Overwintering Survival and Spring Emergence of ... - BioOne

POPULATION ECOLOGY

Overwintering Survival and Spring Emergence of Helicoverpa armigera (Lepidoptera: Noctuidae) in Northern Greece G. K. MIRONIDIS,1 D. C. STAMOPOULOS,2

AND

M. SAVOPOULOU-SOULTANI1,3

Environ. Entomol. 39(4): 1068Ð1084 (2010); DOI: 10.1603/EN09148

ABSTRACT Overwintering survival of Helicoverpa armigera (Hu¨ bner) was studied under Þeld conditions during the winter of 2004 Ð2005 and 2005Ð2006 to clarify whether a local population of this insect is established in northern Greece. Additionally, the postdiapause eclosion times of the overwintering generation was compared with adult male dynamics using pheromone traps. Our study supplies strong evidence that part, if not all, of the late-season generation of H. armigera overwinters as diapausing pupa in northern Greece, suggesting that a local population exists in this area. Pupae enter diapause during September and October in response to declining temperatures and photoperiod. A limited number of degree-days in autumn prevented part of the population from developing to the diapausing pupal stage. Larvae of H. armigera that were placed in Þeld conditions late in October were not able to pupate and died. The termination of diapause and the eclosion of adults in the following spring were determined by the local soil temperatures. Diapause terminated between mid-April and early May, and adult emergence occurred within a 4- to 6-wk period from late April to early June. Females emerged slightly earlier than the males. The emergence of 10, 25, 50, 75, and 90% of adults required 153, 199, 252, 303, and 347 DD, respectively. Differences in timing and degree of overlap between adult emergence from the experimental population and pheromone trap catches suggests that the spring population of this species also includes immigrants. KEY WORDS Helicoverpa armigera, survival rate, diapause, overwintering, traps

Insect overwintering is a fascinating process involving many physiological, epidemiological, biochemical, and behavioral changes (Leather et al. 1993). Insects adapt to low temperature in essentially three ways: they survive by entering into a dormant (diapause or quiescent) state (Tauber and Tauber 1976, Tauber et al. 1986, Danks 1987), by displacement (dispersal or migration) (Drake and Gatehouse 1995), or by evolving to function normally at low temperatures, particularly those species inhabiting alpine and polar regions (Sømme and Block 1991). Insects that enter a dormant state exhibit tolerance to a wider range of low temperatures than those that do not (Lee 1991). Dormancy, one of the most fundamental seasonal adaptations of insects (Tauber et al. 1986, Danks 1987), and the regional displacement of various degrees, ranging from short-distance ßights to long-distance migration, are two alternative strategies used by many insects to escape in space or in time from adverse environmental conditions and/or to avoid their habitat deterioration (Southwood 1977).

1 Aristotle University of Thessaloniki, Faculty of Agriculture, Laboratory of Applied Zoology and Parasitology, 54124 Thessaloniki, Greece. 2 University of Thessaly, School of Agricultural Sciences, Department of Agriculture, Ichthyology and Aquatic Environment, 38446 Volos, Greece. 3 Corresponding author, e-mail: [email protected].

Helicoverpa armigera (Hu¨ bner) (Lepidoptera: Noctuidae) seems to exhibit both strategies: in subtropical and temperate parts of its geographic range, the insect survives winter conditions by entering a facultative diapause in the pupal stage in response to short daylengths and decreasing temperatures of autumn (Fitt 1989). In addition, regular movements between tropical and higher latitudes have been observed, which apparently serve to supplement or re-establish populations (Wilson et al. 1979; Fitt and Daly 1990; Gregg et al. 1995; Zhou et al. 2000a, b). In some regions of Africa (Roome 1979), Australia (Wilson et al. 1979, Kay 1982, Zalucki et al. 1986, Fitt and Daly 1990, Murray 1991, DufÞeld and Dillon 2005), Israel (Zhou et al. 2000a), China (Ge et al. 2005), and Japan (Shimizu et al. 2006), Þeld populations of H. armigera have been found to overwinter successfully as diapausing pupae. Populations in the tropics either breed continuously, as in Uganda (Coaker 1959), or only a tiny percentage 2Ð 4% of their pupae diapauses, as in Tanzania (Reed 1965). In northern Greece, H. armigera completes two or three generations per year and causes annual damage, especially to cotton (EPPO/CABI 1997). Despite the fact that H. armigera is the major insect pest of cotton in northern Greece (Mourikis and Vasilaina-Alexopoulou 1969, Mironidis and Savopoulou-Soultani 2008), no data are available on its overwintering in this speciÞc region and generally in Greece. Questions on

0046-225X/10/1068Ð1084$04.00/0 䉷 2010 Entomological Society of America

August 2010

MIRONIDIS ET AL.: OVERWINTERING AND SPRING EMERGENCE OF H. armigera

whether the population of H. armigera that occurs each year is local or is re-established from southern and, apparently, warmer regions with long-distance immigratory ßight remain unanswered. If local overwintering occurs, a reliable method of predicting the emergence of the overwintered H. armigera population would greatly facilitate the understanding of population dynamics and could be useful in making decisions about H. armigera management in the area of northern Greece. Uncertainty as to which plants serve as alternate hosts during this spring emergence period makes difÞcult the population monitoring in the early season. Problems may also arise when extrapolating results to other locations where slight variations in such conditions as temperature, rainfall, and soil type could inßuence the timing of adult emergence (Potter et al. 1981). The accumulation of heat units (degree-days) is a useful tool to predict developmental events of insects and other poikilothermic organisms in the Þeld (Sevacherian et al. 1977, Logan et al. 1979, Potter et al. 1981, Rummel and HatÞeld 1988, McCann et al. 1989, Broufas and Koveos 2000, Milonas et al. 2001, Son et al. 2007). Because temperature is known to have a major inßuence on diapause termination in Heliothis spp. (Roach and Adkisson 1971, Holtzer et al. 1976), heat summation has value, as in some other heliothines (Logan et al. 1979, Potter et al. 1981, Rummel and HatÞeld 1988), in monitoring H. armigera population built-up in the early season after spring adult emergence from overwintering cells (Wilson et al. 1979, Cunningham et al. 1981, DufÞeld and Dillon 2005). Improved methods of forecasting the emergence of overwintering populations of Heliothis spp. could beneÞt decision making in cotton insect pest management programs (DufÞeld 2004, DufÞeld and Dillon 2005). Williams (1977) suggested that Heliothis spring emergence models were directly useful for the implementation of management strategies such as the pre-emergence soil cultivation to increase mortality of pupae in the soil and the prediction of overwintering mortality. The primary objective of our study was to develop an overwintered H. armigera emergence model based on soil temperature. Such a model would allow a better insight into early season H. armigera population development in crop and wild hosts and might be useful in studies of early seasonÕs moth immigration. Furthermore, we gathered more direct evidence about the overwintering dynamics of H. armigera in northern Greece. SpeciÞcally, we determined the incidence and the timing of diapause induction and the diapause termination of H. armigera, and we compared the postdiapause eclosion time in the experimental population to the Þeld adult male dynamics, monitored using pheromone traps. Materials and Methods Insects. The offspring of the F1 and F2 laboratory generation was used in a 2-yr study (2004 Ð2005 and 2005Ð2006) to evaluate the overwintering potential of H. armigera in northern Greece. Approximately 400

