 defenses in Dn4 who found similar symptoms on plants carrying Dn4 as and Dn6 plants. early as 7 d postinfestation with RWA2 and with The fertility (relative number of nymphs produced 94M370 (Dn7) plants that showed very few foliar per female in 14 d) (F ϭ 18.66; df ϭ 4, 45; P ϭ 0.0001) damage symptoms. Our results for leaf trapping are and daily nymph production per female (F ϭ 18.17; similar to those of Burd et al. (2006). Leaf rolling in the df ϭ 4, 45; P ϭ 0.0001) of RWA2 were signiÞcantly leaves of susceptible plants is an important damage reduced in aphids feeding on Dn7 and KS041149 Dnx criterion because it is related to the biological Þtness plants compared with aphids feeding on Jagger Dn0, of D. noxia. Aphids seek shelter in rolled leaves, avoid- CO960223 Dn6, and Yumar Dn4 plants (Table 2). ing predation, parasitism, and toxicity from insecti- However, the intrinsic rate of increase (rm) of aphids cides. on 94M370 Dn7 plants (Ϫ0.073) was signiÞcantly less

As reported by Haley et al. (2004), there is a clear than rm of aphids feeding on plants of Jagger Dn0 difference in virulence between RWA1 and RWA2. (0.179), Yumar Dn4 (0.168), CO960223 Dn6 (0.171), Damage by RWA2 is more intense than that reported and KS041149 Dnx (0.146) (Table 2). Most females on for RWA1 on RWA1-resistant cultivars carrying Dn4, Dn7 plants died without reproducing or yielded very compared with 94M370 (Dn7). Porter et al. (2005) few nymphs. These results conÞrm that Dn7 nega- also found a high level of resistance to RWA2 in 94M81 tively affects the rm of RWA2, whereas higher aphid (Dn7), similar to 94M370 in our study and to results of populations on plants of all other genotypes yielded

Haley et al. (2004). In addition, Liu et al. (2001) noted positive rm values. Voothuluru et al. (2006) found that a difference between plants containing Dn4 and Dn6 the rm of RWA2 conÞned on resistant CItr 2401 plants in reaction to RWA1. Leaves of Dn4 plants showed (0.08) was signiÞcantly lower than the rm of aphids on very little chlorosis but exhibited moderate leaf roll- susceptible Karl plants (0.11), and Miller et al. (2003) ing, whereas leaves of Dn6 plants exhibited chlorosis determined that rm values for RWA1 ranged from 0.24 but no leaf rolling. In our trials, plants containing both on a resistant wheat genotype to 0.29 on a susceptible Yumar Dn4 and CO960223 Dn6 infested with RWA2 genotype. Our results for the growth rate of RWA2 on exhibited more intense chlorosis and leaf rolling than genotypes containing Yumar Dn4 and CO960223 Dn6 damage caused by RWA1 (Smith et al. 2005). Jyoti et are also similar to those of Jyoti et al. (2006), who al. (2006) also reported greater RWA2-related damage demonstrated that the fecundity of RWA2 is higher to a group of wheat cultivars than damage from RWA1 than that of RWA1 on the Dn4 genotype. to the same cultivars. Tolerance. Leaf DWT from plants infested with Antibiosis. At 14 d postinfestation, there were sig- RWA2 was signiÞcantly different among wheat geno- niÞcantly fewer RWA2 aphids on 94M370 Dn7 and types on the basis of proportional leaf DWT (F ϭ KS041140 Dnx plants than on plants containing Dn4, 13.90; df ϭ 4, 45; P ϭ 0.0001). Plants of CO960223 Dn6 Dn6, or Dn0 (F ϭ 19.58; df ϭ 4, 45; P ϭ 0.0001), and and 94M370 Dn7 had signiÞcantly lower leaf DWT signiÞcantly fewer aphids on 94M370 Dn7 plants than values than Jagger Dn0, Yumar Dn4, and KS041149 1296 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 102, no. 3

Table 2. Fecundity (mean ؎ SE number of nymphs per female), mean ؎ SE daily nymph production per female, and intrinsic rate of natural increase (rm) for D. noxia biotype 2 (RWA2) after feeding for 14 d on ﬁve wheat genotypes under 24°C day:20°C night and a photoperiod of 14:10 (L:D) h

