Crop Protection 29 (2010) 415–421

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Crop Protection

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Influence of drought stress on Mexican rice borer (: ) oviposition preference in sugarcaneq

Allan T. Showler a,*, Boris A. Castro b,1 a USDA-ARS IFNRRU, Kika de la Garza Subtropical Agricultural Research Center, 2413 East Highway 83, Bldg. 201, Weslaco, TX 78596, USA b A&M AgriLife Research and Extension Center, Weslaco, TX 78596, USA article info abstract

Article history: Two sugarcane, Saccharum hybrids, varieties were grown in the greenhouse under well watered or Received 4 June 2009 drought conditions to examine the influences of stress on Mexican rice borer, loftini (Dyar), Received in revised form oviposition preference and selected nutritional components without impinging factors common to field 27 July 2009 conditions that alter responses. Our research revealed that, under controlled conditions, drought induced Accepted 29 July 2009 a wider range and greater uniformity of free amino acid (FAA) accumulations than have been previously reported. Drought stress resulted in increased dry leaf tissue and elevated concentrations of 7 of 9 Keywords: detectable free essential (for health) amino acids in stalks, the chief food of Mexican rice borer Eoreuma loftini Free amino acids larvae. Stressed sugarcane was preferred for oviposition, likely related to greater numbers of dry leaves Host plant selection and heightened host plant nutritional quality. Dry leaf tissue, which is not consumed, may be a cue for Resistance improving chances of larvae encountering nutrient-enhanced living tissue, and for concealing eggs in Water deficit folds. Excised dry leaf tissue from the treatments was indistinguishable for oviposition preference; therefore, biochemical status of living tissue may provide oviposition cues. Varieties exhibited no major genotypic differences in FAA accumulations or oviposition preference. Published by Elsevier Ltd.

1. Introduction It was projected to arrive in during 2008 and to infest all of the state’s rice and sugarcane production areas by 2035 (Reay- The Mexican rice borer, Eoreuma loftini (Dyar), originally from Jones et al., 2008). While insecticides can reduce Mexican rice borer Mexico, has been the key pest of sugarcane, Saccharum spp. hybrids, injury to sugarcane, yield increases have not often been reported in south Texas since spreading there in 1980 (Johnson and Van (Johnson, 1985; Meagher et al., 1994; Legaspi et al., 1999b), and use Leerdam, 1981) where it represents >95% of the stalk borer pop- for Mexican rice borer control in Texas was discontinued (Legaspi ulation (Legaspi et al., 1999a). The pest’s distribution has extended et al., 1997). Classical biological control using parasitoids also failed into the rice, Oryza sativa L., production region of east Texas (Meagher et al., 1998), but varietal resistance has shown some (Browning et al., 1989; Reay-Jones et al., 2007b), presumably promise (Reay-Jones et al., 2003, 2005, 2007a). assisted by >15 species of North American alternate host plants Infestations are especially severe where soil is saline (Reay- (Van Zwaluwenberg, 1926; Osborn and Phillips, 1946; Johnson, Jones et al., 2003), and under conditions of water deficit (Meagher 1984; Browning and Hussey, 1987) including other economically et al., 1993; Reay-Jones et al., 2005). Accumulation of plant free important crops such as corn, Zea mays L., sorghum, Sorghum amino acids (FAAs) and other nutrients is influenced by drought bicolor L. Moench, and rice (Van Zwaluwenberg, 1926; Osborn and (Rabe, 1994; Gzik, 1996; Showler, 2002, 2004; Showler et al., 2007), Phillips, 1946; Youm et al., 1988; Rodriguez-del-Bosque et al., 1996). including in sugarcane (Muquing and Ru-kai, 1998). Availability of amino acids is an important factor for growth, development, host selection, and reproduction of many herbivorous q Mention of trade names or commercial products in this publication is solely for (McNeil and Southwood, 1978; Showler, 2001, 2004; Nation, 2002), the purpose of providing specific information and does not imply recommendation and population buildups of some insect pests are linked to drought or endorsement by the U.S. Department of Agriculture. stress (Brodbeck and Strong, 1987; Ruberson et al., 1994; Mattson * Corresponding author. Tel.: 1 956 969 4882; fax: 1 956 969 5024. þ þ and Haack, 1991). The purpose of this study was to determine, E-mail address: [email protected] (A.T. Showler). 1 Present address: Dow AgroSciences, 7521 West California Ave., Fresno, CA under controlled greenhouse conditions, the influence of water- 93706, USA. deficit stress and two commonly grown cultivars on Mexican rice