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larvae per year of the parental generation (mostly of third and fourth instars) were randomly collected from cotton Þelds (41⬚ ⌵, 023⬚ ⌭) in the Þrst and second 10-d period of August 2004 and 2005. Larvae were placed individually into 9-cm-diameter petri dishes and were provided with cotton squares and small bolls. They were maintained in Þeld conditions until pupation and adult emergence. These adults were used to establish the laboratory colony of H. armigera, which was maintained on artiÞcial diet (Mironidis and Savopoulou-Soultani 2008) and held at 25⬚C, 60 Ð70% RH, under a photoperiod 16:8 (L:D) h. Field Experiments. Newly hatched larvae (Þrst instar) of the laboratory colony were placed in Þeld conditions at the region of their origin and were reared in groups of 25Ð50 individuals in 200-ml plastic cups provided with small cubes of the same artiÞcial diet as the one used for the maintenance of the laboratory colony of H. armigera. The cups with the larvae were maintained inside a large cage (70 by 50 by 30 cm) to be protected from rain, vertebrate predators, and parasitoids. The number of H. armigera larvae that were maintained in Þeld conditions, along with the corresponding dates of placement, are shown in Table 1. After the completion of the third instar, the larvae were transferred into individual 9-cm-diameter petri dishes to prevent cannibalism. Larvae were provisioned with artiÞcial diet whenever this was necessary, and mortality was recorded daily until pupation in the petri dish. Pupal weight was recorded on days 2 and 10 after pupation with an analytical balance (Chyo JK-180; ⫾0.01 mg; Chyo, Osaka, Japan). On pupation, sex was determined and recorded. Pupal diapause status was determined either by the retention of pigmented eyespots (Shumakov and Yakhimovich 1955) or by the maintenance of Þrm round shape lobes of the fat body (Pearson 1958). Using the above criteria, 15 d after pupation, the pupae were checked under a stereoscope (M26, 40⫻; Leica, Wetzlar, Germany) for eyespot retention and for the condition of their liposome to separate diapausing from nondiapausing pupae. After the separation, the nondiapausing individuals were maintained in Þeld conditions in the large cage and inside the plastic petri dishes of their pupation. The number of adults emerging was recorded daily. Nondiapausing pupae were considered dead if they could not rotate their abdomen when disturbed. Larval and pupal survival was calculated as the percentage of individuals that completed the larval and pupal stage, respectively. The incidence of diapause and the survival of larvae and pupae were recorded for each date of larval placement in Þeld conditions. Embedding of Diapausing Pupae Into the Soil. Twenty days after pupation, diapausing pupae from each placement date (Table 1) were embedded into the soil at the region of their origin inside artiÞcial chambers. The depth of embedding (40 Ð 60 mm) was the one preferred by the insect in Þeld conditions to pupate in the soil, creating an appropriate burrow (Wilson 1983, Murray and Zalucki 1994). ArtiÞcial chambers were built to simulate the natural chambers

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ENVIRONMENTAL ENTOMOLOGY

Table 1.

Dates of larval placement in field conditions and diapause incidence (%)

Year

Dates of placement

⌵

Pupation (%)

2004

8 Sept. (Þrst date) 18 Sept. (second date) 12 Oct. (third date) 28 Oct. (fourth date) 21 Aug. (Þrst date) 31 Aug. (second date) 20 Sept. (third date) 1 Oct. (fourth date) 28 Oct. (Þfth date)

600 300 1,000 1,200 160 405 350 390 1,010

2005

Vol. 39, no. 4

␹2 df P

Diapausing pupae (%) Total

么么

乆乆

55.00 (n ⫽ 330) 79.67 (n ⫽ 239) 16.10 (n ⫽ 161) Ñ 33.13 (n ⫽ 53) 21.23 (n ⫽ 86) 46.29 (n ⫽ 162) 25.90 (n ⫽ 101)

65.15 (n ⫽ 215) 30.13 (n ⫽ 72) 62.73 (n ⫽ 101) Ñ 13.21 (n ⫽ 7) 44.19 (n ⫽ 38) 90.12 (n ⫽ 146) 82.18 (n ⫽ 83)

50.70 (n ⫽ 109) 61.11 (n ⫽ 44) 50.50 (n ⫽ 51) Ñ 71.43 (n ⫽ 5) 31.58 (n ⫽ 12) 52.74 (n ⫽ 77) 50.60 (n ⫽ 42)

49.30 (n ⫽ 106) 38.89 (n ⫽ 28) 49.50 (n ⫽ 50) Ñ 28.57 (n ⫽ 2) 68.42 (n ⫽ 26) 47.26 (n ⫽ 69) 49.40 (n ⫽ 41)

Ñ 19.645 1 0.000

Ñ 24.050 1 0.000

Ñ 0.179 1 0.672

Ñ 0.192 1 0.662

Sex ratio (么:乆) 1.03:1 1.57:1 1.02:1 2.50:1 0.46:1 1.12:1 1.02:1

⌵, no. of Þrst instar larvae that were placed in Þeld condition in the corresponding dates; n, no. of pupae that formed the corresponding percentage.

that pupae built in the soil (McCann et al. 1989). On a sterilized polystyrene conical tube with screw cap (17 by 120 mm; Geiner Bio-One, Kremsmu¨ nster, Austria) a small hole, ⬇1 mm in diameter, was drilled through the bottom to permit drainage. The chamber was Þlled with sterilized sand to a depth of 80 mm below the open end. A pupa was placed head up in the chamber on the surface of the sand. A small square of muslin was secured over the open end to prevent predatory insects from devouring the pupa. A vertical hole through the screw cap was drilled to permit air exchange. The complete construction was placed vertically in the ground so that only the screw cap protruded from the soil surface. The result was that the pupae were in darkness, at a controlled depth in the soil, in chambers that approximated their natural burrows. Pupae inside artiÞcial chambers were checked daily for eyespots movement and for adult emergence. Eyespots movement was used as the criterion for diapause termination (Shumakov and Yakhimovich 1955, Wilson et al. 1979, Lopez 1986), as well as to deÞne the onset of postdiapause development for use of degreeday predictions of adult emergence. Sex Pheromone Trapping. Pheromone-baited funnel traps (obtained from Hellafarm, Maroussi, Greece) were used to monitor the ßight activity of male H. armigera. On 1 March 2005 and 1 March 2006, four and Þve traps, respectively, were deployed in Þelds (four Þelds in 2005 and Þve Þelds in 2006, ⬇2 ha each), which later were sown with cotton in late April, on a perimeter of 3Ð 4 km around the Þeld with the embedded chambers containing the diapausing pupae. The distance separating pheromone traps was at minimum 3 km, creating a relatively small network around the Þeld with the embedded chambers. The traps were Þtted with dispensers impregnated with (⌮)-11-hexadecenal and (⌮)-9-hexadecenal, the main sex pheromone of H. armigera (Kehat and Dunkelblum 1990, 1993). The traps were placed just above the crop canopy (30 cm high). Trap height was adjusted according to the crop growth in the Þeld. Dichlorvos (Nuvan 100 EC) at 0.2% was used as a fumigant in the traps to kill trapped moths. Male moths captured in

