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1 University of Nersk - Lincoln DigitlCommons@University of Nersk - Lincoln Pulictions from USDA-ARS / UNL Fculty U.S. Deprtment of Agriculture: Agriculturl Reserch Service, Lincoln, Nersk 2006 Host specificity of divergent popultions of the lef eetle Diorhd elongt (Coleopter: Chrysomelide), iologicl control gent of sltcedr (Tmrix spp.) Lindsey R. Milrth C. Clrk DeLoch Follow this nd dditionl works t: Prt of the Agriculturl Science Commons Milrth, Lindsey R. nd DeLoch, C. Clrk, "Host specificity of divergent popultions of the lef eetle Diorhd elongt (Coleopter: Chrysomelide), iologicl control gent of sltcedr (Tmrix spp.)" (2006). Pulictions from USDA-ARS / UNL Fculty This Article is rought to you for free nd open ccess y the U.S. Deprtment of Agriculture: Agriculturl Reserch Service, Lincoln, Nersk t DigitlCommons@University of Nersk - Lincoln. It hs een ccepted for inclusion in Pulictions from USDA-ARS / UNL Fculty y n uthorized dministrtor of DigitlCommons@University of Nersk - Lincoln.

2 Biologicl Control 36 (2006) Host speciwcity of diverent popultions of the lef eetle Diorhd elongt (Coleopter: Chrysomelide), iologicl control gent of sltcedr (Tmrix spp.) Lindsey R. Milrth, C. Jck DeLoch United Sttes Deprtment of Agriculture, Agriculturl Reserch Service, Grsslnd, Soil nd Wter Reserch Lortory, 808 E. Blcklnd Rod, Temple, TX 76502, USA Received 11 Ferury 2005; ccepted 16 Septemer 2005 Aville online 2 Novemer 2005 Astrct The lef eetle, Diorhd elongt (Brullé) sensu lto, ws relesed in 2001 for the clssicl iologicl control of exotic sltcedrs, complex of invsive Tmrix species nd hyrids. It did not estlish t sites south of 37 N ltitude where summer dylengths re elow the criticl photoperiod of the northern-dpted popultions of the eetle tht were relesed. Therefore, we ssessed the host speciwcity of four D. elongt popultions collected from more southern ltitudes in the Old World (Tunisi, Crete, Uzekistn, nd Turpn, Chin). All popultions were similr to ech other nd the previously relesed popultions of D. elongt in their host speciwcity. Lrvl/pupl survivl for ll popultions ws % on Tmrix test plnts, 0 76% on ntive Frnkeni plnts (oth in the order Tmricles), nd 0% on the remining 28 species of plnts on which ll the lrve died s 1st instrs. D. elongt lid high numers of eggs on sltcedr, generlly fewer eggs on thel ( modertely vlued evergreen species of Tmrix) except for Uzekistn eetles, nd few to no eggs on three species of Frnkeni. Few to no dults were found on Frnkeni plnts which lso were poor mintennce hosts. The relese of ny of the four D. elongt popultions in the southern US nd northern Mexico should pose no risk to plnts outside the order Tmricles nd low risk to ntive, non-trget Frnkeni plnts. Athel my e less dmged thn sltcedr. Pulished y Elsevier Inc. Keywords: Host rnge; Host speciwcity; Clssicl iologicl control; Weeds; Weed iologicl control; Diorhd elongt; Sltcedr; Tmrix; Frnkeni 1. Introduction Sltcedrs (Tmrix spp., Tmricles: Tmriccee) re deciduous shrus or smll trees of riprin res in deserts nd steppes of Eursi nd Afric (Bum, 1978). Ten species of Tmrix, including nine diverent sltcedrs nd the single evergreen species T. phyll (L.) Krsten (thel), were introduced into the United Sttes nd Mexico Mention of trde nmes or commercil products in this rticle is solely for the purpose of providing speciwc informtion nd does not imply recommendtion or endorsement y the US Deprtment of Agriculture. * Corresponding uthor. Present ddress: United Sttes Deprtment of Agriculture, Agriculturl Reserch Service, US Plnt, Soil nd Nutrition Lortory, Tower Rod, Ithc, NY 14853, USA. Fx: E-mil ddress: lrm32@cornell.edu (L.R. Milrth). eginning in the erly 1800s, primrily s ornmentls, for windreks nd shde, nd to stilize strem nks (Bum, 1967; Crins, 1989; DiTomso, 1998). Following the lte 1920s, some of the sltcedr species ecme highly invsive long western riprin res nd lkeshores, with n erly estimte of 600,000 h of ottomlnds infested (Roinson, 1965). The primry species involved in this invsion re Tmrix rmosissim Ledeour nd T. chinensis Loureiro, s well s common nd widespred hyrid tht hs formed etween these two species (Gskin nd Schl, 2002). Additionl invsive tx include T. prvixor de Cndolle, T. cnriensis Willdenow, T. gllic L. (the ltter two species eing diycult to distinguish), nd hyrids involving comintions of T. rmosissim nd T. chinensis with T. prvixor nd T. cnriensis/t. gllic (Gskin nd Schl, 2002, 2003). Sltcedr infesttions currently rnge /$ - see front mtter. Pulished y Elsevier Inc. doi: /j.iocontrol

3 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) from North Dkot to Wshington nd south to northern Mexico. Both nthropogenic chnges in western riprin ecosystems, which creted idel conditions for sltcedr invsion of distured res, s well s the ility of sltcedr to invde nd modify undistured environments, hs creted wht is considered to e n ecologicl disster for riprin res of the West (DeLoch et l., 2000). The Diorhd lef eetle from Fukng, Chin nd Chilik, Kzkhstn, designted s Diorhd elongt (Brullé) deserticol Chen y DeLoch et l. (2003) nd y Lewis et l. (2003,), ws relesed into the open Weld in 2001 in the United Sttes for the clssicl iologicl control of sltcedr. Both dults nd lrve defolite sltcedr; 3rd instrs re the most dmging stge (Lewis et l., 2003). These eetle popultions re estlished nd incresing in undnce t most sites north of 37 N ltitude (DeLoch et l., 2004). However, no estlishment occurred t more southern sites where summer dy lengths re less thn 14 h 30 min, which is the criticl photoperiod for the Fukng popultion (D.W. Ben, personl communiction). In the southern res, dult eetles enter reproductive dipuse in erly summer nd presumly deplete their ft ody reserves nd strve efore the following spring (Lewis et l., 2003). This left lrge res of some of the most serious sltcedr infesttions, from Texs nd Oklhom west to southern Cliforni, without iologicl control gent. Other, more southern, popultions of D. elongt sensu lto hve shorter criticl photoperiods for dipuse induction (DeLoch et l., 