0261-2194/$ – see front matter Published by Elsevier Ltd. doi:10.1016/j.cropro.2009.07.014 416 A.T. Showler, B.A. Castro / Crop Protection 29 (2010) 415–421 borer oviposition preference and on plant nutritional quality in treatment per assay). The juice (10–15 ml) was thoroughly mixed terms of FAAs absent extraneous impinging factors common to field for 2 min using a Virtishear homogenizer (Virtis, Gardiner, NY, conditions. USA). 5 ml from each sample was placed in separate 10 ml tubes and centrifuged at 10 000 rpm for 30 min. The extract samples 2. Materials and methods were stored at 80 C. 1 ml of supernatant from each sample was filtered through a 0.5 mm filter fitted to a 5 ml plastic syringe. 2.1. Large cage experiment – oviposition preference, whole plants Samples were placed in the autosampler of an Agilent 1100 Series (Agilent Technologies, Atlanta, GA, USA) reversed-phase high- Single sugarcane plants were grown from seed pieces in 7.6-l performance liquid chromatograph (HPLC) with a binary pump pots until they reached 1.2 m in height (9 nodes) in a greenhouse at delivering solvent A [1.36 g sodium acetate trihydrate þ 500 ml the USDA-ARS Kika de la Garza Subtropical Agricultural Research purified HPLC grade water þ 90 ml triethylamine (TEA) þ sufficient Center in Weslaco, Hidalgo County, Texas. The soil was Sunshine acetic acid to bring the pH to 7.2] and solvent B [1.36 g sodium mix no. 1 nursery potting soil (w75% sphagnum peat moss, perlite, acetate trihydrate þ 100 ml purified HPLC grade water (acetic acid dolomitic limestone, and gypsum; Sungro Horticulture, Bellevue, added to this mixture to bring the pH to 7.2) þ 200 ml acetoni- Washington). When the plants reached the two-leaf stage, 200 ml trile þ 200 ml methanol] at 100 and 1.0 ml/min on a Zorbax Eclipse of Peters Professional (Scotts-Sierra Horticultural Products AAA 4.6 150 mm 3.5m column (Agilent Technologies). Absor- Company, Marysville, OH) water-soluble general purpose 20–20– bances at 262 and 338 nm were monitored on a variable wave- 20 N–P–K fertilizer at 15.8 g/l of water was applied to the soil. Half length detector for 48 min per sample. The autosampler measured of the plants were variety L97-128 (not regarded as being resistant and mixed 6 ml sodium borate buffer (0.4 N, pH 10.2 in water), 1 ml to the sugarcane borer, Diatraea saccharalis [F.] (White et al., 2008), 9-fluorenylmethylchloroformate (FMOC), and 1 ml ophthalaldehyde and not listed in the literature as being resistant to the Mexican rice (OPA) derivitizing agents, and 2 ml of sample, then injected 2 ml for borer) and the other half were variety CP70-321 (resistant to the chromatographic separation of FAAs. Identification and quantifi- sugarcane borer (Bessin et al., 1990) and to the Mexican rice borer cation of 17 derivitized FAAs (alanine, arginine, aspartic acid, (Pfannenstiel and Meagher, 1991; Legaspi et al., 1999a)). Twelve cystine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, pots of each variety were placed within 3 m 1.2 m 1.8 m methionine, phenylalanine, proline, serine, threonine, tyrosine, and (l w h) cages (36 mesh/cm2) inside the greenhouse. Six plants valine) were achieved by calibrating with a standard mixture of of each variety were well watered and the other six were drought amino acids. Peak integration accuracy was enhanced by manual stressed. Soil in the well watered pots was kept constantly moist establishment of peak baselines using Agilent software. Results are with infusions of water as needed. The drought stressed plants presented as picomoles of FAA per microliter of juice. received only 2000 ml of water at the beginning of each week in Differences among the four treatments were found using one- a single pulse. The two watering regimes were implemented when way ANOVA and means were separated using Tukey’s HSD the plants were 1.2 m tall and for the next 6 wk before the assays (Analytical Software, 1998). Because the assays were conducted in commenced. Six replicates of each variety-irrigation treatment different cages and at different times, the data from the assays were combination were arranged in a randomized complete block not pooled. Overall differences between varieties or irrigation design. The experiment was repeated three times, and each, regimes were detected using a 2 2 factorial design (Analytical referred to as assays 1, 2, and 3, was conducted z1 mo apart from Software, 1998). the other assays and in different cages. Adult Mexican rice borers were collected as larvae in commer- 2.2. Small cage experiment 1 – oviposition preference for dry leaf cial sugarcane fields in Hidalgo County to generate a colony of tissue, all four treatments Mexican rice borers at the Texas A&M University System’s AgriLife Research and Extension Center in Weslaco. The colony was reared Sugarcane plants were grown under the same treatment on artificial diet (Martinez et al., 1988)at25C, 65% RH, and conditions as in the whole plant assays. To test whether dry leaf a photoperiod of 14:10 (L:D). Pupae were separated by sex, and one tissue alone provides chemical or visual cures that affect oviposi- emerged adult female was placed in a 3.8-l plastic container with tion preference, a twenty-cm long distal section of dry (completely two adult males for 48 h to mate. Thirty mated females were brown) leaf from the basal half of each of seven different sugarcane released into each greenhouse cage (7–8 females were released in plants was excised per cultivar per treatment 6 d after the last each of the four cage corners to reduce point-of-release effects). irrigation. The 28 leaf sections were suspended by their basal ends Females were allowed to lay eggs for 5 d. using paper clips attached to a string stretched across the top of Numbers of dry leaves (at least the distal half) on each plant a30 30 60 cm (l w h) cage in a completely randomized were counted on the day of Mexican rice borer release into each design, and a 1 cm3 ball of cotton soaked with water was placed at cage. At the same time, one fully expanded leaf per