traps were counted and removed every 2Ð3 d. The pheromone dispensers were renewed every 3 wk to ensure their effectiveness. Trapping operations stopped the Þrst of November of both years of experiments because no captures of males were observed after 15 October of both years. Temperature Recording. Air temperature in the microclimate of the larvae in the Þeld was recorded by means of an electronic climate parameter recorder (HOBO Pro RH/Temp data logger; Onset, Bourne, MA). The device was mounted 1.3 m above the ground and was programmed to record temperature at 30-min intervals. The temperature of the microclimate of the diapausing pupae embedded into the soil was recorded by means of four sensors (TMCx-HD) plugged directly into an electronic climate parameter recorder (HOBO H8 data logger; Onset). One of the sensors was installed in one of the chambers, adjacent to the pupa, measuring and recording automatically the temperature in the pupal chamber. The other three sensors were installed in the soil at depths of 20, 10, and 5 cm. The device was programmed to record temperature data at hourly intervals. Statistical Analysis. The effect of date of larval introduction into the Þeld on developmental time of larval and pupal stages, as well as on pupal weight (diapausing and nondiapausing individuals), was determined by analysis of variance (ANOVA). The differences between dates of larval placement in the Þeld, in number of days for diapause termination, were also determined by a one-way ANOVA. A logarithmic transformation log10(x ⫹ 1) of the data was used to avoid heterogeneity of variance: untransformed means are presented in the tables. Percentages were compared using the ␹2 test (Sokal and Rohlf 1995). The t-test was used to compare the values of the means from two samples. Calculating Degree-Days. The method of Baskerville and Emin (1969) based on the lower threshold temperature (LTT) was used to calculate daily degree-days from minimum-maximum temperature data for the development of larval and overwintering pupal stages of H. armigera. In an earlier laboratory devel-

August 2010


opmental study, at constant temperatures, we estimated the LTT for development of each immature life stage of the Greek population of H. armigera (Mironidis and Savopoulou-Soultani 2008). The linear approximation method was used to calculate the LTT for larval and pupal stage development of nondiapausing individuals, which was estimated to be 10.5 and 10.2⬚C, respectively. However, the overwintering pupal cycle of H. armigera can be divided into two phases. Phase I is associated with cold exposure requirements of diapausing pupae, whereas phase II is a reactivation development in response to increasing spring temperatures that culminates in diapause termination (eyespots movements) and allows the resumption of normal pupal development, which is completed with adult emergence (Roome 1979, Wilson et al. 1979, Hackett and Gatehouse 1982, McCann et al. 1989). Pupae in either of the two phases cannot be differentiated morphologically, and separation is based primarily on the response of the diapause pupae to temperatures suitable for terminating diapause (Murray and Wilson 1991, Mironidis 2009). Once the requirement of exposing the pupae to cold has been met (phase I), the pupae remain dormant until temperatures become warm enough to begin the phase II (Logan et al. 1979, Danks 1987, McCann et al. 1989). The temperature threshold for the reactivation of development (phase II) of diapausing pupae of H. armigera is higher than that for the normal development of nondiapausing pupae (Fitt 1989). Wilson et al. (1979), with constant-temperatures studies for the Australian population of H. armigera, reported that the second phase of diapause development commences at a temperature of ⬇17⬚C, whereas G.K.M. and M.S.-S. (unpublished data) using laboratory data for diapause development at a range of constant temperatures for the Greek population found that temperatures ⬎15.9⬚C are needed for the completion of this phase. In our degree-day estimations for larval development, we used 10.5⬚C as LTT, whereas the total number of required degree-days for adult emergence from overwintering pupae was calculated using 15.9⬚C as LTT for phase II of diapause development (Þrst April to eyespots movement) and 10.2⬚C as LTT for postdiapause development (eyespots movement to adult emergence). The LTT for postdiapause development is considered to be the same with nondiapause pupae (10.2⬚C), because it has been found for H. armigera that once diapause terminated, postdiapause development proceeds at a rate similar to that for nondiapause pupae (Wilson et al. 1979, Foley 1981, Murray and Wilson 1991, Mironidis 2009). Furthermore, the Þrst of April was used as the beginning point for phase II of diapause development because after this date mean soil temperatures in Þeld conditions in northern Greece (Fig. 5) were above the LTT (15.9⬚C) for phase II. The use of these different LTTs for phase II diapause development/termination and for postdiapause development conÞrms the degree-day estimations. Distribution Model of Developmental Time. The variation in developmental time for larvae and pupae

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on different dates of larval placement in Þeld conditions was described by a cumulative probability distribution of developmental completion of each corresponding stage against degree-days using a Weibull function (Re´ gnie` re 1984). The Weibull function is often used to describe the distribution of insects completing a developmental stage in response to time or temperature (Wagner et al. 1984). The expression of the function is as follows: f共 x兲 ⫽ 100 关1 ⫺ exp(⫺关 x/␤ 兴 ␣兲], where f(x) is the cumulative percentage of insects entering a life stage, x is degree-days, and ␣ and ␤ are model parameters that were estimated using the Marquardt algorithm in SPSS NLN 14 (SPSS 2006). Results Larval Development and Diapause Induction Under Field Conditions. The percentage of H. armigera larvae that pupated under Þeld conditions and the percentage of diapause incidence along with sex ratio of diapausing pupae are shown in Table 1. The percentage of pupation and consequently of survival was signiÞcantly different between the dates of larval placement in Þeld conditions. In 2004, it ranged from 16 to 80 (␹2 ⫽ 496.00; df ⫽ 2; P ⬍ 0.001), and in 2005, it ranged from 21 to 46 (␹2 ⫽ 61.565; df ⫽ 3; P ⬍ 0.001). The percentage of pupation was signiÞcantly different between the 2 yr, irrespective of the date of larval placement (␹2 ⫽ 19.645; df ⫽ 1; P ⬍ 0.001). In both years, H. armigera larvae that were placed in Þeld conditions after mid-October died before reaching the pupal stage. The percentage of pupae that entered diapause showed a signiÞcant relationship with the date of larval placement in the Þeld both in 2004 (␹2 ⫽ 75.908; df ⫽ 2; P ⬍ 0.001) and in 2005 (␹2 ⫽ 141.676; df ⫽ 3; P ⬍ 0.001). In 2004, these percentages were 65, 30, and 63% for the Þrst, second, and third date of placement, respectively. The low percentage (⬇30%) observed at the second date of larval placement can probably be attributed to the high values of temperature that were recorded in the period between 20 September and 10 October 2004 (Fig. 1; Table 1). In 2005, the percentage of diapausing pupae increased gradually with the date of larval placement and showed its highest value (90%) on the third date, whereas on the fourth date, this value was reduced to 82. There was a signiÞcant interaction between the incidence of pupal diapause and the year of experiment (␹2 ⫽ 24.050; df ⫽ 1; P ⬍ 0.001), which was independent of the insect sex (么: ␹2 ⫽ 0.179; df ⫽ 1; P ⫽ 0.672, 乆: ␹2 ⫽ 0.192; df ⫽ 1; P ⫽ 0.662). However, sex ratio of diapausing pupae was found to differ markedly from 1:1 in some dates of placement (second in 2004; Þrst, second, and perhaps third in 2005) for both years of experiments (Table 1). The proportion of diapausing and nondiapausing pupae and the larval developmental time, in relation to the date of larval placement in the Þeld, are shown in Fig. 2 and in Table 2, respectively. The H. armigera larvae that were placed in Þeld conditions on 8 and 18

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Vol. 39, no. 4

Fig. 1. Daily maximum, mean, and minimum air temperature from 1 August to 31 December for 2004 (A), and 2005 (B) and photoperiod throughout the year at northern Greece (C). Daylength was calculated as sunrise to sunset with 0.5 h addition as twilight (Hellenic National Meteorological Service). Horizontal line was drawn through 10.5⬚C as the LTT of larval stage and horizontal distinctive line was drawn through 10.2⬚C as the LTT of pupal stage of nondiapausing individuals. Vertical arrows indicate the dates of larval placement in Þeld conditions.

