Biology of Agrotis malefida Guene (Lepidoptera: Noctuidae: Noctuinae): a true diapausing diapausant cutworm?

Alexandre Specht1,2,*, Andrs O. Angulo3, Tania S. Olivares3, Edegar Fronza1, Eduardo Valduga1, Francine Albrecht2, Graziela Poletto1, Neiva Monteiro de Barros2.

- Laboratrio de Biologia, Universidade de Caxias do Sul, Campus da Regio dos Vinhedos.

Alameda Joo Dall Sasso, 800, Cx. Postal 32. Bairro Universitrio, Bento Gonalves, RS. CEP 95700-000.
2

- Instituto de Biotecnologia, Universidade de Caxias do Sul. Universidad de Concepcin, Chile.

Key-words: Biology; Damage; Development; Larvae; Underground habits;

Biology of Agrotis malefida (Lepidoptera: Noctuidae: Noctuinae): a true diapausant cutworm?

Biological Sciences

*Alexandre Specht- Author for correspondence. spechta@terra.com.br.

Abstract

This study aimed to estimate the biotic potential of Agrotis malefida Guene, 1852 (Lepidoptera: Noctuidae: Noctuinae) under laboratory conditions. The insects were reared in a controlled environment (25 1 C, 70 10% RH and photophase of 14 hours) and observed daily. The larvae were fed Greenes artificial diet and adults were given an aqueous solution of sucrose 10%. The viability and duration of immature stages were assessed. The experiment started from 2,410 eggs. Larvae were isolated shortly after hatching. Longevity, pre-, post- and oviposition, fecundity and fertility of 13 adult 13 couples were also evaluated. The viability of eggs, larvae, pupae and prepupae was 96.72, 91.25, 78.37 and 95.26%, respectively. The average duration of eggs, larvae, prepupae, pupae and adults was 7.93, 54.26, 61.61, 37.43 and 12.85 days, respectively. The immature stage of A. malefida lasted an average of 161.289 days, ranging from 102 to 227 days. The life cycle of A. malefida is much longer than that of other congeners. The mean fecundity was 1,696.769 eggs and fertility 1,641.149 larvae per female. In conditions where the study was conducted, the biotic potential of A. malefida was of 606,666.593 individuals / female / year. The results also indicated that this species goes through larval and pupal diapause (pre-pupae).

Este estudo objetivou estimar o potencial bitico de Agrotis malefida Guene, 1852 (Lepidoptera: Noctuidae: Noctuinae) em condies laboratoriais. Os insetos foram criados em sala climatizada (25 1C, 70 10% UR e fotofase de 14 horas) com observaes dirias. As larvas foram alimentadas com dieta artificial de Greene e os adultos com soluo aquosa de sacarose a 10%. Nas fases imaturas avaliaram-se a viabilidade e a durao, a partir de 2,410 ovos, cujas larvas foram individualizadas logo aps a ecloso. Na fase adulta, avaliou-se a longevidade, perodos de pr, ps e oviposio, fecundidade e fertilidade de 13 casais. A viabilidade das fases de ovo, larva, pr-pupa e pupa foi de 96.72, 91.25, 78.37 e 95.26%, respectivamente. A durao mdia das fases

3 de ovo, larva, pr-pupa, pupa e adulto foi de 7.93, 54.26, 61.61, 37.43 e 12.85 dias, respectivamente. O perodo de desenvolvimento dos imaturos de A. malefida teve durao mdia de 161.289 dias, variando de 102 a 227 dias. O ciclo de vida de A. malefida bem mais longo do que de outras espcies do gnero. A fecundidade mdia foi de 1,696.769 ovos e a fertilidade 1,641.149 larvas por fmea. Nas condies em que foi realizado este estudo o potencial bitico de A. malefida foi de 606.666,593 indivduos/fmea/ano. Os resultados tambm indicaram que esta espcie apresenta diapausa larval (pr-pupal) e pupal.