plant was the bottom of each of three cages (each cage was a separate assay). excised and its water potential measured with a Model 610 (PMS Ten mated adult female Mexican rice borers were released at the Instrument, Corvallis, Oregon) pressure bomb (n ¼ 6 leaves from bottom of each assay cage and permitted to lay eggs for 2 d, after separate plants per treatment per assay). After 5 d, all of the leaves which numbers of egg clusters and total numbers of eggs per leaf on each plant were visually inspected and the numbers of egg were recorded. Data were analyzed with ANOVA and means were clusters and eggs/cluster were recorded. Numbers of egg clusters separated using Tukey’s HSD (Analytical Software, 1998). Differ- and total eggs per dry leaf were calculated. Five weeks later, each ences between varieties and irrigation regimes were analyzed plant was stripped of leaves and leaf sheaths, and the stalk was using a 2 2 factorial design (Analytical Software, 1998). visually inspected for Mexican rice borer entry and exit holes. Sugarcane juice was pressed, using a vise, from freshly cut 2.3. Small cage experiment 2 – oviposition preference for dry leaf internodes from the lower third of the stalks, when the Mexican tissue, two irrigation treatments rice borers were released in the large cages. The samples were taken from extra sugarcane plants that had been grown under the Sugarcane plants (var. CP70-321) were grown under the two same conditions as those used in the large cage assays, and each irrigation regimes as in the whole plant assays. Excised dry leaf sample was taken from a separate plant (5 plants per cultivar per sections from seven different sugarcane plants in each treatment A.T. Showler, B.A. Castro / Crop Protection 29 (2010) 415–421 417 were suspended in the Plexiglass cages as in the first small cage were not different in the first two assays, but in the third assay, experiment, and mated female Mexican rice borers were released clusters per dry leaf were 10-fold more numerous (F ¼ 9.35, df ¼ 1, as in small cage experiment 1. After 2 d, the leaf sections were 20, P ¼ 0.0001) and eggs per dry leaf were 8.6-fold more abun- removed and numbers of egg clusters and total eggs were recorded. dant (F ¼ 6.87, df ¼ 1, 20, P ¼ 0.0039) on drought stressed than on Data were analyzed using the two-sample t test (Analytical Soft- well watered sugarcane. Larval entry and adult exit holes were from ware, 1998). The experiment was repeated three times. 1.8- to 9.3-fold and from 2.8- to 6-fold, respectively, more numerous on drought stressed sugarcane of either variety (Table 1). Factorial analysis indicated that the stress was associated with from 2.4. Small cage experiment 3 – oviposition preference for dry leaf 2- to 15-fold and from 3- to 15-fold more entry and exit holes, tissue, two cultivars respectively (entry holes, F 39.60, df ¼ 1, 20, P < 0.0001; exit holes, F 36.87, df 1, 20, P < 0.0001). Significant interactions Sugarcane plants of both varieties were grown under the ¼ were not detected. drought irrigation regime as described in the whole plant assays. Sixteen FAAs were detected in the samples from each of the four Excised dry leaf sections from seven different sugarcane plants in treatments in all three assays, and effects of drought stress and/or each treatment were suspended in the Plexiglass cages as in the cultivar were apparent in at least two of the three assays, excluding first small cage experiment, and mated female Mexican rice borers aspartic acid (Table 2). Factorial analyses showed that glycine, were released as in small cage experiment 1. After 2 d, the leaf histidine, isoleucine, leucine, methionine, phenylalanine, proline, sections were removed and numbers of egg clusters and total eggs serine, threonine, tyrosine, and valine were more abundant in were recorded. Data was analyzed using the two-sample t test drought stressed sugarcane in all three assays, and alanine, gluta- (Analytical Software, 1998). The experiment was repeated three mic acid, and lysine were more abundant in two of the three assays times. (Table 3). Factorial analyses detected varietal differences in 14 FAAs, but results were inconsistent between assays (Table 4). 3. Results Free essential (for insect growth and development) amino acids detected in this study were arginine, histidine, isoleucine, 3.1. Large cage experiment – oviposition preference, whole plants leucine, lysine, methionine, phenylalanine, threonine, and valine (tryptophan, the only other essential amino acid for , could Water potential in drought stressed sugarcane among the three not be detected with our instrumentation), and seven of those assays was 2.5-fold that of well watered sugarcane of either were more concentrated than the drought stressed sugarcane of variety (Table 1). Dry leaves were from 2.5- to 3.4-fold more both varieties (Table 2). Total free essential amino acids and total abundant on the stressed plants (Table 1). Factorial analysis showed free amino acids in the drought stressed sugarcane were consis- dry leaves to be from 3.4- to 4.9-fold more abundant in all three tently more concentrated than in well watered plants (Tables 2 assays (F value presented is the lowest of the three) (F 82.05, and 3). df ¼ 1, 20, P < 0.0001) on drought stressed plants than on well watered plants. Egg clusters and total eggs per plant ranged from being 3- to 11-fold and from 4.4- to 8.7-fold more numerous, 3.2. Small cage experiments 1–3 respectively, in the drought stressed treatments (Table 1). Factorial analysis determined that clusters and total eggs were from 5.8- to Eggs were laid exclusively on the dry leaf tissue provided in 19.5-fold and from 7.8- to 24.4-fold, respectively, more numerous these experiments, but treatment differences were not detected for (clusters, F 21.05, df ¼ 1, 20, P 0.0002; eggs, F 15.56, df ¼ 1, 20, numbers of egg clusters or for total numbers of eggs per dry excised P 0.0008). Numbers of egg clusters and total eggs per dry leaf leaf. Factorial analysis also failed to detect differences.