September 2004 completed their development in ⬇20 d, whereas those that were placed on 12 October 2004 required signiÞcantly longer time for the completion of larval stage, both those that entered diapause later in pupal stage (mean ⫽ 32.68 d, F ⫽ 2,708.325; df ⫽ 2,385; P ⬍ 0.001) and those that did not (mean 37.12 d, F ⫽ 2,708.325; df ⫽ 2,385; P ⬍ 0.001). The same pattern was also observed in 2005; developmental time

for the larval stage increased signiÞcantly with the date of larval placement in the Þeld (Table 2). For those insects that entered pupal diapause, the mean number of days required to complete the larval stage (20.4 d) was signiÞcantly higher (t ⫽ 5.211; df ⫽ 237; P ⬍ 0.001) than that of nondiapausing ones (19.39 d) for the second date of 2004. On the contrary, on the third date, the duration of larval stage of nondiapaus-

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Fig. 2. Percentage of pupation of nondiapausing (䡺) and diapausing (f) individuals of H. armigera in relation to the total number of pupae at each larval placement in the Þeld conditions in 2004 (A) and in 2005 (B).

ing pupae was signiÞcantly longer by 5 d (t ⫽ ⫺5.951; df ⫽ 159; P ⬍ 0.001) than that of diapausing pupae, whereas on the Þrst date, no signiÞcant differences were observed (t ⫽ ⫺0.639; df ⫽ 328; P ⫽ 0.523; Table 2). In 2005, the differences in mean larval developmental time of diapausing pupae were signiÞcantly higher for the Þrst (t ⫽ ⫺2.179; df ⫽ 51; P ⫽ 0.034) and second (t ⫽ ⫺2.956; df ⫽ 84; P ⫽ 0.004) date in relation to nondiapausing pupae, whereas for the third and fourth dates, the larvae that entered diapause in the pupal stage required less time to complete their larval development; yet the differences were signiÞcant only on the fourth date (t ⫽ 3.980; df ⫽ 100; P ⬍ 0.001; Table 2). The date of larval placement in Þeld conditions signiÞcantly affected the pupal weight in both years of experiments (Table 2). Generally, the later they pupated, the smaller the pupal weight was. SpeciÞcally, when weight was recorded on day 2 after pupation, the mean weight of diapausing pupae decreased signiÞcantly with the date of larval placement in the Þeld

both for 2004 (F ⫽ 22.421; df ⫽ 2,384; P ⬍ 0.001) and 2005 (F ⫽ 12.524; df ⫽ 3,271; P ⬍ 0.001). The same pattern was observed for the nondiapausing pupae, with the exception of the second and fourth date of 2005. The diapausing pupae were signiÞcantly heavier (weight recorded on days 2 and 10 after pupation) in relation to nondiapausing pupae for all dates of larval placement, except for pupal weight recorded on day 2 of the Þrst date of 2004 (t ⫽ 2.744; df ⫽ 328; P ⫽ 0.079) and the second (t ⫽ ⫺0.665; df ⫽ 84; P ⫽ 0.508) and fourth (t ⫽ ⫺0.751; df ⫽ 100; P ⫽ 0.462) dates of 2005. On the contrary, the date of larval placement in the Þeld did not signiÞcantly affect the reduction in pupal weight between the Þrst and the second record, but this reduction was signiÞcant between diapausing and nondiapausing pupae (t-test, P ⬍ 0.05; Table 2). The variation in the required degree-days for the completion of the larval stage in relation to the dates of larval placement in the Þeld was well described with the cumulative Weibull distribution model: r2 ⫽ 0.922Ð 0.993 (Fig. 3; Table 3). The value of shape pa-

2005

Values in parentheses are ranges. Means within a column followed by the same letter (lowercase) are not signiÞcantly different (P ⬍ 0.05, Tukey HSD test). Means between sexes in a particular date followed by the same letter (uppercase) are not signiÞcantly different (P ⬍ 0.05, t-test). n, no. of individuals.

Nondiapausing

43.8 ⫾ 14.2aB (5.6Ð259.3) 48.7 ⫾ 19.4aB (7.4Ð288.6) 41.5 ⫾ 4.7aB (2.8Ð184.8) Ñ 1.787 2, 308 0.000 33.1 ⫾ 7.5aB (4.3Ð292.5) 52.8 ⫾ 12.1aB (7.6Ð339.9) 76.1 ⫾ 24.9aB (4.4Ð320.1) 51.2 ⫾ 15.5aB (2.8Ð295.2) Ñ 1.487 3, 124 0.221 4.3 ⫾ 0.5aA (1.2Ð24.1) 5.3 ⫾ 0.8aA (0.6Ð28.3) 5.9 ⫾ 0.6aA (1.4Ð32.0) Ñ 1.682 2, 384 0.000 4.1 ⫾ 0.5aA (1.8Ð6.0) 5.0 ⫾ 0.7aA (1.0Ð22.0) 5.6 ⫾ 0.3aA (0.3Ð25.1) 4.8 ⫾ 0.3aA (0.9Ð17.9) Ñ 1.035 3, 271 0.378 325.4 ⫾ 5.2aA (191.3Ð429.6) 312.8 ⫾ 3.2bB (229.5Ð382.6) 290.5 ⫾ 4.8cB (221.7Ð393.6) Ñ 19.647 2, 308 0.000 300.1 ⫾ 0.1abB (193.7Ð387.7) 321.4 ⫾ 5.5aA (212.0Ð396.2) 258.8 ⫾ 10.7cB (193.1Ð320.8) 284.4 ⫾ 5.4bcA (190.6Ð450.1) Ñ 9.056 3, 124 0.000 20.25 ⫾ 0.14aA (n ⫽ 115) 19.39 ⫾ 0.11aB (n ⫽ 167) 37.12 ⫾ 0.73bB (n ⫽ 60) Ñ 1,029.491 2, 339 0.000 15.85 ⫾ 0.33aB (n ⫽ 46) 18.29 ⫾ 0.33bB (n ⫽ 48) 30.06 ⫾ 1.23cA (n ⫽ 16) 53.94 ⫾ 2.31dB (n ⫽ 18) Ñ 365.282 3, 124 0.000 2004

8 Sept. (Þrst date) 18 Sept. (second date) 12 Oct. (third date) 28 Oct. (fourth date) F df P 21 Aug. (Þrst date) 31 Aug. (second date 20 Sept. (third date) 1 Oct. (fourth date) 28 Oct. (Þfth date) F df P

20.14 ⫾ 0.11aA (n ⫽ 215) 20.4 ⫾ 0.17aA (n ⫽ 72) 32.68 ⫾ 0.13bA (n ⫽ 101) Ñ 2,708.325 2, 385 0.000 17.71 ⫾ 0.42aA (n ⫽ 7) 19.82 ⫾ 0.40aA (n ⫽ 38) 28.81 ⫾ 0.37bA (n ⫽ 146) 45.19 ⫾ 0.89cA (n ⫽ 83) Ñ 311.473 3, 271 0.000

331.4 ⫾ 3.1aA (201.5Ð449.3) 318.6 ⫾ 4.3bA (211.2Ð405.7) 309.6 ⫾ 3.1cA (231.2Ð368.3) Ñ 22.421 2, 384 0.000 343.3 ⫾ 3.8aA (287.7Ð386.3) 326.9 ⫾ 6.1aA (224.4Ð408.1) 301.1 ⫾ 2.4bA (222.8Ð377.2) 296.2 ⫾ 3.5bA (185.3Ð376.5) Ñ 12.524 3, 271 0.000

Diapausing Nondiapausing

Pupal weight (2 d after pupation)

Diapausing Nondiapausing

Duration of larval stage

Dates of larval placement

Diapausing

Weight loss (between 2 and 10 d)


Year

Table 2.