INTRODUCTION Noctuidae, the largest family of macro-moths, includes several agriculturally important species. Most noctuinae larvae, collectively known as cutworms, have underground habits. They are voracious and are able to feed on a variety of hosts (Kitching & Rawlins 1998). Agrotis Ochsenheimer, 1816 (Noctuinae), for instance, is a cosmopolitan genus with 317 species (Poole 1989). All Agrotis larvae have underground habits, coming to the surface at night to feed. While feeding, they cut the base of their food plants, causing severe damage and rapid crop loss. When disturbed, Agrotis larvae roll up in a very characteristic behavior (ex. Baudino 2004; Angulo et al. 2008).

4 Agrotis malefida Guene, 1852 has been recorded from most of the Americas (Lafontaine 2004) and is likely to occur worldwide (Angulo and Quezada 1975). The species has been widely documented in the United States and Mexico (Lafontaine 2004), Chile (Angulo and Quezada 1975, Angulo et al. 2008), Argentina (Igarzbal et al. 1994; Rizzo et al. 1995; Baudino 2004), Brazil (ex. Silva et al. 1968, Specht and Corseuil 1996; 2002, Specht et al. 2004) and United States of America (Lafontaine 2004; Pogue 2006). Larvae of A. malefida are known in North America as Pale-Sided Cutworms; in Latin America, as gusano spero, oruga spera or oruga cortadora spera (Lafontaine 2004); and in Brazil, as lagarta rosca (Silva et al. 1968). Larvae of A. malefida are polyphagous, attacking various herbaceous, native and cultivated plants, including economically relevant crops such as alfalfa, bean, cabbage, clover, corn, onion, pea, pepper, potato, Sorghum, soybean, sunflower, tobacco and tomato (e.g. Silva et al. 1968, Villata et al. 1988, Rizzo et al. 1995, Pastrana 2004, Specht et al. 2004, Angulo et al. 2008). Larvae often build a subterranean tunnel which is used to storage?? forage for food, and as place to bring food back to eat (Lafontaine 2004). The economic relevance of A. malefida has motivated various studies on the morphology of the adult (Angulo and Quezada 1975) and immature stages (Rizzo et al. 1995, Angulo et al. 2008), and on various biological aspects (Villata et al. 1988; Rizzo et al. 1995; Baudino 2004) of the species. The biological studies, however, lack do not give estimations of the biological potential of A. malefida. In view of the importance of the species, and its great intraspecific variability, we have endeavored to describe the biological parameters and to estimate the biotic potential of A. malefida in the laboratory, from using individual observations on thousands of exemplars. MATERIALS AND METHODS Our laboratory rearing experiment started with 13 couples (F1) obtained from a mass rearing maintained by the Laboratrio de Biologia, Campus Universitrio da Regio dos Vinhedos (CARVI), Universidade de Caxias do Sul (UCS). The mass rearing itself came had originated from

5 38 individuals collected in Vale dos Vinhedos, Bento Gonalves, RS, on July 05, 2008. Our experiments were conducted in a controlled environment (25 1C, 70 10% UR and 14 h. photophase). We conducted daily observations as follows: each adult couple (n=13) was maintained inside a cylindrical plastic container (10 cm diameter, 15 cm height). In order to stimulate oviposition, strings of filter-paper of the same length as the plastic vial were inserted into each container and attached to the lid, made of plastic film. The subjects were fed an aqueous sucrose solution (10%), made available in hydrophilic cotton, . with a piece of cotton. The following parameters were evaluated: fecundity (number of eggs per female), fertility (number of hatched larvae per female), longevity and duration of pre-oviposition, post-oviposition and oviposition. For the egg stage, we assessed the viability and duration of the incubation period of five random oviositions from each couple (including first and last ovipositions), totaling 2,410 eggs. Each egg posture was individualized in a Petri dish, where it remained until the larvae hatched. [In order to avoid desiccation] the bottom of each Petri dish was lined with filter paper moistened with distilled water After hatching, neonate larvae were placed in separate 150 ml plastic cups closed with a lid, and fed an artificial diet (Greene et al. 1976). We evaluated the survival, and the duration of the larval period of 2,331 individuals. The pre-pupal period was considered separately, and was deemed to begin its onset when was recorded when the larvae stopped feeding. During the pre-pupal period, individuals decrease in size and metamorphose into a pupa. Because individuals in that phase period have Due to the hypogean habits of individuals in that period, we added expanded vermiculite moistened with distilled water to each cup. Pupae were kept in the same containers as pre-pupae, and were not offered any food. On the second day after pupation, when the cuticle had hardened, we ascertained the sex of each individual with the help of sketches from Angulo et al. (2008). Besides ascertaining the duration of the pupal period, we measured the weight, the greater width between pterotecae, and the total length of each pupa.