2004; D.W. Ben, personl communiction) nd therefore re more likely to estlish t more southern ltitudes in North Americ. However, vrition in this nd other iologicl trits mong popultions of D. elongt, especilly those distnt from the originl source of eetles relesed, my e ccompnied y vrition in their host rnge s well. A primry concern in the sltcedr iologicl control progrm hs een ssessing the risk posed to ntive, non-trget plnts in the genus Frnkeni L., smll shrus of desert nd slt mrsh hitts (Lewis et l., 2003). In North Americ, six species occur in the southwestern United Sttes nd northern Mexico (Whlen, 1980, 1987). The genus Frnkeni is plced in the Frnkenicee nd together with the Old World fmily Tmriccee comprises the order Tmricles (Spichiger nd Svolinen, 1997). Frnkeni spp. re the only ntive memers of the Tmricles found in North Americ (or even the Western Hemisphere); Tmrix is the only exotic genus of the order present (Whlen, 1980, 1987). A secondry concern involves the introduced thel, which is ntive to prts of southern Asi nd northern nd estern Afric (Bum, 1978). It is cold-intolernt, tree-sized, evergreen species of Tmrix tht is grown s drought-tolernt shde tree nd windrek, especilly in northern Mexico. We report here our evlution of the host speciwcity of four popultions of D. elongt collected cross wide geogrphicl re of Eursi nd North Afric nd elow 43 N ltitude, three of which hd never een tested previously. This informtion, in comintion with other iologicl studies of cndidte popultions, will provide the sis for determining the most promising popultion of D. elongt to relese in the southern res of the sltcedr infesttion in North Americ. 2. Mterils nd methods 2.1. Insect colonies The Diorhd eetles collected on Tmrix in Asi nd the Mediterrnen re were ll identiwed s D. elongt y A.S. Konstntinov (USDA-Agriculturl Reserch Service Systemtic Entomology Lortory, Beltsville, MD) nd/ or I.K. Loptin (Byelorussin University, Minsk, Belrus), lthough vrious nmes hve een proposed in the literture (see DeLoch et l., 2003). Ongoing reserch y our tem (J.L. Trcy, ARS, Temple, TX; D.J. Kzmer, J.F. Gskin, ARS, Sidney, MT; D.W. Ben, J.C. Herr, ARS, Alny, CA; A.A. Cossé, R.J. Brtelt, ARS, Peori, IL; nd D.C. Thompson, New Mexico Stte University, Ls Cruces, NM) indictes the proility of four species involved in our studies. The four popultions of D. elongt included in this study originted from North Afric to western Chin. They were collected 15 km south of Sfx, Tunisi (ltitude N, longitude E, elevtion 10 m); 3 km west of Sfkki, Crete, Greece (ltitude N, longitude 24.6 E, elevtion 7 m); 7 km west of Krshi (Qrshi), Uzekistn (ltitude N, longitude E, elevtion 350 m); nd t the Turpn Eremophyte Botnic Grden of Acdemi Sinic, c. 10 km southest of Turpn, Xinjing Province, Chin (ltitude N, longitude E, elevtion 70 m elow se level). We here refer to these s D. elongt from Tunisi, Crete, Uzekistn, nd Turpn. Although the Wrst host speciwcity tests in involved D. elongt from or ner Turpn, survivl on ll test plnts in the initil lrvl no-choice test ws very low nd no oviposition occurred in the dult test tht included ntive Frnkeni plnts (DeLoch et l., 2003). Therefore, in the present study we evluted this popultion more rigorously. In ddition, severl tests involving D. elongt from Crete included comprison with the previously relesed popultion from Fukng, Chin. Voucher specimens of D. elongt from ll loctions were deposited with the Ntionl Collection of Insects nd Mites of the Ntionl Museum of Nturl History, Smithsonin Institution, Wshington, DC (under Lot Numers GSWRL nd ). All eetles from overses (except from Fukng) were rought into the USDA-ARS, Exotic nd Invsive Weed Reserch Unit qurntine fcility t Alny, Cliforni where prsites, predtors, nd pthogens were eliminted. Beetles (eggs nd/or dults) were susequently sent to the USDA-ARS Arthropod Continment Fcility (qurntine) t Temple, Texs to initite our own colonies or for immedite use in some tests. We otined Fukng eetles from Weld colonies ner Lovell, Wyoming in June 2002 from our coopertor D.J. Kzmer (USDA-ARS, Sidney, Montn).

4 34 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) The Fukng eetles hd originlly een clered through the Temple qurntine fcility. Beetle colonies t Temple were mintined either in the qurntine lortory on potted Tmrix spp. (photoperiod of 16:8 h [L:D] nd 28 C [rnge C]) or in outdoor Weld cges on plnted Tmrix spp Test plnts The test plnts selected for host rnge testing of D. elongt were sed on the revised list of DeLoch et l. (2003), which ws developed ccording to the centrifugl phylogenetic method (Wpshere, 1974) using the ngiosperm phylogeny of Spichiger nd Svolinen (1997). Plnts included US ccessions of the sltcedrs T. rmosissim, T. chinensis, T. cnriensis/t. gllic, nd T. prvixor, vrious sltcedr hyrids, thel (T. phyll), three species of Frnkeni (ll in the order Tmricles), nd nine other relted plnts in the suclss Cryophyllide (see Tle 1 for nmes). Most Tmrix species nd hyrids were identi- Wed y J.F. Gskin (USDA-ARS, Sidney, MT) using the fourth intron of the nucler phosphoenolpyruvte croxylse (pepc) gene (Gskin nd Schl, 2002). In ddition, four unrelted hitt ssocites of sltcedr nd 15 griculturl plnts were included in the tests to hve dt ville to nswer concerns of lndowners should they rise. As in the originl tests, Frnkeni spp. nd thel were considered criticl test plnts, i.e., species within the order Tmricles on which only low level of dmge ws cceptle (DeLoch et l., 2003; Lewis et l., 2003). Plnts were otined from seeds, trnsplnts or cuttings of plnts growing in the Weld, or locl nurseries. Some of the test plnts hd een plnted on the Temple fcility grounds. Seeds, trnsplnts or cuttings were plnted in 8-L pots contining mixture of 10:3:2:1 prts of vermiculite, potting soil, pet moss, nd snd. Cuttings nd trnsplnts were rooted under n utomtic misting mchine. Venus Xytrps (Dione muscipul Ellis) were plced in 4-L pots contining 1:1 mixture of pet moss nd snd nd were not fertilized. Agriculturl test plnts derived from seed were fertilized weekly with N-P-K solule fertilizer. All other plnts were fertilized