Table 1 Mean (SE) water potential (bar), and numbers of dry leaves, Mexican rice borer egg clusters, total eggs, entry holes, and exit holes per stalk of two sugarcane varieties maintained under well watered or drought stressed greenhouse conditions.

Measurement Assay FPa Treatmentb

L97-128 W CP70-321 W L97-128 D CP70-321 D Water potential 1 1177.41 <0.0001 9.2 0.5b 10.3 0.36b 30.0 0.2a 30.0 0.2a No. dry leaves 25.16 <0.0001 2.0 1.0b 3.8 0.8b 9.5 0.8a 10.3 0.4a No. egg clusters 7.26 0.0025 0.3 0.3b 0.5 0.3b 2.7 0.6a 2.0 0.4a No. eggs 6.93 0.0038 4.7 4.7b 5.0 3.2b 48.0 13.7a 29.3 8.5a No. entry holes 20.33 <0.0001 5.8 0.5b 4.8 0.5b 10.2 1.2a 10.8 1.2a No. exit holes 12.28 0.0003 1.7 0.4b 1.5 0.2b 4.7 0.7a 5.0 0.7a

Water potential 2 1036.63 <0.0001 11.5 0.6c 14.3 0.8b 30.0 0.2a 30.0 0.2a No. dry leaves 43.90 <0.0001 1.0 0.4c 3.3 0.3c 12.5 1.2a 8.7 0.2b No. egg clusters 5.81 0.0077 1.0 0.8b 0.2 0.2b 4.8 1.9a 3.2 1.4b No. eggs 7.16 0.0033 10.0 4.5b 1.0 1.0b 62.2 21.4a 44.0 5.3a No. entry holes 5.42 0.0068 0.3 0.2b 0.2 0.2b 3.2 1.1a 2.8 1.1a No. exit holes 5.48 0.0065 0.3 0b 0b 1.0 0.4a 1.2 0.5a

Water potential 3 1082.76 <0.0001 7.6 0.5b 6.0 0.6b 30.0 0.8a 30.0 1.4a No. dry leaves 52.43 <0.0001 1.7 0.5b 2.5 0.8c 12.2 0.9a 8.5 0.2b No. egg clusters 37.57 <0.0001 0b 0.3 0.3b 4.5 1.3a 3.3 0.6a No. eggs 46.49 <0.0001 0b 4.0 4.0b 63.0 20.3a 34.7 9.4a No. entry holes 9.34 0.0005 3.2 1.6bc 0.3 0.2c 10.0 2.3a 5.5 1.2ab No. exit holes 4.68 0.0124 0.2 0.2b 0b 1.8 0.6a 1.2 0.5a

a One-way ANOVA, randomized complete block design, df ¼ 1, 20. b W, well watered; D, drought stressed. 418 A.T. Showler, B.A. Castro / Crop Protection 29 (2010) 415–421

Table 2 Mean (SE) picomoles of free amino acid per microliter of sugarcane stalk juice in two varieties, L97-128 and CP70-321, that were well watered or drought stressed.