Wheat D. noxia resistance Mean Ϯ SE no. nymphs Mean Ϯ SE daily nymph Mean Ϯ SE r genotype gene per female production m Jagger Dn0 (none) 30.1 Ϯ 3.1a 2.2 Ϯ 0.2a 0.179 Ϯ 0.001a Yumar Dn4 24.6 Ϯ 2.5a 1.8 Ϯ 0.3a 0.168 Ϯ 0.001a CO960223 Dn6 26.1 Ϯ 3.7a 1.8 Ϯ 0.2a 0.171 Ϯ 0.001a KS041140 Dnx 15.7 Ϯ 2.8b 1.1 Ϯ 0.2b 0.146 Ϯ 0.001a 94M370 Dn7 0.46 Ϯ 0.2c 0.03 Ϯ 0.02c Ϫ0.073 Ϯ 0.004b

Means within a column followed by the same letter are not signiÞcantly different (␣ ϭ 0.05, PROC GLM; t-test); n ϭ 10. rm,logeMd/d, where d is time (days) required for a newly emerged aphid to produce its Þrst offspring; and Md is total number of progeny ͓ ͔ produced by the mother of F1 P1 . Experiments were conducted in October and November 2006. Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021

Dnx (Table 2). The DWT of Dnx plants was also and proportional plant height changes, and there were signiÞcantly (P ϭ 0.05) less than that of Dn0 (Table 3). signiÞcant differences among genotypes for shoot The same trends held true for root DWT values (F ϭ DWT (F ϭ 8.52; df ϭ 4, 45; P ϭ 0.0001), root DWT (F ϭ 7.53; df ϭ 4, 45; P ϭ 0.0001). The CO960223 Dn6 root 4.96; df ϭ 4, 45; P ϭ 0.0021), and plant height (F ϭ 6.91; DWT value was signiÞcantly less than that of Jagger df ϭ 4, 45; P ϭ 0.0002) (Table 3). However, only Dn0, Yumar Dn4, or KS041149 Dnx plants, and the CO960223 Dn6 showed TI values smaller than those of 94M370 Dn7 root DWT value was signiÞcantly less the susceptible control Jagger Dn0. Thus, CO960223 than that of plants of Jagger Dn0 or KS041149 Dnx Dn6 was the only genotype truly tolerant to RWA2. (Table 3). The variance for 94M370 Dn7 was very high for aphid Changes in proportional plant height were also sig- number, leaf and root DWT, and plant height. A sep- niÞcantly different among the genotypes (F ϭ 8.45; arate analysis, conducted without 94M370 Dn7 data, df ϭ 4, 45; P ϭ 0.0001), with Dn6 and Dn7 plants detected that leaf and root DWT and proportional displaying signiÞcantly less change in height than plant height change were signiÞcantly (P ϭ 0.05) less plants of Dn0 and Dnx (Table 3). Plants of KS041149 for CO960223 Dn6 than for Jagger Dn0 (data not Dnx had signiÞcantly greater changes in proportional shown). In addition, the leaf DWT value for KS041140 plant height than all the other genotypes, suggesting Dnx was signiÞcantly less than that of Jagger Dn0. that growth slowed as plants responded to D. noxia These results also support our Þndings that only feeding with the production of antibiosis defense re- CO960223 Dn6 is tolerant to RWA2. sponse compounds (Boyko et al. 2006). However, the Aphid number and leaf rolling (r ϭ 0.68), aphid proportional height value of CO960223 Dn6 plants was number and chlorosis (r ϭ 0.56), and leaf DWT and negative (infested plants were taller than noninfested root DWT (r ϭ 0.55) were signiÞcantly correlated plants), indicating that D. noxia feeding stimulated (P ϭ 0.05). In contrast, there was a weak negative CO960223 Dn6 plant growth. Puterka et al. (2006) correlation between plant height and aphid number found that in general, D. noxia feeding reduces the (r ϭϪ0.14). These results explain much of effect of growth of most resistant and susceptible barley, Hor- aphid infestation on plant phenotypic response (leaf deum vulgare (L.), genotypes. However, RWA5 rolling and chlorosis) as well as an expected positive caused slight growth stimulation to plants of some relationship between plant leaf and root tissue weight genotypes. change. In contrast, there seems to be little relation The TI values reßected trends similar to those between aphid population increase and plant height shown by the proportional leaf and root DWT changes measurement.

