Biology of Agrotis malefida Guenée (Lepidoptera: Noctuidae: Noctuinae): a true diapausing

diapausant cutworm?

Alexandre Specht1,2,*, Andrés O. Angulo3, Tania S. Olivares3, Edegar Fronza1, Eduardo Valduga1,

Francine Albrecht2, Graziela Poletto1, Neiva Monteiro de Barros2.

1 - Laboratório de Biologia, Universidade de Caxias do Sul, Campus da Região dos Vinhedos.

Alameda João Dall Sasso, 800, Cx. Postal 32. Bairro Universitário, Bento Gonçalves, RS. CEP

95700-000.

2 - Instituto de Biotecnologia, Universidade de Caxias do Sul.

3 – Universidad de Concepción, 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.

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Abstract

This study aimed to estimate the biotic potential of Agrotis malefida Guenée, 1852 (Lepidoptera:

Noctuidae: Noctuinae) under laboratory conditions. The insects were reared in a controlled

environment (25 ± 1 º C, 70 ± 10% RH and photophase of 14 hours) and observed daily. The larvae

were fed Greene’s 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 pre-

pupae was 96.72, 91.25, 78.37 and 95.26%, respectively. The average duration of eggs, larvae, pre-

pupae, 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 biótico de Agrotis malefida Guenée, 1852

(Lepidoptera: Noctuidae: Noctuinae) em condições laboratoriais. Os insetos foram criados em sala

climatizada (25 ± 1ºC, 70 ± 10% UR e fotofase de 14 horas) com observações diárias. As larvas

foram alimentadas com dieta artificial de Greene e os adultos com solução aquosa de sacarose a

10%. Nas fases imaturas avaliaram-se a viabilidade e a duração, a partir de 2,410 ovos, cujas larvas

foram individualizadas logo após a eclosão. Na fase adulta, avaliou-se a longevidade, períodos de

pré, pós e oviposição, fecundidade e fertilidade de 13 casais. A viabilidade das fases de ovo, larva,

pré-pupa e pupa foi de 96.72, 91.25, 78.37 e 95.26%, respectivamente. A duração média das fases

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de ovo, larva, pré-pupa, pupa e adulto foi de 7.93, 54.26, 61.61, 37.43 e 12.85 dias,

respectivamente. O período de desenvolvimento dos imaturos de A. malefida teve duração média

de 161.289 dias, variando de 102 a 227 dias. O ciclo de vida de A. malefida é bem mais longo do

que de outras espécies do gênero. A fecundidade média foi de 1,696.769 ovos e a fertilidade

1,641.149 larvas por fêmea. Nas condições em que foi realizado este estudo o potencial biótico de

A. malefida foi de 606.666,593 indivíduos/fêmea/ano. Os resultados também indicaram que esta

espécie apresenta diapausa larval (pré-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).

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Agrotis malefida Guenée, 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 (Igarzábal 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 áspero”, “oruga áspera” or “oruga cortadora áspera”

(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 Laboratório de Biologia, Campus Universitário da Região dos Vinhedos

(CARVI), Universidade de Caxias do Sul (UCS). The mass rearing itself came had originated from

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38 individuals collected in Vale dos Vinhedos, Bento Gonçalves, RS, on July 05, 2008. Our

experiments were conducted in a controlled environment (25 ± 1ºC, 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.

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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 (λ).

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

(Hübner, 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%).

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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 º C - 4 days; Santos and

Nakano 1982 to 25 º C - 4 days, Bento et al. 2007 - 26 º C - 3.3 days), A. infecta (Teston et al. 2001

- 25 º C - 3.2 days), A. mahalpa (Specht et al. 2008 - 20 º C - 6 days) and P. saucia (Moreno Fajardo

Cardona Serna and 2006 to 23.7 º C - 5 days).

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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

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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 pre-

pupal 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 19ºC and 25ºC, 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 c-

nigrum Linnaeus (e.g. Honěk 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.

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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

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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

(λ) - 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 λ = 2.55, calculated in weeks, as

postulated in Silveira Neto et al. (1976).