twice yer with pellets of slow-relese fertilizer ( N-P-K). Plnts were held under nturl dylengths nd C in greenhouse or nturl tempertures in n outdoor slthouse prior to their use in tests Lrvl no-choice tests We used comintion of lortory tests utilizing excised folige nd Weld-cge tests with sleeve gs on potted test plnts to determine the survivl of D. elongt lrve overed single type of test plnt Vils, lortory, 2002 We conducted no-choice test of lrvl/pupl survivl involving D. elongt from Crete nd Fukng during June 2002 in the qurntine lortory t 28 C (rnge C) nd photoperiod of 16:8 h (L:D). The experimentl design for ech eetle popultion ws one-wy tretment structure in completely rndomized design. Three replictes per test plnt were used, which consisted of eight Tmrix species, ccessions nd hyrids, nd three Frnkeni species for the two popultions. An dditionl nine species of relted plnts nd four species of unrelted hitt ssocites of sltcedr were included for the Crete eetle (see Tle 1 for nmes). Unfed neonte lrve (0- to 24-h old) were plced in seprte 50 ml ventilted plstic vils. Ech lrv ws provided excised leves from single type of test plnt. Leves were replced every 2 3 dys s needed nd lrve or pupe were checked dily for survivl nd development. Between 10 nd 13 lrve of ech eetle popultion were provided ech test plnt species. Groups (replictes) of three to Wve lrve were used to clculte percentge survivl to the dult stge Agriculturl plnts, vils, lortory, 2003 Three series of lrvl no-choice tests were conducted in The Wrst test evluted the lrvl/pupl survivl nd development of D. elongt from Crete, Tunisi, Uzekistn, or Turpn on 15 species of griculturl crops of importnce to Texs nd other sttes. These were: rice Cocodrie (Oryz stiv L.), sorghum ATx2752*RTx430 (Sorghum icolor (L.) Moench), winter whet TAM III (Triticum estivum L.), corn CML325xTx732 (Ze mys L.), penut Tmrun 96 (Archis hypoge L.), pecn (Cry illinoinensis (Wngenh.) K. Koch), wtermelon Chrleston Gry (Citrullus lntus (Thun.) Mtsum. & Nki), ornge Vlenci (Citrus sinensis (L.) Oseck), grpefruit Rio Red (Citrus x prdisi Mcfd.), cntloupe Imperil 45 (Cucumis melo L.), soyen AG4701 (Glycine mx (L.) Merr.), cotton (Gossypium hirsutum L.), sunxower (Helinthus nnuus L.), lflf (Medicgo stiv L.), nd grpe Chmpnel (Vitis vinifer L.). Tmrix rmosissim (Lovelock, NV; GenBnk Accession No. AY090385) served s the control test plnt. The test ws done during June 2003 in the qurntine lortory t 28 C (rnge C) nd photoperiod of 16:8 h (L:D). The experimentl design for ech eetle popultion ws one-wy tretment structure in completely rndomized design. Four replictes were used per test plnt. Groups of four unfed neonte lrve (0- to 15-h old) from ech of the four eetle popultions were plced in seprte 50 ml ventilted plstic vils nd fed excised leves from one of the test plnt species. Leves were replced every 2 3 dys s needed nd the lrve or pupe were checked dily for survivl nd development. Sterilized snd ws plced in the ottom of ech vil when mture 3rd instrs (Wnl instr) were present to provide puption site. Surviving dults were counted upon emergence nd percentge survivl ws clculted for ech group of lrve Non-Tmricles plnts, vils, lortory, 2003 The second no-choice test ssessed the suitility of nine species of relted plnts (not including Frnkeni species)

5 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) Tle 1 Percentge survivl (neonte to dult) of two popultions of D. elongt on 11 Tmricles nd 13 other test plnts: no-choice tests in vils, Temple, TX, June 2002 Txonomic grouping % survivl (men SD) Crete, Greece Fukng, Chin Suclss Cryophyllide, Polygonlin Linege Order Tmricles Fmily Tmriccee Tmrix phyll c,d (L.) Krsten (thel) Uvlde, TX c T. cnriensis Willdenow/T. gllic L; Texs City, TX (AY090398, AY090437) cd T. chinensis Loureiro; Seymour, TX (AY090386) T. chinensis nd/or T. rmosissim T. chinensis; Artesi, NM (AY090386) nd/or (AY090385, c AY090386) T. prvixor de Cndolle; Ls Cruces, NM T. rmosissim Ledeour; Lovell, WY (AY090385, AY090396) c Puelo, CO (AY090385) T. rmosissim nd/or T. rmosissim T. chinensis; Bishop, CA (AY090385) nd/or (AY090385, AY090386) Fmily Frnkenicee Frnkeni jmesii Torrey; Puelo, CO e 0 0 d F. johnstonii Correll; Lredo, TX e 0 0 F. slin d,e (Molin) I.M. Johnston; Point Isel, CA d Order Nepenthles Fmily Drosercee Dione muscipul Ellis (Venus Xytrp) 0 0 e Order Plumginles Fmily Plumgincee Limonium limtum e Smll (ordered se lvender) 0 0 e Plumgo cpensis Thunderg (lue plumgo) 0 0 e Order Polygonles Fmily Polygoncee Rumex ltissimus e Wood (smooth dock) 0 0 e Order Simmondsiles Fmily Simmondsicee Simmondsi chinensis (Link) Schneider (jojo) 0 0 e Suclss Cryophyllide, Cryophylllin Linege Order Cryophyllles Fmily Amrnthcee Amrnthus litoides Wtson (prostrte pigweed) 0 0 e Fmily Chenopodicee AllenrolW occidentlis e (Wtson) Kuntze (pickleweed) 0 0 e Atriplex cnescens e (Pursh) Nutll (4-winged sltush) 0 0 e Fmily Portulccee Portulc olerce L. (purslne) 0 0 e Unrelted Hitt Assocites Fmily Fcee Prosopis glndulos e Torrey vr. glndulos (honey mesquite) 0 0 e Fmily Sliccee Populus fremontii e Wtson (Fremont cottonwood) 0 0 e Slix exigu e Nutll (coyote willow) 0 0 e Fmily Astercee Bcchris slicifoli e (R. & P.) Pers. (seepwillow cchris) 0 0 e n D 3 with 13 (Crete) or (Fukng) lrve smpled per test plnt. Vlues within ech column followed y the sme letter re not signiwcntly diverent (Kruskl Wllis Test on rnks with men rnk vlues seprted y Fisher s protected lest signiwcnt diverence test, P > 0.05). Txonomic groupings follow ngiosperm phylogeny of Spichiger nd Svolinen (1997), nd Tmrix follows the revision of Bum (1978) nd the moleculr identiwctions of test plnt mteril y J.F. Gskin (USDA-ARS, Sidney, MT). X denotes hyrid. Accession numers, if ville, represent sequences of hplotypes of the pepc gene tht re in the Ntionl Institute of Helth s GenBnk genetic sequence dtse. Accession numers re given for ech hplotype of heterozygous pepc genes, while single ccession numer is given for the two identicl hplotypes of homozygous pepc genes. c Not conwrmed y txonomic uthorities. d Criticl test plnts. e Hitt ssocites of Tmrix.