Mean developmental time (days ⴞ SE) of larval stage (diapausing and nondiapausing individuals) and pupal weight (mg ⴞ SE) of H. armigera in field conditions

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Vol. 39, no. 4

rameter (␣) increased with the date of larval placement in the Þeld during the autumn, which indicates that the variability relative to the required degreedays for the completion of larval stage is higher at the early dates of larval placement in relation to the following dates (Table 3). Adult Emergence (From Nondiapausing and Diapausing Pupae) Under Field Conditions. The date of larval placement in the Þeld had no signiÞcant effect (t ⫽ 0.115; df ⫽ 103; P ⫽ 0.909) on pupal developmental times of nondiapausing pupae of H. armigera formed from larvae that had been placed in Þeld conditions on the Þrst and second dates of 2004. No adult eclosion was observed from the nondiapausing pupae of the third date (Table 4). On the contrary, in 2005, the date of larval placement in the Þeld had a significant effect on the pupal developmental time of nondiapausing pupae, which ranged from 13.86 to 32.64 d for the Þrst and third date, respectively (F ⫽ 64.848; df ⫽ 3,108; P ⬍ 0.001; Table 4). The date of larval placement in the Þeld signiÞcantly affected the mean number of days required from pupation until diapause termination (eyespots movement) in both 2005 (F ⫽ 1,033.083; df ⫽ 2,103; P ⬍ 0.001) and 2006 (F ⫽ 345.429; df ⫽ 2,148; P ⬍ 0.001). The diapause development (phase I and phase II) of overwintering pupae of H. armigera ranged from 174.82 to 211.61 d for the third and Þrst date of placement in 2005 and from 175.13 to 224.50 d for the fourth and second date in 2006 (Table 4). The eyespots movement from the diapausing pupae occurred between mid- and late April in 2005 and between late April and early May in 2006. However, it is not yet possible to predict with accuracy the end of phase I of diapause development in Þeld conditions. It is therefore assumed that this phase is completed in northern Greece by early spring when the requirement of exposing the pupae to cold has been met (Fig. 5). The adult emergence from the postdiapause pupae that had been embedded into the soil in the artiÞcial chambers occurred between early and late May in 2005 and between mid-May and early June in 2006 (Fig. 4). This difference in the time of adult eclosion between the 2 yr of experiments is probably attributed to the variation of soil temperature and to the date of larval placement in the Þeld. SpeciÞcally, the mean soil temperature, as recorded in the different depths of soil (20, 10, and 5 cm) and inside the artiÞcial chamber, was ⬇21⬚C during the Þrst 10 d and 24⬚C during the second 10 d of May 2005 and 18 and 21⬚C in the same periods in 2006 (Fig. 5). The date of larval placement signiÞcantly affected the mean number of days required from pupation until adult emergence both in 2005 (F ⫽ 1,390.280; df ⫽ 2,103; P ⬍ 0.001) and in 2006 (F ⫽ 490.603; df ⫽ 2,148; P ⬍ 0.001; Table 4). On all dates of larval placement in the Þeld, female eclosion preceded slightly that of males. Approximately 50% of females eclosed from diapausing pupae before 14 May 2005 and before 25 May 2006, whereas 50% of the males eclosed before 17 May 2005 and before 29 May 2006. By 26 May 2005 and 5 June 2006, all moths of both sexes had eclosed (Fig. 4).

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Fig. 3. Observed and Þtted values of cumulative pupation (%) of H. armigera in the Þeld in a degree-days time scale for each larval placement in the Þeld conditions in 2004 (A), in 2005 (B), and total cumulative pupation (%) in both years of experiments (C). Degree-days were accumulated using 10.5⬚C as the LTT of larval stage and the air temperature.

The mean number of days for the completion of pupal stage of diapausing males was signiÞcantly higher than that of females for all dates of larval placement in the Þeld in 2004 (Þrst date [t ⫽ 4.333; df ⫽ 48; P ⬍ 0.001], second date [t ⫽ 3.904; df ⫽ 22; P ⫽ 0.0013], third date [t ⫽ 5.008; df ⫽ 30; P ⬍ 0.001]), whereas in 2005, signiÞcant differences were observed only for the third date of larval placement (t ⫽ 3.392; df ⫽ 94; P ⬍ 0.001; Table 4). The date of larval placement in both years of experiments did not affect the survival rate of diapausing males and females (Table 4). On the contrary, there was signiÞcant interaction between the survival rate of each insect sex and the year of experiment (么 [␹2 ⫽ 6.518; df ⫽ 1; P ⫽ 0.011], 乆 [␹2 ⫽ 4.709; df ⫽ 1; P ⫽ 0.030]), whereas, between the insect sexes, the survival was independent of the placement

date in autumn of 2004 (Þrst date [␹2 ⫽ 0.160; df ⫽ 1; P ⫽ 0.690], second [␹2 ⫽ 1.717; df ⫽ 1; P ⫽ 0.190], third [␹2 ⫽ 0.007; df ⫽ 1; P ⫽ 0.931]) and in autumn of 2005 (second date [␹2 ⫽ 0.879; df ⫽ 1; P ⫽ 0.348], third [␹2 ⫽ 0.331; df ⫽ 1; P ⫽ 0.565], fourth [␹2 ⫽ 0.013; df ⫽ 1; P ⫽ 0.910]; Table 4). Daily maximum and minimum soil temperatures at depths of 20, 10, and 5 cm and inside the artiÞcial chamber were recorded in both years of experiments, and they are depicted in Fig. 5. The adult emergence of the total overwintering population against degreedays, which was calculated using the soil temperatures at the different depths as well as using the mean soil temperature, was well described by the Weibull function (r2 ⫽ 0.991Ð 0.995) for each year of experiment (Fig. 6; Table 5). In both years, the start date for the

13.86 ⫾ 0.63a (n ⫽ 42) 21.31 ⫾ 0.52b (n ⫽ 45) 32.64 ⫾ 2.59c (n ⫽ 11) 30.01 ⫾ 1.94c (n ⫽ 14)

No adults emerged from nondiapausing pupae on third larval placement date in 2004. Values in parentheses are ranges. Means within a column followed by the same letter (lowercase) are not signiÞcantly different (P ⬍ 0.05. Tukey HSD test). Means between sexes in a particular date followed by the same letter (uppercase) are not signiÞcantly different (P ⬍ 0.05, t-test and ␹2-test for percentages). ⌵, no. of pupae; n, no. of adults that were emerged from pupae.