Free amino acidsa Assay FPb Treatmentc

L97-128 W CP70-321 W L97-128 D CP70-321 D Alanine 1 3.55 0.0480 7323 1858ab 3478 1124b 15 078 1847a 7857 2167ab Arginine 6.45 0.0075 1358 347a 462 129b 1272 188a 855 132ab Aspartic acid 1.34 0.308 2533 257 2443 157 1996 187 2118 289 Glutamic acid 13.07 0.0004 6 6ab 149 69ab 29 13bc 860 206a Glycine 5.36 0.0142 484 144b 441 81b 1161 87a 653 105ab Histidine 11.47 0.0008 424 103b 938 199b 1618 172a 1995 211a Isoleucine 16.68 0.0001 703 173c 1263 145b 1916 332a 2858 215a Leucine 17.75 0.0001 731 150b 946 127b 1939 285a 2639 285a Lysine 6.15 0.0090 708 91a 328 87b 639 92a 514 68ab Methionine 16.18 0.0002 384 57b 228 76b 1224 226a 1241 250a Phenylalanine 19.73 0.0001 239 59b 229 23b 523 84a 1008 123a Proline 16.89 0.0001 421 70b 497 128b 1674 520a 4062 903a Serine 5.51 0.0130 3612 809b 4290 892ab 6875 737ab 8875 1125a Threonine 9.01 0.0021 1464 251b 2568 484ab 2863 353a 3887 306a Tyrosine 32.51 <0.0001 209 24c 138 7c 318 31b 515 58a Valine 12.63 0.0005 1826 391b 3188 470a 3871 490a 5584 351a Free essential amino acids 11.02 0.0009 7841 1248b 10 153 1686b 15 365 1678a 20 585 1474a Total FAAs 6.92 0.0059 22 432 4034b 21 592 3865b 42 498 4143a 45 528 4159a

Alanine 2 6.33 0.0081 3569 193b 5836 1324b 12 257 1423a 9102 2277ab Arginine 18.06 0.0001 1436 185a 677 52b 1084 93a 1324 35a Aspartic acid 2.80 0.0855 2074 158 2827 233 2516 61 2682 311 Glutamic acid 5.26 0.0151 18 18b 7 7b 126 43a 80 8a Glycine 8.25 0.0030 396 52b 505 37b 623 83ab 1072 173a Histidine 10.33 0.0012 598 85b 577 77b 1642 218a 1613 273a Isoleucine 20.09 0.0001 605 57b 565 54b 1293 204a 923 47a Leucine 7.97 0.0034 441 79b 510 52b 776 23a 798 88a Lysine 4.25 0.0291 367 94b 375 57ab 650 80ab 752 65a Methionine 37.52 <0.0001 225 20b 239 64b 771 88a 949 168a Phenylalanine 20.26 0.0001 258 43b 226 49b 908 88a 731 88a Proline 18.46 0.0001 558 36b 518 99b 1198 89a 1455 51a Serine 17.09 0.0001 2306 285c 4315 623b 4468 913b 8109 931a Threonine 6.35 0.0080 1490 95b 2258 192ab 2954 444a 3431 510a Tyrosine 2.58 0.1025 209 24 186 26 357 84 311 60 Valine 5.42 0.0137 1193 260b 1563 30ab 2797 502a 2298 259a Free essential amino acids 23.52 <0.0001 6616 145b 6992 68b 12 878 1472a 12 823 983a Total FAAs 19.17 0.0001 15 749 540c 21 189 2108bc 24 424 2837b 35 576 2656a