6 36 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) nd four unrelted hitt ssocites of sltcedr (ll outside the order Tmricles, see Tle 1 for nmes) for immture development nd survivl of D. elongt from Tunisi, Uzekistn or Turpn. The control tretment, experimentl design, nd methods used were the sme s descried in the previous section Tmricles, sleeve gs, outdoors, 2003 The third 2003 lrvl test utilized seven species nd hyrids of Tmrix nd three species of Frnkeni tht were fed to D. elongt lrve from Tunisi, Uzekistn or Turpn (see Tle 2 for nmes). Tests were conducted outdoors on the fenced grounds of the Temple ARS lortory in m (length width height, 18 m 3 ) Weld cges t men minimum nd mximum temperture of 22 nd 36 C, respectively, nd photoperiod of 16:8 h (L:D) (nturl light supplemented with hlogen Xoodlight ove ech cge). The re immeditely surrounding the Weld cges ws treted periodiclly with hydrmethylnon (tetrhydro-5,5-dimethyl-2(1h)-pyrimidinone[3-[4-(trixuoromethyl)phenyl]-1-[2-[4-(trixuoromethyl)phenyl]ethyenyl]-2-propenylidene]hydrzone, Amdro, Amrnds, Atlnt, Georgi) to control Wre nt (Solenopsis invict Buren) infesttions. A permit uthorizing testing in outdoor cges ws received from USDA Animl nd Plnt Helth Inspection Service in 1998 nd ws renewed in The experimentl design for ech eetle popultion ws one-wy tretment structure in completely rndomized design, with Wve replictes per test plnt. Groups of 10 neonte lrve (0- to 15-h old) from Turpn (July 2003), Tunisi or Uzekistn (August 2003) were plced in individul cm polyester orgnz sleeve gs tht were securely tied to the rnches of potted test plnts. Two or three gs were used per plnt. Plnts were plced inside the lrge Weld cges nd wtered with drip irrigtion system. Surviving dults were counted upon emergence nd percentge survivl ws clculted for ech group of eetles. Some replictes were omitted from nlysis due to nt predtion Adult tests We studied dult D. elongt host plnt selection nd oviposition preferences mong diverent Tmrix nd Frnkeni species nd hyrids using vrious pired- nd multiple-choice tests. All other test plnts, which did not support ny lrvl development, were not included in these host speciwcity tests. We conducted ll tests in outdoor cges on the fenced grounds of the Temple ARS lortory. The primry focus of these tests ws to investigte host cceptnce y D. elongt of the ntive Frnkeni species, in prticulr F. jmesii Torrey nd F. johnstonii Correll. F. slin (Molin) I.M. Johnston lso ws included in some of these tests ut ws more fully ssessed y the ARS lortory in Alny, CA (J.C. Herr, unpulished dt). A second focus involved the introduced thel. This plnt ws more fully tested in seprte study nd dditionl results will e reported elsewhere Pired-choice test (Tmrix:Frnkeni), smll cges, outdoors, 2003 This test focused solely on dult coloniztion of nd oviposition on the ntive species F. jmesii nd F. johnstonii when pired with sltcedr y the vrious popultions of D. elongt originting from Crete, Tunisi, Uzekistn, nd Turpn. Tests were conducted outdoors during July 2003 in cm (length width height, 0.3m 3 ) luminum screen cges plced inside 3 3 2m Weld cges, under 16:8 h (L:D) photoperiod (nturl light supplemented with hlogen Xoodlight ove ech Weld cge) nd men minimum nd mximum temperture of 22 nd 38 C, respectively. One potted sltcedr plnt (T. rmosissim T. chinensis, Puelo, CO) nd one potted Frnkeni species (F. jmesii or Tle 2 Percentge survivl (neonte to dult) of three popultions of D. elongt on vrious Tmrix nd Frnkeni plnts: no-choice tests in sleeve gs outdoors, Temple, TX, July nd August 2003 Test plnts % survivl [men SD (n)] Sfx, Tunisi Krshi, Uzekistn Turpn, Chin Tmrix prvixor Ls Cruces, NM (5) c (5) (5) T. chinensis nd/or T. rmosissim T. chinensis Artesi, NM (4) cd (5) (5) (AY090386) nd/or (AY090385, AY090386) T. rmosissim T. chinensis Puelo, CO (AY090385, AY090386) (5) cd (4) (5) T. cnriensis/t. gllic Glveston, TX (AY090398, AY090437) (5) (5) (5) T. rmosissim nd/or T. rmosissim T. chinensis Bishop, CA (5) (3) (5) (AY090385) nd/or (AY090385, AY090386) T. chinensis Pecos River/I-10, TX (AY090386) (5) c (5) (5) T. phyll c (thel) Encino, TX c (5) (4) (4) Frnkeni jmesii c Puelo, CO c (3) d (3) (5) F. johnstonii c Lredo, TX c (4) cd (5) (5) F. slin c Owens Vlley, CA c (3) cd (5) (5) Ten lrve were smpled per replicte of ech eetle popultion. Vlues within ech column followed y the sme letter re not signiwcntly diverent (Kruskl Wllis Test on rnks with men rnk vlues seprted y Fisher s protected lest signiwcnt diverence test, P >0.05). Moleculr identiwctions of Tmrix, including GenBnk ccession numers if ville. X denotes hyrid. c Criticl test plnts.