Table 3. Mean ؎ SE proportional change in leaf and root dry weight (DWT); plant height; and leaf, root, and plant height tolerance indices (TI) for ﬁve wheat genotypes after 14 d of D. noxia biotype 2 (RWA2) infestation under 24°C day:20°C night and a photoperiod of 14:10 (L:D) h

Wheat D. noxia resistance Mean Ϯ SE % proportional change Mean Ϯ SE tolerance index genotype gene Leaf DWTa Root DWTa Plant htb Leafc Rootc Plant htd Jagger Dn0 (none) 44.4 Ϯ 4.6a 30.3 Ϯ 5.9a 18.0 Ϯ 2.9b 0.34 Ϯ 0.07a 0.24 Ϯ 0.06a 0.13 Ϯ 0.03a Yumar Dn4 47.2 Ϯ 4.7a 10.7 Ϯ 8.8ab 16.3 Ϯ 3.2bc 0.41 Ϯ 0.06a 0.10 Ϯ 0.08a 0.16 Ϯ 0.05a CO960223 Dn6 4.5 Ϯ 6.7c Ϫ15.4 Ϯ 7.6c Ϫ3.1 Ϯ 7.1d 0.05 Ϯ 0.04b Ϫ0.18 Ϯ 0.09b Ϫ0.02 Ϯ 0.05c KS041140 Dnx 25.1 Ϯ 5.3b 24.5 Ϯ 6.4a 27.9 Ϯ 2.5a 0.39 Ϯ 0.09a 0.35 Ϯ 0.12a 0.45 Ϯ 0.09b 94M370 Dn7 5.4 Ϯ 5.8c Ϫ4.1 Ϯ 9.4bc 8.9 Ϯ 3.0cd 3.13 Ϯ 3.8ab 2.72 Ϯ 3.6ab 4.23 Ϯ 2.2abc

Means within a column followed by the same letter are not signiÞcantly different (␣ ϭ 0.05, PROC GLM; LSD). Experiments were conducted in October and November 2006. a DWT ϭ͓(dry weight (mg) of control plant Ϫ dry weight (mg) of infested plant)/dry weight (mg) of control plant͔ϫ100. b Proportional change in plant ht ϭ͓(ht (cm) of control plant Ϫ ht (cm) of infested plant)/ht (cm) of control plant͔ϫ100. c TI, DWT/number of aphids on infested plants. d Plant ht TI, proportional change in plant ht/number of aphids on infested plants. June 2009 LAZZARI ET AL.: FEEDING BEHAVIOR OF RUSSIAN WHEAT APHID 1297

2,4

1, 6

0,8

-2,4 -1,6 -0,8 0,8 1, 6 2,4 3,2 4

-0,8 Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021 Component 2 -1,6

-2,4

Component 1

Fig. 1. Principal component analysis of wheat genotypes Dn0 (ϩ), Dn4 (Ⅺ), Dn6 (Œ), Dn7 (E), and Dnx (छ). Components 1 (number of Diuraphis noxia biotype 2) and 2 (D. noxia induced leaf rolling) explained Ϸ80% of the variance (aphid number, 54.4%; leaf rolling, 25.5%). Dn7 antibiosis resistance was completely separate from all other genotypes. Dn6 tolerance was partially separate from other genotypes.

In contrast to the signiÞcant reductions in RWA1- RWA2 feeding). Although PCA responses of Jagger related leaf DWT loss on both Dn4 and Dn6 plants Dn0, Yumar Dn4, and KS041140 Dnx overlapped, some reported by Smith et al. (2005), our results indicated signiÞcant differences were found among these geno- that only CO960223 Dn6 plants exhibited reduced types for aphid number, leaf rolling, and leaf chlorosis. leaf DWT loss to RWA2. Despite the moderate Electronic Penetration Graph Analyses. There RWA2-related chlorosis and leaf rolling to were no signiÞcant differences (P ϭ 0.05) in RWA1 or