Alanine 3 28.64 <0.0001 1246 91b 694 310b 7173 718a 5322 889a Arginine 7.78 0.0038 211 10ab 166 37b 356 28a 416 55a Aspartic acid 1.99 0.1695 1135 24 907 90 1310 192 1373 175 Glutamic acid 2.99 0.0734 525 118 354 30 577 85 365 40 Glycine 26.53 <0.0001 354 33b 236 52c 778 77a 547 43a Histidine 9.15 0.0020 153 13b 222 33b 535 22a 861 108a Isoleucine 25.50 <0.0001 246 22b 250 125b 1597 115a 2537 370a Leucine 27.41 <0.0001 205 27b 206 96b 1271 102a 1988 280a Lysine 11.47 0.0008 122 13b 118 29b 320 43a 383 49a Methionine 49.12 <0.0001 59 3b 83 28b 774 68a 858 149a Phenylalanine 44.21 <0.0001 41 1b 64 13b 203 26a 369 44a Proline 95.77 <0.0001 322 35c 211 17d 584 29b 1371 144a Serine 22.72 <0.0001 502 35b 301 124b 2392 299a 2510 457a Threonine 21.80 <0.0001 469 28b 265 117b 1657 119a 2379 323a Tyrosine 222.47 <0.0001 181 8b 70 6c 384 5a 481 49a Valine 20.20 0.0001 918 58b 492 230b 2792 218a 4032 579a Free essential amino acids 23.18 <0.0001 2427 167b 1710 706b 9509 741a 13 827 1971a Total FAAs 30.27 <0.0001 6693 333b 4486 1266b 22 711 1976a 25 797 3309a

Means within each row followed by different letters are significantly different (P < 0.05). a Cystine was detectable but not found in the samples. b One-way ANOVA, randomized complete block design, df ¼ 3, 12. c W, well watered; D, drought stressed.

4. Discussion help explain observed differences in leaf desiccation, physi- ochemical aspects, and Mexican rice borer oviposition preference. Drought stress was associated with relatively high numbers of Because excised dry leaf tissue (small cage experiments) presented dry leaf tissue and Mexican rice borer eggs. Although plants with to Mexican rice borers in the absence of living tissue did not 30 bar water potential were alive, they were highly stressed as influence oviposition, it appears that dry leaf tissue alone does not indicated by the abundance of dry leaf tissue. While we could not influence oviposition preference regardless of whether or not the perform correlative tests because exact measures of water poten- tissue originated on drought stressed plants or on one or the other tial over 30 bar were unavailable, the difference between the well varieties used in this study. While the quantity of dry leaf tissue watered and drought stressed treatments was substantial and can offers more potential oviposition sites, and the tight folds and curls A.T. Showler, B.A. Castro / Crop Protection 29 (2010) 415–421 419

Table 3 Factorial analyses of FAA accumulations (pmoles/ml sugarcane stalk juice) between well watered and drought stressed treatments, 3 assays, n ¼ 5 replications in each assay.

FAA Assay 1a Assay 2a Assay 3a

Differenceb Highc FP Differenceb Highc FP Differenceb Highc FP Alanine 2.1 D 11.55 0.0034 – – 0.48 0.4983 6.4 D 78.11 <0.0001 Arginine – – 0.49 0.4931 – – 0.84 0.3712 2.0 D 27.38 0.0001 Aspartic acid – – 3.77 0.0690 – – 0.46 0.5062 1.3 D 5.37 0.0333 Glutamic acid 5.7 D 7.42 0.0144 5.9 D 5.40 0.0328 – – 0.18 0.6786 Glycine 2.0 D 14.04 0.0016 1.9 D 13.80 0.0017 2.2 D 45.92 <0.0001 Histidine 2.3 D 24.41 0.0001 2.8 D 33.83 <0.0001 6.5 D 60.72 <0.0001 Isoleucine 2.4 D 38.60 <0.0001 1.9 D 20.43 0.0003 8.3 D 63.51 <0.0001 Leucine 2.7 D 4.36 <0.0001 1.7 D 23.47 0.0002 7.9 D 65.64 <0.0001 Lysine – – 0.44 0.5178 1.9 D 19.82 0.0003 2.9 D 40.25 <0.0001 Methionine 4.0 D 29.23 <0.0001 3.7 D 39.43 <0.0001 11.4 D 83.87 <0.0001 Phenylalanine 3.3 D 28.72 0.0001 3.4 D 66.66 <0.0001 5.5 D 56.72 <0.0001 Proline 6.2 D 17.09 0.0007 2.5 D 72,44 <0.0001 3.7 D 29.23 <0.0001 Serine 2.0 D 19.40 0.0004 1.9 D 16.11 0.0009 6.1 D 55.51 <0.0001 Threonine 1.7 D 15.19 0.0012 1.7 D 14.49 0.0014 5.5 D 51.31 <0.0001 Tyrosine 2.4 D 26.41 0.0001 1.7 D 6.61 0.0198 3.4 D 76.34 <0.0001 Valine 1.9 D 28.06 0.0001 1.8 D 12.85 0.0023 4.8 D 50.78 <0.0001 Free essential amino acids 2.0 D 34.48 <0.0001 1.9 D 48.97 <0.0001 5.6 D 59.81 <0.0001 Total FAAs 2.0 D 30.88 <0.0001 1.6 D 25.83 0.0001 4.3 D 80.8 <0.0001