6 We calculated the biological potential from the biological variables, considering the environmental resistance to be null. The fertility life table was calculated using the equations of Silveira Neto et al. (1976) and the following parameters were determined: net reproductive rate (Ro); mean generation time (T); natural intrinsic growth rate (rm) and infinite growth rate (). The temporal and morphometric parameters were analyzed using descriptive statistics to calculate the means and standard errors. When necessary, the means were compared by t test assuming unequal variances at a significance level of 95%. Voucher specimens (10 couples) were deposited in the collection of the CARVI-UCS.

RESULTS AND DISCUSSION Adult phase Longevity, representing 7.384% of the development cycle of A. malefida, did not significantly differ between genders (p = 0.076) (Figure 1, Table 1). When compared with results from other studies, the longevity of A. malefida, in our data, was significantly higher longer than in the results obtained by Villata et al. (1988) (4.86 days), and within the range of those obtained by Rizzo et al. (1995) (3 -9 days) for the same species. When reared at temperatures near those used in our experiment, the longevity of the congener A. ipsilon was equivalent to that of A. malefida (eg Bento et al. 2007 ~ 13 days). However, in our experiment, individuals of A. malefida spent more time in the larval and pupal phases with respect to individuals of A. ipsilon (e.g. Santos & Nakano 1982 ~ 24.3%; Bento et al. 2007 ~ 24.5%), who spent relatively more time in the adult stage (Figure 1). The proportion of the lifecycle spent as an adult was much lower in A. malefida than in [other] species whose adults experience aestivation, for instance A. infusa (Boisduval, 1832) (Common 1954). It was also much lower than in noctuids with underground habits such as Peridroma saucia (Hbner, 1080) (Moreno Fajardo and Cardona Serna 2006 ~ 17.41%) and epigean species such as Anicla infecta (Ocsenheimer, 1816) (Teston et al. 2001 ~ 24.25%) and Anicla mahalpa Schaus, 1898 (Specht et al. 2008 ~ 17.61%).

7 The relatively short pre-oviposition period (Table 2) indicates that, under the conditions of this study, adults of A. malefida reach sexual maturation soon after emergence. Females of A. malefida, as those of A. ipsilon, are able to lay eggs two days after having emerged (eg Harris et al. 1962, Swier et al. 1976, Santos and Nakano 1982, Bento et al. 2007). The egg laying period, which occurs from the second to the tenth day after sexual maturation, and the highest fecundity, between the seventh and the tenth days (Figure 2), also resemble the values obtained for A. ipsilon (eg Harris et al. 1962, Swier et al. 1976, Santos and Nakano 1982, Bento et al. 2007). The mean fecundity of A. malefida, approximately 1,700 eggs per female (Table 2), resembles that obtained by Rizzo et al. (1995), who reported between 1,000 and 1,600 eggs per female; it differs from the results of Villata et al. (1988), who reported an average of only 474.5 eggs per fertile female. Our results are also consistent with values obtained for other Noctuinae, for instance A. ipsilon (eg Swier et al. 1976 - 1,423.53; Archer et al. 1980 - Maximum average 1.984; Santos and Nakano 1982 to 1.263, Bento et al. 2007 - 1.806 eggs), A. ignicans (Foester and Mello 1996 - 1.807) and A. mahalpa (Specht et al. 2008 - 2.014). Egg phase The average duration of the embryonic period corresponded to 4.557% of the development cycle (Figure 1, Table 1) of our A. malefida subjects, longer than previously reported by Rizzo et al. (1995), who found that this phase lasts five to six days under lower temperatures (22-23 C). Our results, however, resemble those of Villata et al. (1988), who reported that the egg phase in A. malefidathat species lasted eight days when experiments were conducted at 25 C and relative humidity of 60%. The embryonic period of A. malefida is relatively long when compared with that of several other noctuidsNoctuinae representatives ???, representatives of Noctuinae reared under similar temperatures, for instance A. ipsilon (eg Harris et al. 1962 - 25 C - 4 days; Santos and Nakano 1982 to 25 C - 4 days, Bento et al. 2007 - 26 C - 3.3 days), A. infecta (Teston et al. 2001 - 25 C - 3.2 days), A. mahalpa (Specht et al. 2008 - 20 C - 6 days) and P. saucia (Moreno Fajardo Cardona Serna and 2006 to 23.7 C - 5 days).