CO960223 Dn6, the proportional leaf DWT loss was RWA2 feeding on the F5 resistant (Dnx) or suscepti- very low and did not differ statistically from that of ble (Dn0) plants from the (PI 220127/SandoÕs Selec- 94M370 Dn7. However, the leaf TI value of tion) cross, or plants of CItr 2401 for several param- CO960223 Dn6, which was signiÞcantly less than eters, including mean time until initiation of Þrst that of the susceptible control, reinforced the va- probe, mean Þrst probe plus 8 h, mean difference in lidity of the tolerance of CO960223 Dn6 to RWA2. time from Þrst probe to Þrst SEP, mean available SEP, Smith et al. (2005) speculated that different grow- mean duration of E-waves, and duration of total path- ing environments could favor growth and related way phase. Although there were no signiÞcant differ- aphid resistance of plants containing Dn6 over that ences in the mean interval from the Þrst probe to the of plants containing Dn4. Also, plants possessing Þrst SEP event for either biotype, intervals were Dn6, which originated in Iran, may have been co- shorter for RWA1 (63 min on Dn0 plants, 110 min on evolutionarily exposed to greater levels of biotic Dnx plants) than RWA2 (193 min on Dn0, 123 min on stresses, resulting in more durable D. noxia resis- Dnx) (data not shown). tance than plants possessing Dn4. However, RWA1 aphids feeding on leaves of Dn0 Principal Component Analyses. The PCA of aphid plants produced signiÞcantly (P Ͻ 0.001) more total number, chlorosis, leaf rolling, leaf DWT, root DWT, SEP bouts than on leaves of Dnx plants or CItr 2401 and proportional plant height separated 94M370 Dn7, resistant control plants (Fig. 2). Individual RWA1 re- CO960223 Dn6, and KS041140 Dnx from Jagger Dn0 peatedly sampled phloem sap and engaged in com- and Yumar Dn4 (Fig. 1). Components 1 and 2 (aphid mitted phloem ingestion on all genotypes, but they number and leaf rolling) explained Ϸ80% of the vari- either received and or accepted less sap from leaves of ance among genotypes, which was signiÞcant (Jolliffe Dnx and CItr 2401 plants. Feeding of RWA2 yielded cutoff ϭ 0.7), whereas aphid number contributed the opposite result, producing signiÞcantly (P Ͻ 54.4% of the variance, and leaf rolling contributed 0.002) fewer total SEP bouts on Dn0 plants than on 25.5% of the variance. Numbers of aphids and plant leaves of Dnx or CItr 2401 plants (Fig. 2). damage symptoms, especially leaf rolling, were the The mean proportion (sum of all SEP bouts/avail- variables that explained the principal component able SEP activity) produced by RWA1 on leaves of groupings. The PCA results support results of antibi- Dn0 plants was also signiÞcantly (P Ͻ 0.005) greater osis and tolerance experiments (i.e., 94M370 Dn7 ex- than on Dnx plants, but RWA1 invested a signiÞ- hibits antibiosis and CO960223 Dn6 is tolerant of cantly greater proportion of sum SEP bouts per 1298 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 102, no. 3

8 b 7 CItr2401 Dnx 6 Dn0 a 5 a a a 4

3 a b SE total no. SEP events SEP total no. SE 2 ±

1 Mean

0 Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021 1 2 Biotype

Fig. 2. Mean Ϯ SE total number of E-wave (salivation and phloem sap ingestion with continuous salivation) events produced by D. noxia biotypes 1 and 2 on leaves of F5 plants of the bulk resistant population from [PI 220127 (Dnx)/SandoÕs Selection (Dn0)], from F5 plants of bulk susceptible population from [PI 220127 (Dnx)/SandoÕs Selection (Dn0)], and the resistant control genotype CItr 2401, containing two resistance genes, during an 8-h recording period. Means followed by a different letter differ signiÞcantly at P ϭ 0.05. RWA1: Dn0, n ϭ 15; Dnx, n ϭ 13; CItr 2401, n ϭ 8. RWA2: Dn0, n ϭ 5, Dnx, n ϭ 13; CItr 2401, n ϭ 11. available SEP on CItr 2401 plants than on Dnx or Dn0 plants. Our results are also in agreement with those plants (Fig. 3). Thus, RWA1 invested signiÞcantly cited in the review by Reese et al. (1994a), which less of the available SEP in either E1 (salivation) or noted that aphid resistance is often related to re- E2 (phloem sap ingestion) on leaves of Dnx plants duced ingestion of phloem contents. than on the either the resistant CItr 2401 or sus- The marked decrease in RWA1 phloem ingestion ceptible Dn0 control plants. As with RWA1, RWA2 on Dnx plants (Fig. 2) demonstrated that RWA1 invested signiÞcantly more sum E-wave duration may encounter any of several feeding inhibitors, time per available SEP on CItr 2401 plants than on digestibility reducing substances, or toxins shown to Dnx or Dn0 plants (Fig. 3). However, the results for be produced in Dnx plants during the initial6hof Dnx or Dn0 plants were reversed; RWA2 committed plant response to RWA1 feeding (C.M.S. et al., un- signiÞcantly (P Ͻ 0.001) more of the available SEP published). Alternatively, RWA1 may not perceive to feeding on Dnx than to feeding on Dn0. suitable or sufÞcient gustatory stimuli to maintain The reduced mean number of RWA1 SEP bouts committed phloem ingestion from leaves of Dnx and reduced sum of all SEP bouts/available SEP plants (Smith 2005). The lack of differences be- activity we observed in RWA1 feeding on Dnx plants tween Dnx and Dn0 plants in total pathway time are similar to results of Brewer and Webster (2001), required by either RWA1 or RWA2 is a strong in- who determined that the total duration of the sieve dication that Dnx resistance factors are likely not element phases was signiÞcantly shorter on RWA1- intercellular or intracellular within the leaf tissue resistant barley plants than on susceptible barley but rather in the sieve element cells or phloem sap,