a df ¼ 1, 19. b Expressed as the greater treatment mean across both cultivars divided by the lower mean; –, no significant difference (P > 0.05). c The treatment factor with the greatest accumulation of the FAA(s); D, drought stressed; –, no difference between factors detected. characteristic of dry sugarcane leaf tissue provides some protection changes in water-deficit stressed plants can favor insect feeding, for eggs (Van Leerdam et al., 1986; Reay-Jones et al., 2007a,b; growth, or reproduction (White, 1984; Mattson and Haack, 1991), Showler, unpublished data), the presence of dry leaves might also and balances among water, protein, carbohydrates, lipids, essential signal the existence of associated concentrated nutrients, such as amino acids and other nutrients are important to host plant FAAs, in attached living tissue. As an example of the connection selection and for insect survival (Vanderzant, 1958, 1963; Blaney between Mexican rice borer oviposition preference and nutritional and Simmonds, 1988; Mattson and Haack, 1991; Nation, 2002; quality of the host plant absent dry leaves, young sugarcane Chapman, 2003). Drought stress-associated FAA accumulations, for (5 nodes), despite having fewer dry leaves than older sugarcane example, have been associated with high populations of herbivo- (10 nodes), was preferred by the Mexican rice borer, a trend rous insects (Jayaraj and Seshadri, 1967; White, 1984; McQuate and associated with higher levels of free essential amino acids in the Connor, 1990; Blua et al., 1994) and may promote insect outbreaks young plants which had no dry leaves (Reay-Jones et al., 2007a). (Brodbeck and Strong, 1987; Mattson and Haack, 1991). In sugar- Many biotic and abiotic stresses on plants are associated with cane, Reay-Jones et al. (2005) reported elevated FAA accumulations accumulations of FAAs (Joshi and Naik, 1980; Singh et al., 1993; in drought stressed field conditions, but our study found 42% Rabe, 1994; Gzik, 1996; Bussis and Heineke, 1998; Showler et al., more diversity in free essential amino acids with consistently 1990, 2007; Showler, 2002) that heighten the plant’s nutritional elevated accumulations. value and affect herbivorous insect preference and performance While we recognize the importance of carbohydrates, lipids, and (Helms et al., 1971; McNeil and Southwood, 1978; Rhoades, 1983; other nutritional constituents, amino acids are as well important to White, 1984; Brodbeck and Strong, 1987). Protein and FAA content insect growth and development (Vanderzant, 1958, 1963; Nation,

Table 4 Factorial analyses of FAA accumulations (pmoles/ml sugarcane stalk juice) in varieties L97-128 and CP70-321, 3 assays, n ¼ 5 replications in each assay.

FAA Assay 1a Assay 2a Assay 3a

Differenceb Highc FP Differenceb Highc FP Differenceb Highc FP Alanine 2.0 L 9.61 0.0065 2.6 C 10.42 0.0049 – – 4.05 0.0604 Arginine 2.0 L 8.94 0.0082 – – 2.62 0.1239 – – 0.04 0.8364 Aspartic acid – – 0.01 0.9438 – – 4.44 0.0504 – – 0.36 0.5569 Glutamic acid 27.9 C 3.06 0.0021 5.3 L 6.83 0.0388 1.5 L 6.52 0.0206 Glycine 1.5 L 5.39 0.0329 1.5 C 6.83 0.0182 1.4 L 10.34 0.0051 Histidine 1.9 C 15.48 0.0011 – – 0.02 0.8892 – – 2.40 0.1400 Isoleucine 1.6 L 11.06 0.0040 – – 3.14 0.0942 – – 4.28 0.0542 Leucine – – 4.12 0.0584 – – 0.51 0.4840 – – 4.18 0.0568 Lysine 1.6 L 8.15 0.0110 – – 0.54 0.4726 – – 0.64 0.4331 Methionine – – 0.16 0.6901 – – 0.23 0.3476 – – 0.44 0.5164 Phenylalanine 1.6 C 5.75 0.0282 – – 2.17 0.1587 1.8 C 9.25 0.0074 Proline 2.2 C 4.47 0.0495 – – 1.37 0.2576 1.7 C 6.61 0.0198 Serine – – 2.26 0.1511 1.8 C 4.51 0.0014 – – 0.02 0.8821 Threonine 1.5 C 9.30 0.0072 – – 3.23 0.0903 – – 1.26 0.2772 Tyrosine – – 1.79 0.1990 – – 0.42 0.5248 – – 0.04 0.8357 Valine 1.5 C 13.45 0.0019 – – 0.04 0.8457 – – 1.15 0.2986 Free essential amino acids 1.3 C 6.07 0.0247 – – 0.03 0.8550 – – 2.10 0.1652 Total FAAs – – 0.08 0.7854 1.4 C 13.37 0.0020 – – 0.04 0.8349