50.00 61.54〈 71.01〈 80.00〈

53.49〈 46.43〈 32.61〈

46.55〈 25.00〈 31.48〈 2 ␹ ⫽ 1.625 2 0.444 Ñ 33.33〈 61.04〈 76.00〈 2 ␹ ⫽ 0.327 2 0.849 225.87 ⫾ 0.48aB (n ⫽ 23) 211.61 ⫾ 0.62a (200Ð219) 216.38 ⫾ 1.38bB (n ⫽ 13) 203.83 ⫾ 1.11b (189Ð212) 186.33 ⫾ 0.57cB (n ⫽ 15) 174.82 ⫾ 0.39c (165Ð219) 1,033.083 1,033.083 2, 103 2, 103 0.000 0.000 252 (n ⫽ 1) 235.00 240.88 ⫾ 1.10aA (n ⫽ 16) 224.50 ⫾ 0.72a (213Ð232) 218.08 ⫾ 0.90bB (n ⫽ 49) 202.33 ⫾ 1.09b (185Ð210) 186.69 ⫾ 2.29cA (n ⫽ 16) 175.13 ⫾ 1.18c (166Ð206) 345.429 345.429 2, 148 2, 148 0.000 0.000 228.74 ⫾ 0.45aA (n ⫽ 27) 222.18 ⫾ 0.54bA (n ⫽ 11) 190.06 ⫾ 0.49cA (n ⫽ 17) 720.189 2, 48 0.000 Ñ 244.00 ⫾ 1.78aA (n ⫽ 4) 221.60 ⫾ 0.49bA (n ⫽ 47) 192.47 ⫾ 1.84cA (n ⫽ 19) 269.678 2, 78 0.000 101 72 100 1,390.280 2, 103 0.000 7 38 146 45 490.603 2, 148 0.000

227.42 ⫾ 0.39a (221Ð234) 219.04 ⫾ 0.98b (210Ð225) 188.31 ⫾ 0.49c (183Ð193) 1,759.770 2, 52 0.000 252.00 241.50 ⫾ 0.97a (249Ð235) 219.80 ⫾ 0.55b (235Ð203) 189.83 ⫾ 1.51c (206Ð171) 293.454 2, 67 0.000

乆乆么么 Total 乆乆么么 Total

24.13 ⫾ 0.36a (n ⫽ 46) 24.07 ⫾ 0.39a (n ⫽ 59) Ñ

8 Sept. (Þrst date) 115 18 Sept. (second date) 167 12 Oct. (third date) 60 F df P 2005 21 Aug. (Þrst date) 46 31 Aug. (second date) 48 20 Sept. (third date) 16 1 Oct. (fourth date) 18 F 64.848 df 3, 108 P 0.000

The comparison of the patterns of the diapause development and emergence of the overwintering insects with the Þeld activity of adult moths is important to determine the role of local populations in the population dynamics of Heliothis species in a speciÞc area

2004

Discussion

N

model was the 1 April because, before this date, mean soil temperatures were below the lower threshold for pupal diapause development (15.9⬚C; Fig. 5). This LTT was used until 30 April 2005 and 10 May 2006 because, by those dates, 100% of the overwintering pupae terminated diapause, whereas 10.2⬚C was used as LTT for postdiapause development as mentioned above. The overall accumulated percentage of adult emergence in both years against degree-days was well described by the Weibull function (r2 ⫽ 0.925), which accounted for 92.5% of year to year variation in the emergence pattern (Fig. 6C; Table 5). The model also indicated that 10, 25, 50, 75, and 90% of cumulative emergence of adults of H. armigera in northern Greece occurs at 153, 199, 252, 303, and 347 DD after 1 April, respectively (Fig. 6C). Sex Pheromone Trapping. Captures of H. armigera male moths in pheromone traps appeared initially on 6 May 2005 and 13 May 2006, which are relatively close to the adult emergence of the overwintering population that had been placed in the artiÞcial chambers into the soil. Female moths started to eclose in the Þeld on 7 May 2005 and 17 May 2006, whereas male moths started to eclose on 9 May 2005 and 21 May 2006. In both years, adult emergence from the overwintering population preceded peak trap catches, but the difference was greater in 2006 (Fig. 7). The trap catches of the overwintering generation (Þrst ßight) of H. armigera lasted for ⬃1.5 mo and was completed on 17 and 24 June 2005 and 2006, respectively, ⬇2 wk after the completion of adultsÕ emergence from the artiÞcial chambers (Figs. 4 and 7). In this Þrst ßight, low levels of male moth captures (maximum ⬇11 males/trap) were observed, which was followed by three peaks in 2005 and two peaks in 2006, which suggests the occurrence of three or four generations per year of this insect in northern Greece. The end of H. armigera ßight in this region was observed in mid-October, because no male moths were captured in traps after this date (Fig. 7).

Total

0.978 0.986 0.939 0.968 0.984 0.993 0.975 0.922 0.969 0.968

N

221.752 ⫾ 1.406 206.478 ⫾ 0.820 160.831 ⫾ 0.503 196.354 ⫾ 0.909 239.663 ⫾ 1.846 302.376 ⫾ 1.186 219.389 ⫾ 1.470 178.648 ⫾ 0.283 235.019 ⫾ 1.196 218.448 ⫾ 1.073

Diapause termination

15.221 ⫾ 2.122 29.162 ⫾ 3.747 44.294 ⫾ 7.175 29.559 ⫾ 4.348 9.221 ⫾ 0.842 9.449 ⫾ 0.469 10.200 ⫾ 0.860 39.109 ⫾ 3.413 16.995 ⫾ 1.396 22.379 ⫾ 2.661

r2

Diapausing

␤

Duration of pupal stage

8 Sept. (Þrst date) 18 Sept. (second date) 12 Oct. (third date) Total 2005 21 Aug. (Þrst date) 31 Aug. (second date) 20 Sept. (third date) 1 Oct. (fourth date) Total Total

␣

Nondiapausing

2004

Parameter ⫾ SE



Year

Year

Table 3. Estimated value of parameters in Weibull distribution model 关f(t)兴 of H. armigera larval stage in field conditions

Vol. 39, no. 4

Survivorship (%) diapausing

ENVIRONMENTAL ENTOMOLOGY Table 4. Mean developmental time (days ⴞ SE) of pupal stage (diapausing and nondiapausing individuals), no. of days ⴞ SE for diapause termination, and survivorship (%) of H. armigera diapausing pupae in field conditions

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Fig. 4. Adult spring emergence pattern of diapausing male (F) and female (E) pupae that had been placed into the soil in the artiÞcial chambers in autumn of 2004 (A) and of 2005 (B). For the Þrst larval placement (21 August) in 2005, no adult emergence was observed.

(Lopez et al. 1984, Rummel et al. 1986). The survival and the overwintering patterns of H. armigera under Þeld conditions in northern Greece has not been determined, even though the insectÕs presence and the damage it causes have been recorded for many years (Isaakides 1941, Pelekassis 1962). Furthermore, there were no data on the insectÕs diapause incidence and termination under natural outdoor conditions in northern Greece, although temperate zone populations of H. armigera exhibit diapause ability both in Þeld and laboratory conditions (Qureshi et al. 1999, 2000; Zhou et al. 2000a, b; Shimizu and Fujisaki 2002; Ge et al. 2005; Shimizu et al. 2006). Similarly, it has not been clear whether local populations of H. armigera were established in Greece. It is probable that populations in Greece re-establish each year by immigration from the subtropics or the tropics because this species has outstanding ßight ability (Pedgley 1985;

Pedgley et al. 1987; Gregg and Wilson 1991; Coombs et al. 1993; Gregg 1993; Gregg et al. 1995; Casimero et al. 1999; Feng et al. 2004, 2005). The aim of our research was to clarify the above subjects by placing H. armigera larvae in outdoor conditions at different time intervals and assessing their phenological responses. Our study provides evidence that H. armigera overwinters as diapausing pupae in northern Greece, at least a part of the late-season generation. This strongly suggests that a local population exists in this area as opposed to the speculation that the annual recolonization happens entirely from immigrant moths. Pupae enter diapause during September and October in response to declining temperatures (from ⬇24 to 15⬚C) and photoperiod (from ⬇13 to 10 h light) in this area (Fig. 1). In this study, we found that the part of the population that did not enter pupal diapause during autumn either died as larvae or pupae or a small