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Final considerations

Even though Angulo and Quezada (1975) observed morphological and behavioral

similarities between adults of A. malefida and A. ipsilon¸ our results demonstrated that, the life

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 relação ao ciclo de vida, both species

são 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 Guenée, 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 º 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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Adults7,38%

Eggs4,56%

Larvae31,17%

Pre-pupae35,39%

Pupae21,50%

16

Figure 1. Percentage of each developmental periods for Agrotis malefida reared at 25 ± 1°C, 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 º C, 70 ± 10% RH and 14 hour photophase.

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Table 1 – Survival and duration of the life cycle of Agrotis malefida in different stages of development, under controlled conditions (25 ± 1 º C, 70 ± 10% RH and 14 hour photophase).

Developmental phase

Inicial-final N Survival (%) Duration (days) Minimum-Maximum

Adult 26 ----- 12.853 ± 3.783 5 – 22

Egg 2,410-2,331 96.722 7.933 ± 1.221 6 – 10

Larvae 2,331-2,127 91.248 54.251 ± 6.391 44 – 77

Pre-pupae 2,127-1,667 78.373 61.611 ± 17.559 10 – 131

Pupae 1,667-1,588 95.261 37.427 ± 4.132 11 – 99

Total immature 1,588 ----- 161.289 ± 17.352 102 - 227

Total ----- 65.892 174.075 -----

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Table 2: Length of pre- post-, and oviposition (days) and mean fecundity of A. malefida in controlled conditions (25 ± 1 º C, 70 ± 10% RH and photophase of 14 hours). N = 13 couples

Parameter ± EP Minimum-Maximum

Pre-ovipostion 2.538 ±1.050 1 - 4

Ovipostion 9.692 ± 3.038 5 - 15

Post-ovipostion 0.154 ± 0.376 0 - 1

Fecundity 1696.769 ± 412.730 992 - 2170

X

20

Table 3 – Duration of larval and pupal phases, considering males and females separately. The pre-pupae period is separated from the larval period.

Females (n= 813) Males (n= 775)

± EP Minimum-

Maximum

± EP Minimum-

Maximum

Larvae 54.771±6.637 (44 – 77) 53.703±6.048 (46 – 77)*

Pre-pupae 62.230±17.282 (10 – 128) 60.927±17.830 (18 – 131)ns

Pupae 37.782±4.24 (16 – 99) 37.053±3.983 (11 – 71)*

Total* 162.813±17.339 (117 – 227) 159.684±17.231 (102 – 225)** P < 0,001; ns = P> 0,05

X X

21

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 º C, 70 ± 10% RH and photophase of 14 hours).

LARVAE PRE-PUPAE PUPAEInterval

days Female Male Female Male Female Male

N % N % N % N % N % N %

10-19 --- --- --- --- 1 0.12 3 0.39 3 0.37 3 0.387

20-29 --- --- --- --- 13 1.60 22 2.84 5 0.62 5 0.645

30-39 --- --- --- --- 59 7.26 49 6.32 579 71.22 611 78.839

40-49 190 23.37 92 11.87 143 17.59 166 21.42 222 27.31 151 19.484

50-59 473 58.18 574 74.07 144 17.71 133 17.16 --- --- 2 0.258

60-69 109 13.41 76 10.19 178 21.89 161 20.77 3 0.37 1 0.129

70-79 41 5.04 30 3.87 148 18.20 124 16.00 --- --- 2 0.258

80-89 --- --- --- --- 85 10.46 79 10.19 --- --- --- ---

90-99 --- --- --- --- 27 3.32 21 2.71 1 0.12 --- ---

100-109 --- --- --- --- 7 0.86 11 1.42 --- --- --- ---

110-119 --- --- --- --- 7 0.86 4 0.52 --- --- --- ---

120-129 --- --- --- --- 1 0.12 1 0.13 --- --- --- ---

130-139 --- --- --- --- --- --- 1 0.13 --- --- --- ---

Total 813 100.00 775 100.00 813 100.00 775 100.00 813 100.00 775 100.00

22

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 º C, 70 ± 10% RH and photophase of 14 hours).

Weight (g) Width (mm) Length (mm)

Females (n= 194) 0.888 ± 0.151 8.037 ± 0.512 26.389 ± 1.799

Males (n= 131) 0.806 ± 0.109 7.684 ± 0.424 25.916 ± 1.555

** ** **P< 0,01; **P< 0,001