a df ¼ 1, 19. b Expressed as the greater treatment means across both drought and well watered treatments divided by the lower mean; –, no significant difference (P > 0.05). c The treatment factor with the greatest accumulation of the FAA(s); L, L97-128; C, CP70-321; –, no difference between factors detected. 420 A.T. Showler, B.A. Castro / Crop Protection 29 (2010) 415–421

2002). It is unlikely, however, that carbohydrates are a limiting cultivar in their study, LCP85-384, was more susceptible to Mexican factor in a plant like sugarcane, although other kinds of nutrients rice borer injury than another, HoCP85-845, which also had greater might be associated with Mexican rice borer preference. We concentrations of FAAs, based on percentages of bored sugarcane focused on free amino acids as indicators of the nutritional quality internodes and emergence. Judicious selection of water- of the plant. Most dietary nitrogen is absorbed by insects through retentive sugarcane varieties with relatively little dry leaf tissue the gut, and FAAs are nutritionally superior to proteins, more and low nutritional quality for Mexican rice borer larvae, or available for rapid assimilation than proteins, and more physically amendment of soil to enhance water conservation in the field to accessible because of increased solubility (Brodbeck and Strong, achieve the same conditions might be fruitful tactics for suppres- 1987; Cockfield, 1988). Acceptance of a particular host plant by sion of injury caused by this invasive pest. many is mediated by contact chemoreceptors on the antennae, proboscis, tarsi, and ovipositor which detect presence or absence of attractive or repellent primary and secondary Acknowledgments compounds, including amino acids (Sta¨dler, 1984; Blaney and Simmonds, 1988; Ramaswamy, 1988) by contact with the leaf We thank Veronica Abrigo, Jaime Cavazos, Matthew Rector, and surface or by sensing plant volatiles. Detection of host plant Jennifer Plata for technical assistance, and Francis Reay-Jones, biochemicals and nutritional status can affect oviposition prefer- Robert Meagher, Jesusa Legaspi, and Mamoudou Setamou for crit- ence (Singer, 1972, 1983; Showler, 2001), and levels of free essential ical reviews. amino acids in living tissue are associated with the greater numbers of eggs found on drought stressed sugarcane. Decreasing water- deficit stress by irrigating might assist in suppressing Mexican rice References borer oviposition by minimizing numbers of dry leaves and Analytical Software, 1998. Statistix for Windows. Analytical Software, Tallahassee, diminishing the nutritional quality of living tissues. FL, USA. Drought stressed sugarcane varieties LCP85-384 and HoCP85- Bessin, R.T., Reagan, T.E., Martin, F.A., 1990. A moth production index for evaluating 845 under field conditions at Ganado, Jackson County (center: 28 sugarcane cultivars for resistance to sugarcane borer (Lepidoptera: Pyralidae). J. 570 N, 96 350 W), Texas, had elevated levels of four free essential Econ. Entomol. 83, 221–225. Blaney, W.M., Simmonds, M.S.J., 1988. Food selection in adult and larvae of three amino acids, as well as four ‘‘nonessential’’ amino acids (Reay-Jones species of Lepidoptera: a behavioral and electrophysiological study. Entomol. et al., 2005). In a greenhouse study (Reay-Jones et al., 2007a), using Exp. Applic. 49, 111–121. those varieties, found drought stress was associated with accu- Blua, M.J., Perring, T.M., Madore, M.A., 1994. Plant virus-induced changes in aphid population development and temporal fluctuations in plant nutrients. J. Chem. mulations of three free essential amino acids. Overlap between the Ecol. 20, 691–707. two studies (Reay-Jones et al., 2005, 2007a) in terms of specific free Brodbeck, B., Strong, D., 1987. Amino acid nutrition of herbivorous insects and stress essential amino acids consisted of leucine only. Differences in to host plants. In: Barbosa, P., Schultz, J.C. (Eds.), Insect Outbreaks. Academic, London, pp. 346–364. specific FAAs affected by water-deficit stress in the Reay-Jones et al. Browning, H.W., Hussey, M.A., 1987. Susceptibility of ‘Tifton 68’ and ‘Coastal’ ber- (2005, 2007a) studies were probably related to impinging envi- mudagrass to the Mexican rice borer. 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