8 The elevated viability of eggs of A. malefida (Table 1), including those from the first oviposition, suggests that females were fertilized soon after emergence. These results differ from those obtained for A. ipsilon females, which are less attractive to males in their first [few] days after emergence (Swier et al. 1976), resulting in infertile first ovipositions, and consequently low average values of relatively viability (Santos and Nakano 1982 to 64.45%, Bento et al. 2007 to 81.00%).

Larval phase The average duration of the larval stage (including the pre-pupa) of A. malefida, in our experiment, was almost four months (Table 1), accounting for two thirds of the entire life cycle (Figure 1) of the species. This result was similar to the 93.2 days reported for the same species when reared at 25 C and 60% humidity, but very different from the 36.14 days found for specimens reared under the same temperature and 50% relative humidity (Villata et al. 1988). In the present study, larvae remained in separate plastic cups in an incubator with relative humidity of 70 10%. They were fed an artificial diet rich in water. After the third or fourth instars, larvae dug a tunnel in their food diet , where they remained. Therefore, the effective experimental humidity was above that used by Villata et al (1988), and the added moisture was likely responsible for increasing the duration of the larval phase. The direct relationship between increased moisture and increased duration of the larval phase in A. malefida (Villata et al. 1988) contrasts with results described for A. ipsilon. Moisture generally does not influence the duration of the developmental phases of the latter specids, except in the first five stages, when it significantly accelerates the larval stage (Archer et al. 1980). The larval phase of A. malefida, corresponding to 66.56% of the species entire life cycle, lasts longer than in other noctuds such as A. ipsilon (eg Harris et al. 1962 - 48.99%; Santos and Nakano 1982 - 43.42%, Bento et al. 2007 - 46.65%), P. saucia (Moreno Fajardo Cardona Serna and 2006 - 43.15%), A. infecta (Teston et al. 2001- 44.09) and A. mahalpa (Specht et al. 2008 48.18%). Furthermore, the larval phase of A. malefida shows greater variability (Tables 1, 3), a fact