7 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) Tle 3 Presence of dult D. elongt from four diverent popultions 1, 2, nd 5 dys post-relese on Tmrix nd Frnkeni plnts: pired-choice (Tmrix:Frnkeni) tests in smll cges outdoors, Temple, TX, July 2003 No. dults per loction (men SE) P vlue c Loction of dults: Crete, Greece Sfx, Tunisi Krshi, Uzekistn Turpn, Chin Tmrix rmosissim T. chinensis d vs. Frnkeni jmesii 1 dy post-relese P D 0.06 T. rmosissim T. chinensis F. jmesii d Cge wlls c Ded/unccounted dys post-relese P D 0.52 T. rmosissim T. chinensis F. jmesii d Cge wlls c Ded/unccounted dys post-relese P <0.01 T. rmosissim T. chinensis F. jmesii e e e e Cge wlls de cd de cd Ded/unccounted cde cde de c Tmrix rmosissim T. chinensis vs. Frnkeni johnstonii 1 dy post-relese P <0.01 T. rmosissim T. chinensis cd F. johnstonii fg g g g Cge wlls defg efg efg de Ded/unccounted de def de c 2 dys post-relese P <0.01 T. rmosissim T. chinensis F. johnstonii d d d d Cge wlls cd cd d cd Ded/unccounted cd c cd dys post-relese P <0.01 T. rmosissim T. chinensis F. johnstonii cd d cd d Cge wlls cd c cd cd Ded/unccounted cd cd cd Tests in screen cges (ht) cm, ech cge with 20 eetles (10 mles, 10 femles) nd 2 plnts (1 Tmrix nd 1 Frnkeni); n D 3 (Crete) or 5 for ech test plnt pir nd eetle popultion. Within ech plnt piring nd dy post-relese, individul mens (when there ws signiwcnt Loction Beetle interction) followed y the sme letter within nd etween columns re not signiwcntly diverent. Otherwise, the verges of the mens for the four eetle popultions within loction (verges not listed) followed y the sme letter re not signiwcntly diverent (Fisher s protected lest signiwcnt diverence test, P > 0.05). c P vlue for Loction Beetle interction. In ll cses, the min evect of Beetle Popultion ws not signiwcnt (P D 1.0) wheres Loction ws highly signiwcnt (P < 0.01). d Moleculr identiwction; GenBnk Accession Nos. AY nd AY F. johnstonii) were rndomly rrnged in ech cge. Averge heights (including the 20 cm tll pot) of the plnts were 66 cm (sltcedr), 37 cm (F. jmesii), nd 42 cm (F. johnstonii). Pots rested on 3 cm lyer of potting soil tht sored excess wter within the cges to prevent the entrpment nd drowning of eetles. Plnts were wtered with drip irrigtion system (Tle 3). The experimentl design for ech of the two test plnt comintions ws two-wy fctoril tretment structure in completely rndomized design. Three (Crete only) or Wve replictions were used, with four diverent D. elongt popultions nd either four possile loctions for dults (sltcedr, Frnkeni plnt, cge wlls nd ded/unccounted) or three loctions for eggs (sltcedr, Frnkeni plnt, nd cge wlls). Beetles (10 mles nd 10 femles per cge) were relesed into the center of the cge. Adults were counted 1, 2, or 5 dys post-relese nd their loction ws recorded. Adults not esily oserved for the 1 nd 2 dy post-relese counts were scored s unccounted to void unnecessrily disturing the eetles. Ded dults were sexed nd replced if found. Eggs were collected, noting their loction, on the Wfth dy post-relese to llow for mximum oviposition to occur ut just prior to htching of the oldest eggs. All plnts were thoroughly wshed with

8 38 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) Tle 4 Presence of dult D. elongt from three diverent popultions 1, 2, nd 5 dys post-relese on Tmrix, Frnkeni, nd willow plnts: pired-choice (potentil host:non-host) tests in smll cges outdoors, Temple, TX, August 2003 No. dults per loction (men SE) Loction of dults: Sfx, Tunisi Krshi, Uzekistn Turpn, Chin P vlue c Frnkeni jmesii vs. Coyote willow 1 dy post-relese P D 0.20 F. jmesii c Coyote willow c Cge wlls Ded/unccounted dys post-relese P <0.01 F. jmesii d cd cd Coyote willow d d cd Cge wlls c Ded/unccounted dys post-relese P D 0.68 F. jmesii Coyote willow Cge wlls Ded/unccounted Frnkeni johnstonii vs. Coyote willow 1 dy post-relese P D 0.23 F. johnstonii c Coyote willow c Cge wlls Ded/unccounted dys post-relese P D 0.26 F. johnstonii c Coyote willow c Cge wlls Ded/unccounted dys post-relese P D 0.63 F. johnstonii Coyote willow c Cge wlls c Ded/unccounted Tmrix rmosissim T. chinensis d vs. Coyote willow 1 dy post-relese P D 0.33 T. rmosissim T. chinensis Coyote willow Cge wlls Ded/unccounted dys post-relese P D 0.85 T. rmosissim T. chinensis Coyote willow c Cge wlls c Ded/unccounted dys post-relese P D 0.09 T. rmosissim T. chinensis Coyote willow c Cge wlls c Ded/unccounted Coyote willow vs. Coyote willow 1 dy post-relese P D 0.30 Coyote willow # c Coyote willow # c Cge wlls Ded/unccounted

9 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) Tle 4 (continued) No. dults per loction (men SE) P vlue c Loction of dults: Sfx, Tunisi Krshi, Uzekistn Turpn, Chin 2 dys post-relese P D 0.15 Coyote willow # Coyote willow # Cge wlls Ded/unccounted dys post-relese P D 0.98 Coyote willow # Coyote willow # Cge wlls Ded/unccounted Tests in screen cges (ht) cm, ech cge with eetles (2 10 mles, 10 femles) nd 2 plnts (1 potentil host plnt nd 1 coyote willow [non-host]); n D 2 (Turpn) or 3 for ech test plnt pir nd eetle popultion. Within ech plnt piring nd dy post-relese, individul mens (when there ws signiwcnt Loction Beetle interction) followed y the sme letter within nd etween columns re not signiwcntly diverent. Otherwise, the verges of the mens for the three eetle popultions within loction (verges not listed) followed y the sme letter re not signiwcntly diverent (Fisher s protected lest signiwcnt diverence test, P > 0.05). c P vlue for Loction Beetle interction. In ll cses, the min evect of eetle popultion ws not signiwcnt (P > 0.65) wheres loction ws highly signiwcnt (P < 0.01). d Moleculr identiwction; GenBnk Accession Nos. AY nd AY wter nd their loctions rndomized etween replictes. New dults were used for ech replicte Pired-choice test (potentil host:non-host), smll cges, outdoors, 2003 This test, conducted in August 2003, ws similr to the previous one in design except tht dult D. elongt presence nd oviposition were ssessed on F. jmesii nd F. johnstonii in the sence of sltcedr ut with known non-host, coyote willow (Slix exigu Nutll) present. Men