J. Genet. & Breed. 56: 165-169 (2002) Southwestern corn borer damage and aflatoxin accumulation in a diallel cross of maize W.P. Williams, F.M. Davis, -G.L. Windham and P.M. Buckley USDA-ARS Corn Horst Plant Resistance Research Unit, Box 9555, • Mississippi State, MS 39762, USA. Fax: (662) 325-8441. • Received September 18, 2001 ABSTRACT Southwestern corn borer, Diatraea grandiosella Dyar, is a serious pest of maize, Zea mays L., in the southern USA. When plants are infested during and after anthesis, larvae feed on the husks and developing cars before tunneling into the stalk. Larval feeding also provides potential sites for fungi to enter developing ears. Aflatoxin, produced by the fungus Aspergillus flavus Link: Fr, is a potent carcinogen, and its presence at levels exceeding 20 ng g restricts maize from interstate commerce. Aflatoxin contamination is a chronic problem in maize produced in the southern USA. Little is cur- rently known about the value of resistance to southwestern corn borer in reducing aflatoxin accu- mulation. This investigation was undertaken to compare aflatoxin accumulation in crosses among in- bred crosses with different levels of southwestern corn borer resistance and to study the importance of general and specific combining ability in the inheritance of resistance to southwestern corn bor- er and aflatoxin accumulation in an eight-parent diallel cross. Our results indicated that general com- bining ability was a highly significant source of variation in the inheritance of resistance to stalk tun- neling and ear damage by southwestern corn borer and resistance to aflatoxin accumulation. Stalk tunneling, ear damage, and aflatoxin accumulation were lowest in hybrids with the inbred line MP496 as a parent. Key words: Aspergillus flavus, Diatraea grandiosella, Zea mays. Abbreviations: AF, Aspergillus flavus; GCA, general combining ability, SCA, specific combining ability, SWCB, southwestern corn borer. INTRODUCTION Drought and high temperatures have fre- quently been linked with high levels of aflatox- Aflatoxin, a naturally occurring toxin pro- in accumulation in corn (DIENER, 1989; PAYNE, düced by the fungus Aspergillus flavus Link: Fr., 1992). Insect damage to developing maize ears is a potent carcinogen (CASTEGNARO and Mc- and kernels has also been associated with high GREGOR; PITrET, 1998). Aflatoxin contamination aflatoxin levels (GUTHRIE et al., 1981; MCMILLIAN, of maize, Zea mays L., is a sporadic problem in 1983; MCMILLIAN et al., 1985; WINDHAM et al., the Midwest, but a frequent problem in the 1999). Increased aflatoxin contamination of har- Southeast (PAYNE, 1992; WIDSTROM, 1996). Afla- vested grain has been attributed to several in- toxin B 1 is the most commonly found form of sects including corn earworm, Helicoverpa zea aflatoxin in maize. The U.S. Food and Drug Ad- (Boddie); European corn borer, Ostrinia nubi- ministration has a tolerance of 20 ng g for B1 lalis (Hubner), fall armyworm, Spodoptera in maize grain; grain with higher levels is re- frugiperda Q.E. Smith); and southwestern corn stricted from interstate commerce (GoUltiviA and borer, Diatraea grandiosella Dyar. BULLERMAN, 1995). Many countries have set even Both insect damage and aflatoxin accumula- lower tolerances for aflatoxin in maize. tion vary widely from year to year, and the re- \," Corresponding author. E-mail: [email protected] Joint contribution of USDA-ARS and the Mississippi Agricultural and Forestry Experiment Station, Mississippi Agric. And Forestry Exp. Stn. Journal No. J-9759. Purchase BN of Agriculture Use Only 166 lationship between the two is difficult to define. morillonitic, nonacid, thermic vertic Haplaquept) soil. WINDHAM et al. (1999) reported that aflatoxin lev- The single-row plots were approximately Sm long, els were significantly higher in when com- spaced I m apart, and thinned to 20 plants. 1995 In 1998, 7 d after 50% of the plants in a plot had mercial hybrids were both infested with south- silked, plants were infested with 60 southwestern corn western corn borer larvae and inoculated with borer larvae. Fifteen neonates were placed in the leaf A. flavus spores than when only inoculated with axil above the top ear and 45 in the axil of the top- A. flavus spores. In 1996, however, aflatoxin con- ear leaf (DAvis and WILLIAMS, 1994) using a portable plastic dispensing device (MIHM, 1983). On the fol- tamination was generally lower and was not in- lowing day, the top ear of each plant was inoculated creased significantly by infesting developing ears with a 3.4-ml suspension containing 3 x 108 A. flavus with southwestern corn borer larvae. Substantial conidia in distilled water using a treemarking gun effort has been invested in developing technique (ZUMMO and Scorr, 1989). Inoculum was prepared as for identifying maize germpiasm with resistance described by WINDHAM and WILLIAMS (1999) with A. flavus isolate NRRL 3357, which is known to produce to southwestern corn borer by USDA-ARS sci- aflatoxin (Scorr and ZUMMO, 