9 that had been also mentioned by Villata et al. (1988) and Rizzo et al. (1995), though to a lesser extent. However, the percent time larvae are active and feeding (Table 1, Figure 1) (31.165%) is similar among A. malefida, A. ipsilon (eg Harris et al. 1962 - 45.37%, Santos and Nakano 1982 38.82%), P. sauce (Moreno Fajardo Cardona Serna and 2006 - 40.06%), A. infecta (Teston et al. 2001 - 37.48%) and A. mahalpa (Specht et al. 2008 - 42.14%). Females spent more time in the larval stage with respect to males (Table 3). This difference can be explained by the fact that females are in on average larger than their male counterparts (Table 5). Despite the great difference in the duration of the larval phase between sexes, excluding the pre-pupal period, the variation in the duration of this phase was similar, and did not exceed 33 days for either sex. Most females (81.43%) developed between 40 and 59 days, and only 41 (5.04%) took longer than 70 days (Table 4) to develop. Similarly, most males (74.06%) completed their larval phase between 50 and 59 days, and only 30 individuals (3.89%) took more than 70 days to complete their development. The duration of the pre-pupal stage of A. malefida was also longer for females (118 days) than for males (118 days). After rearing the same species under four different temperature regimens and humidity between 50% -60%, Villata et al. (1988) found that individuals remained in the prepupal stage between 3-25 days. In their study, however, the last instar (presumably the pre-pupa) lasted longer in averageon average when temperatures were set between 19C and 25C, and the relative humidity was highest. Their results corroborate our findings that higher humidity levels lengthen the larval period of A. malefida. We have not found reports of other species of Agrotis with long pre-pupal periods. However, some noctuid species in other genera, for instance Xestia cnigrum Linnaeus (e.g. Honk 1979, Oku 1984), Helicoverpa armigera (e.g. Wilson et al. 1979, Qureshi et al. 2000, Feng et al. 2010), and Sesamia nonagrioides (e.g. Fantinou et al. 1996, Gadenne et al. 1997, Eizaguirre et al. 2008) experience diapause, overwintering in the larval or pupal stages.

10 The highest frequencies (above 15%) of males and females in the pre-pupal period occurred between 40 and 80 days into the experiment. A few pre-pupae were found between days 0-20 (n= 4 ~ 0.25%), and also after day 100 (n= 32 ~ 2.02%) (Table 4). The regression analysis (n= 1,583; r2 = 0.066; p< 0.001) indicated a direct correlation between the number of days a larva remained active, and the duration of the pre-pupal stage. Even thought we have not investigated the number of larval instars of our subjects, Villata et al (1988) reported that individuals of A. malefida go through seven to 11 instars. Additionally, according to them, the greater number of instars occurs in when the longer larval phasesphase is longer. In view of the fact that the number of instars is greater in noctuids that undergo diapause (Esperk et al. 2007, Gadenne et al. 1997), and given the results found in our study, combined with those of Villata et al. (1988), we hypothetizehypothesize that A. malefida larvae undergo larval diapause. Despite our precautions, only 71.26% of the larvae survived our experiment. Mortality was greater in the pre-pupal period (Table 1). Our values are lower than those obtained for A. ipsilon fed an artificial diet (eg Bento et al. 2007). The lower larval survival rate of A. malefida, especially in the pre-pupal stage, must be associated with the longer duration of the larval period in this species. A similar observation was made by Eizaguirre et al. (2008), who noted that diapausing larvae of S. nonagrioides had low survival rates. Puppal phase The pupal period of A. malefida accounted for 21.501% of the development cycle (Figure 1, Table 1) and is in the range reported by Rizzo et al. (1995) (29 to 44 days) and Baudinos (2004) (41.0 20 days) for the species. However, the variation obtained in this study was much higher (Tables 1, 3, 4), as 98.423% of subjects had a larval period lasting 31 to 50 days. These results demonstrate that, like other Noctuids such as H. armigera (eg Wilson et al 1979, Qureshi et al. 2000), the pupa of A. malefida may also go through diapause. The survival of A. malefida pupae was relatively high (Table 1) and comparable with that of