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Fig. 5. Daily maximum and minimum soil temperatures at depths of 20, 10, and 5 cm and inside the artiÞcial chamber. Horizontal line was drawn through 15.9⬚C as the LTT of diapausing pupal stage (phase II).

proportion of it eclosed in the winter (Tables 1 and 4), providing evidence that this part of the population is maladapted for overwintering survival. It seems that local nondiapausing individuals are unable to reproduce during the winter. During the second half of the scotophase, when H. armigera females release sex pheromone (Rafaeli and Soroker 1989), the average temperature in the winter in northern Greece is ⬇10⬚C. Under such conditions, only very low levels of copulation, oogenesis, and egg hatch occur (Zhou et al. 2000a, b,; Mironidis and Savopoulou-Soultani 2008). In relation to the total number of H. armigera larvae that completed the larval stage and pupated, the percentage of pupae that entered diapause ranged from 65 to 30% (mean: 52.7%) in 2004 and from 13 to 90% (mean: 57.4%) in 2005 and presented signiÞcant correlation with the date of larval placement in the Þeld and with the year of the experiment but not with the

insect sex (Table 1). The fact that only a portion of the population undergoes diapause strongly suggests that genetic variation exists in H. armigera population in northern Greece. However, the variation in diapause response both within and between years also seems to be in direct relation to variation in temperature and photoperiod, both of which are known to inßuence diapause induction. In 2005, a progressive increase was observed in the percentage of diapausing pupae when the larval placement in the Þeld took place later in autumn (Table 1; Figs. 3 and 4).This fact seems to be typical of H. armigera diapause in temperate regions, because it was also observed in other temperate regions such as in central (Jiang et al. 1999) and northern China (Ge et al. 2005), in Japan (Shimizu et al. 2006), in Australia (Wilson et al. 1979, Kay 1982, Murray 1991), and in Israel (Zhou et al. 2000a). The different proportion of the population that entered diapause at

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Fig. 6. Observed and Þtted values of total cumulative (%) adult emergence of H. armigera in the Þeld in a degree-days time scale in spring of 2005 (A), of 2006 (B), and in both years of experiments (C). The required degree-days for the adult emergence from the overwintering pupae were calculated using 15.9⬚C as the LTT of diapausing pupal stage (phase II) and 10.2⬚C as the LTT of postdiapausing pupal stage and the soil temperatures (depths of 20, 10, and 5 cm and inside the artiÞcial chamber) as well as the average soil temperature.

each placement explains the variation that was observed in the required degree-days for the completion of larval stage in relation to the dates of larval placement (Fig. 3), probably because diapausing and nonTable 5. Estimated value of parameters in Weibull distribution model 关f(t)兴 of H. armigera adult emergence in field conditions

Year 2005

2006

Total

Depth of temperature recorded 20 cm 10 cm 5 cm Chamber interior Average 20 cm 10 cm 5 cm Chamber interior Average

Parameter ⫾ SE

␣

␤

3.406 ⫾ 0.113 3.636 ⫾ 0.128 5.013 ⫾ 0.212 4.253 ⫾ 0.170 4.077 ⫾ 0.156 3.140 ⫾ 0.115 3.655 ⫾ 0.136 3.374 ⫾ 0.125 3.560 ⫾ 0.133 3.432 ⫾ 0.127 3.755 ⫾ 0.141

196.616 ⫾ 1.503 224.201 ⫾ 1.716 328.550 ⫾ 2.222 288.550 ⫾ 2.161 259.479 ⫾ 1.901 256.241 ⫾ 2.203 324.288 ⫾ 2.432 287.900 ⫾ 2.324 315.267 ⫾ 2.434 295.924 ⫾ 2.348 277.701 ⫾ 2.125

r2 0.995 0.994 0.992 0.992 0.993 0.991 0.995 0.994 0.992 0.993 0.925

diapausing individuals exhibit different thermal characteristics or that the thermal threshold could be changing as diapause progresses (Tauber and Tauber 1976). Additionally, in our results, sex ratio differs markedly from 1:1 in some dates of placement, which indicates that the population is stressed and suggests that the conditions were not optimal for these larvae (Table 1). Son et al. (2007) claimed that the dynamic nature of Þeld temperatures and the microclimate are two factors that should be taken into account to understand the thermo-biology of some insect species under Þeld conditions. In the temperate zone, autumn is characterized by unpredictably decreasing temperatures and predictably shortening daylengths (Fig. 1). Under these conditions, H. armigera is subjected to high larval mortality, as shown in our experiments (Table 1). The decrease in temperature during autumn caused a part of the population to develop too late, because of degree-day limitation, to reach the pupal stage (Table

1080


Fig. 7. Seasonal phenology of adult H. armigera in northern Greece. Adult emergence of the pupae that had been placed into the soil in the artiÞcial chambers in autumn of 2004 (A), and of 2005 (C), and captures of H. armigera male moths in pheromone traps in 2005 (B) and 2006 (D). The number of adults in the graphs plotted (A and C) are derived from 273, in 2004, and 236, in 2005, Þeld-exposed laboratory pupae (note different scales).

2). SpeciÞcally, H. armigera larvae that were placed in Þeld conditions after mid-October did not pupate and died because this life stage is not adapted for overwintering (Tables 1 and 2). Furthermore, the continuous exposure to extreme low temperatures prevailing during winter months (Fig. 5) has a negative effect on the survival of different life stages of H. armigera. The results from our study show that extreme low temperatures apart from larvae are highly detrimental to nondiapausing overwintering pupae and to a variable portion of those that successfully undergo diapause (Table 4).

Vol. 39, no. 4

The pupation pattern of H. armigera in our experiments was characterized by the presence of diapausing and nondiapausing pupae (Figs. 2 and 4). The mean developmental time of larval stage, the pupal weight, and the mean developmental time of pupal stage of nondiapausing individuals were related to the date of the Þrst instar larvae placement in the Þeld (Tables 2 and 4). The duration of larval stage in both years does not seem to differ dramatically, even though at certain placements, differences between diapausing and nondiapausing individuals are statistically, but not dramatically, signiÞcant. The most striking results are the differences in weight loss: in nondiapausing pupae, the degree of energy expended is high during the transition to adult stage, whereas in diapausing pupae, all metabolic activity is held to a minimum (Table 2). The presence of those nondiapausing pupae during autumn looks similar both to the Þfth “suicidal” generation of H. armigera in China (Ge et al. 2005) and to the appearance of local nondiapausing H. armigera individuals in Israel, which were incapable of reproducing during the winter because of low temperatures (Zhou et al. 2000a). In contrast, the diapausing pupae of H. armigera undergo a process of cold acclimation during which their trehalose content as a cryoprotectant increases (Izumi et al. 2005). If pupae are produced too late in autumn, there may not be enough time for this cold acclimation to occur. In corresponding experiments, Shimizu et al. (2006) found that the local population of H. armigera in Japan requires relatively low temperatures (⬍20⬚C) for diapause induction because the photoperiodic responses in diapause induction of H. armigera in the speciÞc region are observed only at temperatures around 20⬚C (Qureshi et al. 1999, 2000), whereas temperatures ⬍13.6⬚C, the lowest temperature threshold for Japan population (Qureshi et al. 1999), can affect the survival before pupation. Similarly, in northern Greece the mean daily air temperature during September and October ranges between 15 and 20⬚C and decreases sharply below 15⬚C in early November (Fig. 1). In this period (SeptemberÐOctober) the only suitable host and probably the main source of the overwintering population of H. armigera in this region are cotton crops, which allow larvae of the last generation to complete their larval development and to pupate into the soil (unpublished data). The applications of defoliants to facilitate the mechanical harvesting of cotton start in mid-October in northern Greece, whereas by this time other host crops such as maize have already completed their life cycle and they are not suitable hosts for the insect any more. In the following spring, adult emergence from diapausing pupae that had been placed into the soil inside the artiÞcial chambers lasted ⬃3 wk in both years of the experiments (Figs. 4 and 7), and it is well described by the Weibull model, which accounts for 92.5% of year to year variation in the emergence pattern (Table 5; Fig. 6). One consequence of diapause is that the overwintering pupae attain a uniform physiological state before morphogenesis resumes. Consequently, the emergence of pupae formed over a long period

