Toxicity toSpodoptera exigua andTrichoplusia niofIndividual P1 Protoxins andSporulated Cultures ofBacillus thuringiensis subsp. kurstaki HD-1andNRD-12

Thetoxicities toneonate Spodoptera exigua andTrichoplusia nioflyophilized powders obtained from sporulated liquid cultures (referred toassporulated cultures) andEscherichia coli-expressed P1[crylA(a) crylA(b) crylA(c)] protoxins fromthree-gene strains ofNRD-12andHD-1ofBacillus thuringiensis subsp. kurstaki weredetermined byusing diet incorporation bioassays. Although sporulated cultures frombothstrains weremoretoxic toT.nithanS.exigua, there werenodifferences intoxicity between NRD-12andHD-1. Toxicities ofthethree individual P1protoxins against S.exigua varied byatleast fivefold, withthecrylA(b) protein being themosttoxic. Thesesameprotoxins varied intoxicity against T.nibyatleast 16-fold, withthe cryIA(c) protein being themosttoxic. However, whentested against either S.exigua orT.ni,there wereno differences intoxicity between anNRD-12P1protoxin andthecorresponding HD-1P1protoxin. Comparing thetoxicities ofindividual protoxins withthatofsporulated cultures demonstrates thatnoindividual protoxin wasastoxic toS.exigua asthesporulated cultures. However, this samecomparison against T.nishowsthat boththecrylA(b) andcryIA(c) proteins areatleast astoxic asthesporulated cultures. Results fromthis study suggest thatNRD-12isnotmoretoxic toS.exigua thanHD-1,thatdifferent protein types havevariable host activity, andthatotherB.thuringiensis components arenotrequired forT.nitoxicity butthatother components suchasspores mightberequired forS.exigua toxicity. Although theinsecticidal bacterium Bacillus thuringiensis subsp. kurstaki (serotype H 3a:3b) HD-1caneffectively control someagriculturally important lepidopterous pests, suchasthecabbage looper (Trichoplusia ni[Hubner]), many keylepidopterous pests, suchasthebeetarmyworm (Spodoptera exigua[Hbfiner]), arerelatively tolerant and causeeconomic damage(14). Commercial formulations containing arelatively newstrain ofthis B.thuringiensis subspecies, NRD-12,wereshowntobethree tofour times more toxic toS.exigua thancommercial formulations ofHD-1 (13). Mostoftheinsecticidal activity against lepidoptera in B.thuringiensis subsp. kurstaki HD-1andNRD-12isattributedtotheP1crystal, whichiscomposed ofthree proteins belonging tothecryIA(a), cryIA(b), andcryIA(c) protein types(7,8,15). Moaretal.(15)reported thattheNRD-12 lyophilized powderobtained fromsporulated liquid cultures (referred tohereassporulated cultures) containing both proteinaceous crystals andspores was2.5-fold moretoxic against S.exigua thanwasHD-1.Mostofthis increased activity wasfoundtoreside intheP1crystal. However, subsequent studies revealed thattheHD-1strain usedin those tests wasmissing thecrylA(b) gene,whichhasbeen showntoberequired formaximumtoxicity toS.exigua (15, 19). Although these results demonstrated thatthecryIA(b) protein confers someinsecticidal activity, otherproteins alsomustbepartly responsible. Additionally, itisstill unclear whether NRD-12isinfactmoretoxic toS.exigua thanastrain ofHD-1containing allthree P1protoxin genes and,ifso,whatfactor isresponsible forthis activity. Recently, thethreeP1protoxin proteins ofHD-1and NRD-12havebeencloned andexpressed inEscherichia coli (L.Masson, G.Prefontaine, L.Peloquin, P.C.K.Lau,and