1 a CItr2401 SE a b b Dnx ± 0.8 Dn0 c 0.6 c

0.4

0.2 Mean proportion (sum SEP bouts/available SEP)

0 1 2 Biotype

Fig. 3. Mean Ϯ SE sum of all E-waves (salivation and phloem sap ingestion with continuous salivation)/available SEP

(sieve element phase) produced by D. noxia biotypes 1 (RWA1) and 2 (RWA2) on leaves of F5 plants of the bulk resistant population from [PI 220127 (Dnx)/SandoÕs Selection (Dn0)], from F5 plants of bulk susceptible population from [PI 220127 (Dnx)/SandoÕs Selection (Dn0)], and the resistant control genotype CItr 2401, containing two resistance genes, during an 8-h recording period. Means followed by a different letter differ one signiÞcantly at P ϭ 0.05. RWA1: Dn0, n ϭ 15; Dnx, n ϭ 13; CItr 2401, n ϭ 8. RWA2: Dn0, n ϭ 5; Dnx, n ϭ 13; CItr 2401, n ϭ 11. June 2009 LAZZARI ET AL.: FEEDING BEHAVIOR OF RUSSIAN WHEAT APHID 1299 as indicated by the differences in the mean number phloem sap in Dnx plants ingested by RWA2 fail to and duration of salivation and phloem sap ingestion provide a diet that promotes growth of RWA2 at the events in Figs. 2 and 3. levels of plants containing Dn4, Dn6, and Dn0. Successful data sets for both biotypes on the Dnx and Although the speciÞc biochemical mechanism un- CItr 2401 genotypes were more difÞcult to acquire be- derlying the observed D. noxia feeding responses and cause of the aphids either not probing or becoming resulting antibiosis to plants containing the Dnx gene unglued from their wire tether. This trend also suggests is unknown, this behavior resembles that observed for the possibility of a feeding deterrent as a factor in re- D. noxia feeding on plants containing the Dn7 gene duced probing and feeding of RWA1 on Dnx plants and from rye. In sum, results of the current study indicate the lack of feeding stimulants for both biotypes feeding that plants containing Dnx exhibit antibiosis resistance on KS041140 Dnx and CItr 2401 plants. to RWA2 similar to that in plants containing the rye- In conclusion, our results indicate that RWA2- based Dn7 gene. The Dnx gene from wheat thus pro-