minimum nd mximum tempertures during the experiment were 22 nd 35 C, respectively. One potted Frnkeni species (F. jmesii or F. johnstonii), sltcedr plnt (positive control, T. rmosissim T. chinensis, Puelo, CO), or coyote willow (negtive control) were pired with potted coyote willow plnt in ech cge. The loction of ech pot within cge ws rndomly ssigned. Averge heights (including the pot) of the plnts were 66cm (sltcedr), 37cm (F. jmesii), 42cm (F. johnstonii), nd 65 cm (willow). The experimentl design for ech test plnt comintion ws two-wy fctoril tretment structure in completely rndomized design. Two (Turpn only) or three replictes were used, with three diverent eetle popultions (Tunisi, Uzekistn or Turpn) nd four possile loctions for dults (primry test plnt, coyote willow, cge wlls, nd ded/unccounted) or three loctions for eggs (primry test plnt, coyote willow, nd cge wlls). Groups of 10 mle nd 10 femle eetles were relesed into ech cge; few replictes of Uzekistn eetles contined fewer mles. Adults nd eggs were counted s previously descried, except tht ded dults were not replced (Tle 4) Multiple-choice tests, smll nd lrge cges, outdoors, Six multiple-choice tests of dult D. elongt host preference were conducted from 2002 to Tests A1 A5 (see Tles 5 7) involved comprisons etween D. elongt from Crete nd the previously relesed popultion from Fukng. These tests were conducted from July to August 2002 t men minimum nd mximum temperture of 23 nd 34 C, respectively. Test A6 (Tle 8) consisted of comprison etween D. elongt from Tunisi nd Uzekistn nd ws conducted in Septemer 2003 t men minimum nd mximum temperture of 18 nd 30 C, respectively. All tests were done in the 3 3 2m Weld cges under 16:8 h (L:D) photoperiod (nturl light supplemented with hlogen Xoodlight ove ech cge). This ws done primrily to prevent the induction of dipuse in the Fukng popultion. The experimentl design for ech test ws two-wy fctoril tretment structure in completely rndomized design, with two D. elongt popultions nd seven to nine possile loctions for dults or eggs (5 7 diverent test plnts, cge wlls, nd ground). We used 4 24 replictes depending on the prticulr test. Tests A1, A2, A5, nd A6 included vrious species or hyrids of Tmrix nd the Frnkeni species F. slin, F. johnstonii nd (except for A6) F. jmesii. Test A3 included one Tmrix hyrid, the three Frnkeni species nd two species of non-hosts. This test used Tmrix plnts tht were similr in height to the Frnkeni plnts. Test A4 ws similr to test A3 except tht the Tmrix plnt ws replced y third non-host plnt to ssess the eetles response in the sence of the trget weed (see Tles 5 8 for test plnt nmes). For tests A1 A4 nd A6, which were conducted in the lrge Weld cges, ech cge ws sudivided into four qudrts, ech qudrt contining one ech of the test plnts in pots. The plnts were rndomly rrnged in ech qudrt nd sunk into the ground to the top of ech pot. For ech eetle popultion, unsexed dults were relesed in

10 Tle 5 Presence of dults nd oviposition y D. elongt from two popultions on Tmrix nd Frnkeni plnts: multiple-choice tests in lrge outdoor cges, Temple, TX, July-August 2002 Loction (test plnt, cge wlls, Percent dults oserved or eggs lid (men SD) per loction (totl per test) nd ground) Test A1 (n D 8) Test A2 (n D 8) Adults Eggs Adults Eggs Crete, Greece Fukng, Chin Crete, Greece Fukng, Chin Crete, Greece Fukng, Chin Crete, Greece Fukng, Chin Tmrix rmosissim Puelo, CO c (184) (211) cd (702) (595) (142) (150) (530) (473) T. chinensis Seymour, TX (253) (195) (922) cd (281) T. chinensis nd/or T. rmosissim (146) (117) (388) (282) XT. chinensis Artesi, NM T. rmosissim nd/or (306) (202) (870) (377) T. rmosissim T. chinensis Bishop, CA T. cnriensis/t. gllic Texs (255) cd (122) c (1,258) d (399) City, TX T. prvixor Ls Cruces, NM (147) (118) (697) (392) T. phyll (thel) Phoenix, AZ de (106) e (98) ef (269) e (202) Frnkeni slin Bishop (Polet gh (3) gh (1) g (0) g (0) (1) (2) c (28) (5) cd Cnyon), CA F. johnstonii Lredo, TX gh (2) gh (2) g (0) g (0) (0) (0) c (0) (10) d F. jmesii Puelo, CO h (0) h (0) g (0) g (0) (2) (1) c (0) (0) d Cge wlls f (45) cd (194) g (13) f (159) (125) (203) (113) (111) c Ground/weeds gh (7) gh (1) g (0) g (0) (4) (4) c (16) (0) d Fctor P vlue P vlue P vlue P vlue Loction of dults/eggs <0.01 <0.01 Beetle popultion <0.01 < Loction Beetle <0.01 < Within ech test nd life stge, individul mens (when there ws signiwcnt Loction Beetle interction) followed y the sme letter within nd etween the Crete nd Fukng columns re not signiwcntly diverent. Otherwise, the verges of the mens for the two eetle popultions within loction (verges not listed) followed y the sme letter re not signiwcntly diverent (two-wy Kruskl Wllis Test on rnks with men rnk vlues seprted y Fisher s protected lest signiwcnt diverence test, P > 0.05). Ech replicte (representing 24 h period) is the verge of four qudrts in cge. Moleculr identiwctions of Tmrix. Accession numers re given in Tle L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) 32 48

11 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) Tle 6 Presence of dults nd oviposition y D. elongt from two popultions on Tmrix, Frnkeni nd non-host plnts: multiple-choice tests in lrge outdoor cges, Temple, TX, July August 2002 Loction (test plnt, cge wlls, nd ground) Percent dults oserved or eggs lid (men SD) per loction (totl per test) Test A3 (nd4) Test A4 (nd8 Crete or 5 Fukng) Adults Eggs Adults Eggs Crete, Greece Fukng, Chin Crete, Greece Fukng, Chin Crete, Greece Fukng, Chin Crete, Greece Fukng, Chin (58) (52) (139) (132) T. chinensis T. cnriensis/t. gllic Pecos River/I-10, TX Frnkeni slin Bishop (Polet Cnyon), CA (3) (1) c (0) (0) c (9) (2) c (85) (41) F. johnstonii Lredo, TX (0) (0) d (0) (0) c (1) (0) de (0) (0) c F. jmesii Puelo, CO (0) (0) d (0) (0) c (0) (0) e (0) (0) c Limonium limtum (0) (0) d (0) (0) c (0) (0) e (0) (0) c Plumgo cpensis (0) (0) d (0) (0) c (6) (0) cd (0) (0) c Slix gooddingii (5) (0) cde (0) (0) c Cge wlls (267) (242) (449) (155) (670) (430) (1,114) (391) Ground/weeds (3) (0) c (0) (0) c (23) (2) (0) (0) c Fctor P vlue P vlue P vlue P vlue Loction