11 most noctuids reared in the laboratory, for instance A. ipsilon (eg Reese et al. 1972, Bento et al. 2007) and A. mahalpa (Specht et al. 2008). The length and width of A. malefida larvae (Table 5) were compatible to those obtained by Rizzo et al. (1995), and greater than those described by Angulo et al. (2008) [for the species]. The fact that female pupae were significantly larger and heavier than their male counterparts (Table 5) had been previously described as sexual dimorphism ( Angulo et al. 2008 ) Biotic potential and life and fertility table The sex ratio calculated for the 1,588 immatures (813 females and 775 males) was 0.512. Each female oviposited, on average, 1,696.769 eggs,eggs; the overall egg survival was 65.892%, yielding 1,118.035 viable individuals per female. The average duration of the life cycle (174.075 days) corresponded to 2.097 generations per year (n). Thus, considering the environmental resistance as null, we obtained the following result for the equation PB = (rs * d) n - RA - PB = (0.512 x 1118.035) 2.097 to 0 = 6067 x 105 individuals per female. In other words, each female generates more than half a million young youth a year. This value number is very low when compared to A. ipsilon, which presents a the biotic potential of A. ipsilon, approximately 4.120 x 1018 (calculated using the data from Bento et al., 2007). The net reproductive rate (Ro) was 322.6 times per generation, and the mean generation time (T) was 23.97 weeks. The intrinsic rate of increase (rm) was 0.24, with a finite rate of increase () - the number of females added to the population, per female that will generate another female 1.27. The maximum rates of population increase occurred between days 168 and 170, on the 24th week of life (Figure 2). These values numbers are also lower than those obtained for A. ipsilon by Bento et al. (2007), who reported a Ro of 616.9 times. The discrepancy between our results and those mentioned above can be explained by the shorter generation time of A. ipsilon with respect to A. malefida (6.86 weeks, compared to 23.97 weeks), rm = 0.94 and = 2.55, calculated in weeks, as postulated in Silveira Neto et al. (1976).

12 Final considerations Even though Angulo and Quezada (1975) observed morphological and behavioral our results demonstrated that, the life

similarities between adults of A. malefida and A. ipsilon

clycle of both species are very different, being longer in A. malefida. Angulo and Quezada (1975) salientarem Despite many morphological and behavioral similarities between adultos de A. malefida and other species such as A. ipsilon (Angulo and Quezada, 1975), nossos resultados demonstraram que, com relao ao ciclo de vida, both species so muito have different life cycles, which is longer in A. malefida. The long pre-pupal and pupal periods described for many individuals differentiates this species from A. ipsilon (eg Harris et al. 1962, Benedict et al. 2007) and other noctuids. In warmer regions there is more than one generation per year, and adults can be collected throughout the entire year (Lafontaine 2004). The results of this study have shown that A. malefida, like other representatives of Noctuidae, can go through larval and pupal diapause (pre-pupal), highlighting the need to develop studies to assess their response to environmental variations such as photoperiod, humidity, temperature or a combinations thereof . The observation that A. malefida goes through diapause indicates that other species with similar distribution such as Agrotis brachystria (Hampson, 1903) and Agrotis gypaetina Guene, 1852 (Specht et al. 2004) may also present this type of physiological behavior. Although larval or pupal diapause were not expected to occur in A. malefida under the rearing conditions of our experiment (25 1 C, 70 10% RH and 14 hours photoperiod), a study on S. nonagrioides (Eizaguirre et al. 2008) mentions a few individuals expressing diapause when reared under similar environmental conditions.