induced combined chlorosis and leaf rolling are vides a new potential source of antibiosis-based resis- Downloaded from https://academic.oup.com/jee/article/102/3/1291/2199217 by guest on 01 October 2021 signiÞcantly reduced in plants containing the Dn7 tance to RWA2, and is a signiÞcant improvement over and Dnx genes compared with those containing Dn4 the Dn7 from rye, previously the only candidate gene or Dn6. In addition, RWA2 population increase is known to confer RWA2 resistance in wheat. signiÞcantly reduced on 94M370 Dn7 and KS041140 Dnx plants compared with plants containing Yumar Dn4, CO960223 Dn6, or susceptible Jagger Dn0 con- Acknowledgments trol plants. In contrast to the results of Smith et al. (2005), who demonstrated antibiosis to RWA1 in We thank George Milliken (Department of Statistics, KSU); Susete C. Penteado, and Edilson B. Oliveira (Embrapa-Brazil); Dn4 and Dn6 plants at 21 d postinfestation, we found and Crisleide Lazzarotto and Josiane Cardoso (Department of no signiÞcant differences in the mean number of Zoology, Universidade Federal do Parana) for helping with RWA2 per plant between Jagger Dn0, Yumar Dn4, statistical analyses. We thank Ming-Shun Chen and Brian Mc- and CO960223Dn6. Cormack for critical reviews of the manuscript. This research Tolerance to RWA1 (reduced proportional change in was supported by a fellowship to from Conselho Nacional leaf DWT) was documented in Dn4, Dn6 and Dnx de Desenvolvimento Cientõ´Þco e Tecnolo´gico-Brazil (to S.L.) plants (Smith et al. 2005, Boyko et al. 2006). How- and by funding from the Department of Zoology, Universidade ever, we found that proportional changes in leaf and Federal do Parana´-Brazil, USDAÐARS Areawide Wheat Pest root DWT as well as plant height were also signif- Management grant AR-2472, and Kansas State University. This article is contribution 09-120-J from the Kansas Agricultural icantly reduced on RWA2-infested CO960223 Dn6 Experiment Station. and 94M370 Dn7 plants compared with Yumar Dn4 and KS041140 Dnx plants. When these measures were adjusted for differences in aphid population References Cited size, only CO960223 Dn6 plants exhibited tolerance to plant biomass reduction by RWA2. The PCA of all Backus, E. A. 1994. History, development, and applications of phenotypic variables indicated that aphid popula- the AC electronic monitoring system for insect feeding, pp. tion number and plant leaf rolling accounted for 1Ð51. In M. M. Ellsbury, E. A. Backus, and D. L. Ullman [eds.], Ϸ80% of the variance among the four genotypes History, development, and application of AC electronic insect feeding monitors. Thomas Say Publications in Entomol- evaluated, reinforcing the fact that RWA2 resis- ogy, Entomological Society of America, Lanham, MD. tance is primarily related to decreased aphid pop- Basky, Z. 2003. Biotypic and pest status differences between ulation development. Hungarian and South African populations of Russian The results of RWA2 EPG feeding analyses are wheat aphid, Diuraphis noxia (Kurdjumov) (Homoptera: less deÞnitive than those for RWA1. The fact that Aphididae). Pest Manag. Sci. 59: 1152Ð1158. RWA2 completed signiÞcantly more phloem sap Boyko, E. V., C. M. Smith, T. Vankatappa, J. Bruno, Y. Deng, ingestion events on Dnx plants than on Dn0 plants S. R. Starkey, and D. Klaahsen. 2006. The molecular ba- and expended a longer duration of SEP time per sis of plant gene expression during aphid invasion: wheat available SEP on Dnx plants is in strong contrast to Pto- and Pti-like sequences modulate aphid-wheat inter- action. J. Econ. Entomol. 99: 1430Ð1445. RWA1 feeding. However these biotype-speciÞc dif- Brewer, M. J., and J. A. Webster. 2001. Probing behavior of ferences, coupled with a lack of differences in total Diuraphis noxia and Rhopalosiphum maidis (Homoptera: time devoted to pathway behavior, support the con- Aphididae) affected by barley resistance to D. noxia and tention that Dnx resistance factors inßuencing plant water stress. Environ. Entomol. 30: 1041Ð1046. RWA2 are also phloem-based. Increased phloem Burd, J. D., D. R. Porter, G. J. Puterka, S. D. Haley, and F. B. ingestion of RWA2 on Dnx plants may be related to Peairs. 2006. Biotypic variation among North American the absence of a gustatory stimulant or to the pres- Russian wheat aphid (Homoptera: Aphididae) popula- ence of a RWA1-speciÞc feeding inhibitor that tions. J. Econ. Entomol. 99: 1862Ð1866. forces RWA2 to commit to longer periods to phloem Collins, M. B., S. D. Haley, F. B. Peairs, and J. B. Rudolph. 2005. Biotype 2 Russian wheat aphid resistance among ingestion (Powell et al. 2006). Although few behav- wheat germplasm accessions. Crop Sci. 45: 1877Ð1880. ioral observations of RWA2 have been documented, Diaz, J. D.-M., J. C. Reese, J. Louis, L. R. Campbell, and W. T. apertous RWA2 disperse faster by walking and ini- Schapaugh. 2006. Feeding behavior by the soybean tiate feeding more rapidly than RWA1 (C.M.S. et al., aphid (Hemiptera: Aphididae) on resistant and suscep- unpublished). Nevertheless, the components of tible soybean genotypes. J. Econ. Entomol. 100: 984Ð989. 1300 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 102, no. 3

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