of dults/eggs <0.01 <0.01 <0.01 <0.01 Beetle popultion Loction Beetle Within ech test nd life stge, the verges of the mens for the two eetle popultions within loction (verges not listed) followed y the sme letter re not signiwcntly diverent (two-wy Kruskl Wllis test on rnks with men rnk vlues seprted y Fisher s protected lest signiwcnt diverence test, P>0.05). Ech replicte (representing 24h period) is the verge of four qudrts in cge. Moleculr identiwction; GenBnk Accession Nos. AY nd AY the center of cge. After pproximtely 24 h, or 2 4 dys for test A6 only, the numer of dults nd eggs on ech test plnt nd the cge wlls nd ground were recorded for ech qudrt. Eggs were removed nd dults were spirted ov the plnts. Adults, previously used nd new, for ech popultion were then relesed ck into the sme cge (tests A3 nd A6) or were rotted etween diverent tests tht were run simultneously (tests A1, A2, nd A4). The dt nlyzed were the verge of the four qudrts for ech oservtion dte. Test A5 (smll cge test) used three luminum screen cges, cm, plced in lrge Weld cge. Five potted test plnt species were rndomly rrnged in ech smll cge. Pots were set on 3 cm lyer of dried grss overlying 3 cm lyer of soil. Between 36 nd 40 dults were plced in the center of ech smll cge for pproximtely 24 h, fter which ll dults nd eggs were removed nd their loctions recorded. Adults, previously used in other dult tests, were relesed ck into the smll cges for dditionl replictes. For ll tests, due to the vrile numer of dults ville etween eetle popultions nd replictes nd the unknown proportion of femles, dult coloniztion nd oviposition dt were converted to the percent of dults or eggs, respectively, found t ech loction for given replicte. The verge height of plnts ove the soil line for tests A1 A5 ws 80 cm for Tmrix (20 cm for test A3), 20 cm for Frnkeni, 13 cm for Limonium, 60 cm for Plumgo, nd 130 cm for Slix. In test A6, the verge height of plnts ws 76 cm for Tmrix nd 27 cm for Frnkeni Sttisticl nlyses Dt on percentge lrvl/pupl survivl nd percentge of dults or eggs per loction (ech test plnt species, cge wll, nd ground) did not generlly conform to the ssumptions of prmetric tests nd were nlyzed using protected Kruskl Wllis test performed on the rnks of the percentge dt (PROC GLM, SAS Institute, 1999). Dt on the numer of dults or eggs per loction (ech test plnt, cge wlls, nd ded/unccounted) were sujected to nlysis of vrince (PROC MIXED, SAS Institute, 1999). Mens (numers or rnks) were seprted using Fisher s protected lest signiwcnt diverence test (LSMEANS, SAS Institute, 1999). 3. Results 3.1. Lrvl no-choice tests Vils, lortory, 2002 (Tle 1) Percentge survivl (neonte to dult) of lrvl D. elongt from Crete restricted to single test plnt ws generlly similr mong the vrious Tmrix species, ccessions nd hyrids, including etween the deciduous sltcedrs (except for T. prvixor) nd the evergreen thel (T. phyll). Survivl ws higher on Tmrix plnts thn on ll other test plnts,

12 42 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) Tle 7 Test A5, presence of dults nd oviposition y D. elongt from two popultions on Tmrix nd Frnkeni plnts: multiple-choice test in smll outdoor cges, Temple, TX, July August 2002 Loction (test plnt, cge wlls, nd ground) Percent (men SD) oserved per loction (totl per test) Adults Eggs Crete, Greece (n D 24) Fukng, Chin (n D 18) Crete, Greece (n D 24) Fukng, Chin (n D 18) Tmrix rmosissim Puelo, CO (381) (274) (1,392) (674) T. phyll (thel) Phoenix, AZ (369) (226) (1,921) (585) Frnkeni slin Bishop (Polet Cnyon), CA d (23) ef (6) c (212) d (15) F. johnstonii Lredo, TX ef (4) f (0) d (24) d (0) F. jmesii Puelo, CO f (0) ef (3) d (0) d (24) Cge wlls c (140) (142) (613) c (145) Ground/weeds e (11) ef (10) d (18) d (6) Fctor P vlue P vlue Loction of dults/eggs <0.01 <0.01 Beetle popultion Loction*Beetle For ech life stge, individul mens followed y the sme letter within nd etween the Crete nd Fukng columns re not signiwcntly diverent (Two-wy Kruskl Wllis Test on rnks with men rnk vlues seprted y Fisher s protected lest signiwcnt diverence test, P > 0.05). Ech replicte (representing 24 h period) is one cge. Moleculr identiwction; GenBnk Accession No. AY except etween Crete lrve fed T. cnriensis/t. gllic nd F. slin. Survivl lso ws higher on F. slin compred to F. jmesii, F. johnstonii nd the remining test plnts. All Crete lrve provided plnts other thn Tmrix or Frnkeni (order Tmricles) died s 1st instrs within 3 dys. No diverences were found in percentge survivl of D. elongt from Fukng mong the vrious Tmrix test plnts. Lrvl/pupl survivl on Tmrix ws higher thn on the three Frnkeni species, except for T. prvixor. Only one lrv of 10 survived to the dult stge on F. slin nd ll Fukng lrve died s 1st instrs on F. jmesii nd F. johnstonii. We performed seprte two-wy Kruskl Wllis Test on the rnked percentge dt tht included eetle popultion nd Tmricles plnts (Tmrix nd Frnkeni species only) s fctors. D. elongt from Crete hd higher verge survivl rte thn eetles from Fukng (62 nd 49%, respectively; P < 0.032). The pttern of survivl mong test plnts ws similr etween the two eetle popultions, with greter survivl on Tmrix plnts thn on Frnkeni plnts (P < 0.020, Fisher s Protected LSD test) Agriculturl plnts, vils, lortory, 2003 All lrve of D. elongt, regrdless of eetle popultion, died within 4 dys s 1st instrs when fed excised leves from the vrious species of griculturl plnts. Percentge survivl of lrve fed T. rmosissim ws moderte to high for ll eetle popultions in comprison [ % (Crete), % (Tunisi), % (Uzekistn), nd % (Turpn)] Non-Tmricles plnts, vils, lortory, 2003 No lrve survived more thn 3 dys, remining s 1st instrs, when provided leves from diverent txonomiclly relted (non-tmricles) plnts of sltcedr (i.e., not including the Frnkeni species) or unrelted hitt ssocites. As in the previous test, percentge survivl to the dult stge ws moderte to high for ll popultions of lrve fed T. rmosissim [ % (Tunisi), % (Uzekistn), nd % (Turpn)]. Although feeding dmge ws not exmined in this nd the previous tests, lrve of D. elongt provided non-host plnts generlly were oserved to wnder