1988). The experiment, entists at Mississippi State (DAvIs et al., 1989; was repeated in 1999. In 1999, three additional repli-, DAvis and WILLIAMS, 1997). Several germplasm cations were grown and inoculated with A. flavus. , but lines with resistance to leaf feeding by south- not artificially infested with southwestern corn borer. western corn borer when plants are in the whorl No insecticides were applied to prevent natural insect infestation. stage of growth have been developed and re- Mature ears in all experiments were hand har- leased (WILLIAMS and DAVTS, 1997). Although vested approximatel y 56 d after inoculation with A. germplasm with resistance to southwestern corn flavus and dried for 7 d at 38° C. In 1999, ears were borer after anthesis has not yet been released, visually rated for feeding damage by southwestern inbred lines Mp305, Mp496, and SC213 have cx- corn borer and other Lepidoptera before shelling. Ten ears from each plot were scored using the following hibitedreduced damage when infested 7 to 21d scale: (1) no Lepidoptera damage to any ears, (2) ear after anthesis (unpublished data). tip damage only to three or fewer ears, (3) ear tip This research investigates the association be- damage to four to six ears and no damage below tips, tween resistance to southwestern corn borer (4) ear tip damage to most ears and light additional damage and aflatoxin accumulation in maize. damage (2 or 3 kernels at I or 2 sites destroyed) to three or fewer ears, (5) ear tip damage to most ears Specific objectives were to (i) evaluate, in hy- and light damage to kernels below tips of four to six brids, inbred lines of corn with different levels cars, (6) ear tip damage and light damage below tips of resistance to southwestern corn borer dam- of most ears or moderate damage (4 to 6 kernels de- age and aflatOxin accumulation, (ii) obtain in- stroyèd at I to 3 sites) below tips of three or fewer formation on the relative importance of general ears, (7) ear tip damage and moderate to heavy dam- age (kernels destroyed at more than four sites or more and specific combining ability in the inheritance than six kernels destroyed at fewer sites) to four to of resistance to southwestern corn borer dam- six ears, (8) car tip damage and moderate to heavy1 age and aflatoxin accumulation, and (iii) com- damage below tips of seven or eight ears, and (9) ear pare ear damage and aflatoxin accumulation in tip damage and moderate to heavy damage below tips a diallel cross when infested and not infested of nine or ten ears. Ears from each plot were shelled, and the grain c with southwestern corn borer. was mixed by pouring through a sample-splitter twice. Grain was ground using a Romer mill (Union, MO). Afiatoxin contamination in 50-g subsamples from MATERIALS AND METHODS each plot was determined by the Vicam Aflatest (Wa- tertown, MA). This procedure detects aflatoxin (B1, Eight inbred lines of maize, Ab24E, GA209, 132 , G 1 , G2) at concentrations as low as 2 ng g. GE333, Mp305, Mp488, Mp496, SC213, and T202, In both 1998 and 1999, stalks in plots that had - were selected as parents of a diallel cross. Previous been infested with southwestern corn borer were dis- evaluations indicated that among a group of inbred sected after harvest. Tunneling from the top ear node lines infested with southwestern corn borer larvae af- to the base of the plant was measured. ter anthesis, Mp305, Mp496, and SC213 sustained less Plot means for all traits were calculated and used damage than other lines evaluated; GE333, Mp488, in an analysis of variance. To equalize variances and and, Ab24E were heavily damaged (unpublished da- because some values were zero, the data for aflatox- ta). The 28 single cross hybrids were planted in three in contamination were transformed by adding I and replications on 4 May 1998 and six replications on 3 taking the logarithm of each number [log (y+l)] pri- May 1999 in a randomized complete block design at or to analysis. Data on tunneling and aflatoxin for the Starkville, MS in a Leeper silty clay loam (fine mont- two years were combined for analyses over years. A. 167 combined analysis of data on ear damage and afla- TABLE 2 toxin accumulation with and without southwestern Estimates of general combining ability effects for corn borer infestation in 1999 was performed..Varia- stalk tunneling and aflatoxin accumulation following tion among hybrids was partitioned into"general infestation with sotuhwestern corn borer and (GCA) and specific (SCA) combining ability using inoculation with A. flavus in 1998 and 1999 GRIFFINGS Method 4, Model 1 (1956). Hybrids, years, and treatments (with and without southwest- Inbred Tunneling Aflatoxin ern corn borer infestation) were considered as fixed cm Log(ng g- + 1) effects. Mp305 4.56 -0.07 Ah24E 3.92 0.18 T202 2.14 0.32 RESULTS AND DISCUSSION Mp488 1.41 -0.19 GE333 0.55 0.00 Stalk tunneling following southwestern corn GA209 -1.20 0.38 borer infestation was twice as extensive in 1999 SC2 13 _4.68 -0.01 - as in 1998.