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14 IGARZBAL D, FICHETTI P AND TOGNELLI M. 1994. Claves practicas para la identificacion de larvas de lepidoptera em cultivos de importancia agricola em cordoba (Argentina). Gayana Zool 58: 99-142. KITCHING IJ AND RAWLINS JE. 1998. 19 - The Noctuoidea, p. 355-401. In: In: Kristensen, N.P. (ed.) Lepidoptera, Moths and Butterflies vol. 1: Evolution, Systematics and Biogeography. Berlin: Walter de Gruyter, 491p. LAFONTAINE JD. 2004. Noctuoidea, Noctuinae, Agrotini. In: HODGES RW, DAVIS DR, FERGUSON DC, MUNROE EG AND POWELL JA. (Eds) The moths of America North of Mexico. Fasc 25 -1. Allen Press, Lawrence, Kansas. MORENO FAJARDO ML AND SERNA CARDONA FJ. 2006. Biologa de Peridroma saucia (Lepidoptera: Noctuidae: Noctuinae) en flores cultivadas del hbrido comercial de Alstroemeria spp. Rev Fac Nac Agron - Medelln, 59 (2): 3435-3448. OKU T. 1984. Larval diapause in the spotted cutworm, Xestia c-nigrum Linn (Lepidoptera: Noctuidae. Appl Ent Zool, 19(4): 483-490. PASTRANA JA. (Coord.) 2004. Los lepidpteros argentinos: sus plantas hospedadoras y otros sustratos alimentcios. 1 ed. Buenos Aires: Sociedad Entomolgica Argentina, 350p. POGUE M. 2006. The Noctuinae (Lepidoptera:Noctuidae) of Great Smoky Mountains National Park, U.S.A. Zootaxa 1215: 1-95. POOLE RW. 1989. Noctuidae. In: HEPPNER JB. (Ed). Lepidopterorum Catalogus. New York, Brill, v. 2. p. 501-1013 QURESHI MH MURAI T YOSHIDA H AND TSUMUKI H. 2000. Populational variation in diapause-induction and -termination of Helicoverpa armigera (Lepidoptera: Noctuidae). Appl Entomol Zool, 35 (3): 357-360 REESE JC ENGLISH LM YONKE TR AND FAIRCHILD ML. 1972. A method for rearing Black Cutworms. J Econ Entomol, 65(4): 1047-1050. RIZZO HF, LA ROSSA FR AND FOLCIA AM. 1995. Aspectos Morfolgicos Y Biologicos del Gusano Aspero (Agrotis malefida (Guene)) (Lep.: Noctuidae). Rev Facul Agron 15 (2-3): 199206. SANTOS HR AND NAKANO O. 1982. Dados Biolgicos sobre a lagarta rosca Agrotis ipsilon (Hufnagel, 1776) (Lepidoptera, Noctuidae). An Soc Entomol Bras, 11: 33-48. SILVA AGDA, GONALVES CR, GALVO DM, GONALVES AJL, GOMES J, SILVA MM AND SIMONI L. 1968. Quarto catlogo dos insetos que vivem nas plantas do Brasil, seus parasitos e predadores. Parte II 1 Tomo. Rio de Janeiro, Ministrio da Agricultura, 622p. SILVEIRA-NETO S, NAKANO O, BARBIN D AND VILLA NOVA NA. 1976. Manual de ecologia dos insetos. So Paulo: Editora Agronmica Ceres, 420p. SPECHT A AND CORSEUIL E. 1996. Lista documentada dos noctudeos (Lepidoptera; Noctuidae) ocorrentes no Rio Grande do Sul, Brasil. Biocincias 4: 131-170. SPECHT A AND CORSEUIL E. 2002. Diversidade dos noctudeos (Lepidoptera, Noctuidae) em Salvador do Sul, Rio Grande do Sul, Brasil. Revista Brasileira de Zoologia (Supl.1): 281-298.

15 SPECHT A, SILVA EJE AND LINK D. 2004. Noctudeos (Lepidoptera, Noctuidae) do Museu Entomolgico Ceslau Biezanko, Departamento de Fitossanidade, Faculdade de Agronomia Eliseu Maciel, Universidade Federal de Pelotas, RS. Rev. Bras. Agrocincia 10 (4): 389-409. SPECHT A, FORMENTINI AC AND CORSEUIL E. 2008. Bionomy of Anicla mahalpa Schaus, 1898 (Lepidoptera: Noctuidae: Noctuinae), in the laboratory. Braz J Biol, 68(2): 415-418. SWIER SR, RINGS RW AND MUSICK GJ. 1976. Reproductive behavior of the black cutworm Agrotis ipsilon. Ann ent Soc Am 69:546-550. TESTON JA SPECHT A AND CORSEUIL E. 2001. Biology of Anicla infecta (Ochsenheimer, 1816) (Lepidoptera, Noctuidae, Noctuinae), under laboratory condictions. Braz J Biol, 61(4): 661666. VILLATA CA, LIMONTI MR AND CASTELLANO SR. 1988. Estudio biolgico de Agrotis malefida (Guen.). Revista Agropecuria de Manfredi Y Marcos Juarez 6(2): 37-51. WILSON AGL, LEWIS T AND CUNNINGHAM RB. 1979. Overwintering and spring emergence of Heliothis armigera (Hbner) (Lepidoptera: Noctuidae) in the Namoi Valley, New South Wales. Bull Entomol Res 69: 97-109.