continuously in the vils until deth. In contrst, lrve provided sltcedr settled on the leves nd egn feeding within n hour of the strt of the experiment Tmricles, sleeve gs, outdoors, 2003 (Tle 2) The gretest diverences in survivl were oserved with D. elongt from Tunisi. Lrvl/pupl survivl for Tunisi eetles ws generlly similr mong the vrious Tmrix test plnts. Survivl rtes on Frnkeni plnts were much less thn those on the diverent sltcedrs ut were similr to tht on thel. In contrst, D. elongt from Uzekistn only displyed some diverences in survivl rtes mong the vrious Tmrix nd Frnkeni plnts. In prticulr, percentge survivl on thel ws greter thn on two of the sltcedr ccessions nd the three Frnkeni species. No diverences were found in percentge survivl rtes, which were moderte to high, for Turpn lrve fed vrious Tmrix nd Frnkeni plnts Adult tests Pired-choice test (Tmrix:Frnkeni), smll cges, outdoors, 2003 The presence of dult D. elongt on sltcedr (T. rmosissim T. chinensis) nd Frnkeni plnts ws generlly not diverent mong the eetles from Crete, Tunisi, Uzekistn nd Turpn for ech smpling dte. An unexplined higher mortlity of Turpn eetles in some replictes involving F. johnstonii led to generlly lower

13 L.R. Milrth, C.J. DeLoch / Biologicl Control 36 (2006) Tle 8 Test A6, presence of dults nd oviposition y D. elongt from two popultions on Tmrix nd Frnkeni plnts: multiple-choice test in lrge outdoor cges, Temple, TX, Septemer 2003 Percent (men SD) oserved per loction (totl per test) (n D 5) Adults Eggs Loction (test plnt, cge wlls, nd ground) Sfx, Tunisi Krshi, Uzekistn Sfx, Tunisi Krshi, Uzekistn Tmrix prvixor Ls Cruces, NM (107) (87) (1254) (961) T. chinensis T. cnriensis/t. gllic Big Spring, (106) (79) (1195) (1344) TX (AY090386, AY090437) T. rmosissim Slt Creek, CA (AY090385) (81) (93) (713) (1331) T. phyll (thel) Encino, TX (71) (64) cd (395) (1023) T. cnriensis/t. gllic Glveston, TX (55) (122) c (667) c (122) (AY090398, AY090437) Frnkeni slin Owens Vlley, CA (0) (4) d f (0) def (107) F. johnstonii Lredo, TX (0) (2) d f (0) f (0) Cge wlls (35) (42) c ef (10) cde (178) Ground (7) (1) d f (0) ef (71) Fctor P vlue P vlue Loction of dults/eggs <0.01 <0.01 Beetle popultion Loction Beetle 0.28 <0.01 For ech life stge, individul mens (eggs, signiwcnt Loction Beetle interction) followed y the sme letter within nd etween the Tunisi nd Uzekistn columns re not signiwcntly diverent. Otherwise, the verges of the mens for the two eetle popultions within loction (dults, verges not listed) followed y the sme letter re not signiwcntly diverent (two-wy Kruskl Wllis test on rnks with men rnk vlues seprted y Fisher s protected lest signiwcnt diverence test, P > 0.05). Ech replicte (representing 2 4 dy period) is the verge of four qudrts in cge. Moleculr identiwctions of Tmrix with GenBnk ccession numers, if ville. X denotes hyrid. numers of Turpn dults on sltcedr plnts compred to the other eetle popultions. The numer of dult eetles, when given choice etween sltcedr nd one of two Frnkeni species, ws higher on the sltcedr plnt 1, 2, nd 5 dys post-relese thn on the Frnkeni plnts or cge wlls, with only one exception on the Wrst dy (Turpn eetles on cge wlls with F. johnstonii present) (Tle 3). No to very few dults were counted on the Frnkeni plnts t ny time, which ws not diverent or less thn the numer of dults on the cge wlls (Tle 3). Oviposition y ll popultions of D. elongt ws much greter on the sltcedr plnts, verging eggs per plnt, thn on either Frnkeni species, on which zero to mximum verge of Wve eggs were lid per plnt (Fig. 1). Also, oviposition on the Frnkeni plnts ws not diverent thn on the surrounding cge wlls (Fig. 1). A three-wy fctoril nlysis, with eetle popultion, Frnkeni species nd loction of eggs s fctors, reveled signiwcnt Frnkeni loction interction (P < 0.009). This ws due to greter oviposition y D. elongt on sltcedr plnts pired with F. jmesii thn on sltcedr pired with F. johnstonii. Therefore, ll eetle popultions responded similrly in this test regrding the cceptility of the two ntive Frnkeni plnts for oviposition Pired-choice test (potentil host:non-host), smll cges, outdoors, 2003 The presence of dult D. elongt on the diverent plnts within ech of the four test plnt comintions ws similr (one exception) mong the three eetle popultions we tested. The numer of dult eetles ws gretest on the sltcedr plnt (T. rmosissim T. chinensis, positive control cges) thn elsewhere in the cge eginning one dy fter relese of the dults nd continuing until the end of the experiment (Tle 4). Survivl of dults ws high in the presence of sltcedr (85 92%). In contrst, in the remining cges which contined only Frnkeni (potentil host) nd willow (non-host) plnts, more dults were found on the cge wlls 24 h following relese thn on the Frnkeni or willow plnts (Tle 4). By the 2nd dy post-relese, the numers shifted towrd more ded or unccounted dults with continued low numers or no dults on the plnts. After 5 dys, most dult eetles were ded or were not recovered nd presumly hd died (57 93% of dults in Frnkeni cges; 92 98% in willow only cge). The numer of dults on the Frnkeni plnts ws less thn the numer tht hd died ut similr to the numer still live on the cge wlls (Tle 4). Some feeding y dult D. elongt ws noted on the Frnkeni plnts. Oviposition y ll popultions of D. elongt, in the sence of sltcedr, ws low on either Frnkeni species (verging 0 50 eggs) nd not diverent thn tht on the non-host coyote willow or on the cge wlls (Fig. 2). In contrst, on the sltcedr plnts (positive control), the dults lid lrge numers of eggs, verging eggs per plnt (Fig. 2). A seprte comprison of oviposition mong sltcedr, the two Frnkeni species nd coyote willow only for ech eetle type showed tht, regrdless of the originl source of eetles, sltcedr received more eggs thn ll other test plnts, wheres the numer of eggs lid on the Frnkeni or willow plnts ws not diverent (Fig. 2). Additionlly, oviposition on the two Frnkeni species did not diver mong eetle popultions (P > 0.51, two-wy fctoril nlysis with eetle popultion nd egg loction s fctors).

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