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Pupae 21,50% Adults 7,38% Pre pupae 35,39% Eggs 4,56%

Larvae 31,17%

Figure 1. Percentage of each developmental periods for Agrotis malefida reared at 25 1C, UR 70% 10% UR and 14 hours photophase.

17

Figure 2. Relationship between fecundity (mx) and survival rate (lx) of Agrotis malefida reared on an artificial diet at 25 1 C, 70 10% RH and 14 hour photophase.

18 Table 1 Survival and duration of the life cycle of Agrotis malefida in different stages of development, under controlled conditions (25 1 C, 70 10% RH and 14 hour photophase). Developmental phase Adult Egg Larvae Pre-pupae Pupae Total immature Total Inicial-final N 26 2,410-2,331 2,331-2,127 2,127-1,667 1,667-1,588 1,588 ----Survival (%) ----96.722 91.248 78.373 95.261 ----65.892 Duration (days) 12.853 3.783 7.933 1.221 54.251 6.391 61.611 17.559 37.427 4.132 161.289 17.352 174.075 MinimumMaximum 5 22 6 10 44 77 10 131 11 99 102 - 227 -----

19 Table 2: Length of pre- post-, and oviposition (days) and mean fecundity of A. malefida in controlled conditions (25 1 C, 70 10% RH and photophase of 14 hours). N = 13 couples Parameter Pre-ovipostion Ovipostion Post-ovipostion Fecundity X EP 2.538 1.050 9.692 3.038 0.154 0.376 1696.769 412.730 Minimum-Maximum 1-4 5 - 15 0-1 992 - 2170

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Table 3 Duration of larval and pupal phases, considering males and females separately. The prepupae period is separated from the larval period. Females (n= 813)
X EP

Males (n= 775) MinimumMaximum (44 77) (10 128) (16 99) (117 227)
X EP

MinimumMaximum (46 77)* (18 131)ns (11 71)* (102 225)*

Larvae Pre-pupae Pupae

54.7716.637 62.23017.282 37.7824.24

53.7036.048 60.92717.830 37.0533.983 159.68417.231

Total* 162.81317.339 * P < 0,001; ns = P> 0,05

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Table 4 Frequency of females and males of Agrotis malefida, at intervals of ten days, with larvae actively feeding, pre-pupae and pupae under controlled conditions (25 1 C, 70 10% RH and photophase of 14 hours).
LARVAE Interval days 10-19 20-29 30-39 40-49 50-59 60-69 70-79 80-89 90-99 100-109 110-119 120-129 130-139 Total Female N ------190 473 109 41 ------------813 % ------23.37 58.18 13.41 5.04 ------------100.00 N ------92 574 76 30 ------------775 Male % ------11.87 74.07 10.19 3.87 ------------100.00 PRE-PUPAE Female N % 1 0.12 13 1.60 59 7.26 143 17.59 144 17.71 178 21.89 148 18.20 85 10.46 27 3.32 7 0.86 7 0.86 1 0.12 ----813 100.00 Male N % 0.39 3 2.84 22 6.32 49 166 21.42 133 17.16 161 20.77 124 16.00 79 10.19 2.71 21 1.42 11 0.52 4 0.13 1 0.13 1 775 100.00 PUPAE Female N % 3 0.37 5 0.62 579 71.22 222 27.31 ----3 0.37 --------1 0.12 ----------------813 100.00 Male N % 3 0.387 5 0.645 611 78.839 151 19.484 2 0.258 1 0.129 2 0.258 ------------------------775 100.00

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Table 5: Mean and standard error of length and width, correlation coefficient and t test of measures of the pupae of Morfometric data of A. malefida pupae under controlled conditions (25 1 C, 70 10% RH and photophase of 14 hours). Weight (g) Females (n= 194) Males (n= 131)
*P< 0,01; **P< 0,001

Width (mm) 8.037 0.512 7.684 0.424 **

Length (mm) 26.389 1.799 25.916 1.555 *

0.888 0.151 0.806 0.109 **