August 2010


becomes concentrated, and in general, the date of emergence does not relate to the date of pupation (Wilson et al. 1979, Lopez et al. 1984). Fitt (1989) claimed that, in all species of the genus Heliothis, postdiapause adult emergence occurs over a period of 3Ð 6 wk, with females emerging slightly earlier than males. In our results for all the dates of larval placement in the Þeld, the females were observed to eclose Þrst because the mean pupal developmental time was shorter in females than in males for all dates of placement (Table 4). Two factors probably contribute to this effect. First, there are intrinsic differences in developmental rates between the sexes, with female pupae developing faster than male pupae (Mironidis and Savopoulou-Soultani 2008). Second, female pupae are not found as deep in the ground as male pupae are (Murray and Zalucki 1994), and therefore, they are exposed to higher temperatures, resulting in faster development. Higher daily average temperatures usually occur closer to the soil surface because of the soil thermocline. Murray (1991) reported that, for every 1 mm increase in pupal depth, emergence of H. armigera from diapause is delayed by 0.78 d. Emergence patterns of naturally diapausing pupal populations have been described for all species of Heliothis (Pearson 1958, Roome 1979, Wilson et al. 1979, Kay 1982, Lopez et al. 1984), and simple models for predicting the emergence of adults in spring are available for H. armigera (Wilson et al. 1979, Cunningham et al. 1981, DufÞeld and Dillon 2005). In the Mediterranean region, it has been observed that the postdiapause adult emergence of H. armigera begins in late April, and 50% of females eclose before 6 May, which is 9 d before 50% of the males do (Zhou et al. 2000a). In central Japan, most of the overwintering pupae of H. armigera complete their diapause development, and the adults emerge in early June (Qureshi et al. 1999). In our results, 50% of females eclosed from diapausing pupae before 14 May 2005 and 25 May 2006, whereas 50% of males eclosed before 17 May 2005 and 29 May 2006 (Fig. 4). In southern New South Wales of Australia, DufÞeld and Dillon (2005) reported that the spring emergence of H. armigera from overwintering pupae occurs over a period of ⬇4 wk, from midOctober to mid-November, depending on local climatic conditions, whereas no moths emerged before October. Additionally, Kay (1982) described a consistently bimodal emergence pattern of H. armigera in southeastern Queensland (27⬚35⬘ S) of Australia; some adults emerged in late September and October (early spring), and a second, larger emergence occurred in late November and early December. Wilson et al. (1979) attributed this early peak to the emergence of nondiapausing individuals. Whatever the explanation may be, the extended period of emergence of most species ensures that the entire overwintering population does not encounter unfavorable conditions for reproduction, such as unseasonal weather or lack of synchrony with spring hosts. Lopez et al. (1984) claimed that synchrony between the patterns of spring emergence from overwintering population of Heliothis and their capture in phero-

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mone traps, under similar environmental conditions in the Þeld, showed the feasibility of using pheromone traps to monitor overwintering emergence. Furthermore, the same authors noted that asynchrony between the capture and emergence indicated that movement of the species into the area may occur before emergence from overwintering. In our results, the synchrony between the captures of H. armigera in the pheromone traps and the adult eclosion of the overwintering population that had been placed inside the artiÞcial chambers into the soil is correlative. In both years, the emergence of overwintered adults precedes pheromone trap catches (Fig. 7). Thus, pheromone trap catches may represent adults from the overwintering population, immigrants, or both. The increased number of captures that was observed in the pheromone traps afterward, during July and August, probably can be attributed to a numerical increase in the local and early spring immigrant populations, as well as to new immigrants that come into the region with migratory ßight. If immigration into northern Greece occurs, the origin of migrating moths is unclear. We hypothesize that these moths are immigrants from warmer, more southern regions. It is known that the ability for long distance movements of seasonal migration from low to higher latitudes in summer, usually on warm winds preceding cold fronts, is a characteristic of H. armigera (Fitt 1989). In Cyprus, catches of H. armigera adult moths in a light trap net during winter months are evidence that this species undergoes winter migration in the Middle East with the aid of southern winds (Pedgley 1986). The same author showed that this species migrates up to 1,000 km to reach Britain and other parts of Europe from sources in southern Europe and North Africa (Pedgley 1985). Furthermore, captures of H. armigera male moths in pheromone traps in Israel, before the appearance of the local overwintering diapausing population, are evidence of the spring migration in this area from regions with lower latitude (Zhou et al. 2000a). Recently, the comparison of genetic structure of populations of H. armigera from Turkey, Israel, Ethiopia, and Egypt showed low levels of genetic differentiation and consequently a high degree of gene ßow between those populations (Zhou et al. 2000b). This fact supports the claims for the existence of migratory ßights of this insect in these regions (Zhou et al. 2000a). However, experimental data related to migratory behavior of this species are not available for the regions of northern Greece. Timing of diapause termination and of postdiapause spring emergence gain additional signiÞcance for those insects whose life histories require close synchrony with phenological development of a particular host plant complex (Logan et al. 1979). This synchronization assumes anthropocentric implications when the preferred or accepted host plant happens to be an agricultural crop. Such is the case of Heliothis spp., whose impact as a pest on several crops is dependent on synchrony of ovipositional life stages with attractive host plant stages (Johnson et al. 1975). In the area of northern Greece, the seeding of corn, which seems

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to be the primary host crop allowing the early development of H. armigera populations (Mironidis 2009), begins in mid-April, and the corn crop forms silk approximately in middle June, whereas the seeding of cotton begins in late April. Consequently, adult eclosion from the overwintering generation of H. armigera takes place in a period when a suitable host crop that can support the increase of the local population is available. In summary, our data supply strong evidence that a local population of H. armigera exists in northern Greece. In this area, H. armigera enters pupal diapause in autumn, at least a part of the late-season generation, probably in cotton crop residues. Diapause termination and eclosion of the adults in the following spring are determined by the local soil temperature conditions, with the adult emergence being concentrated in a period of 4 Ð 6 wk between late April and early June. The correlation between postdiapause adult emergence and trap captures indicate that spring population of this species may also include immigrants. Thus, our study improves the understanding of dormancy and phenology of H. armigera in northern Greece relative to what is known in other parts of its geographic range. References Cited Baskerville, G. L., and P. Emin. 1969. Rapid estimation of heat accumulation from maximum and minimum temperatures. Ecology 50: 515Ð517. Broufas, G. D., and D. S. Koveos. 2000. Threshold temperature for post-diapause development and degree-days to hatching of winter eggs of the European red mite (Acari: Tetranychidae) in northen Greece. Environ. Entomol. 29: 710Ð713. Casimero, V., R. Tsukuda, F. Nakasuji, and K. Fujisaki. 1999. The pre-calling period and starting time of calling by females of three Japanese populations of the cotton bollworm, Helicoverpa armigera (Hu¨ bner) (Lepidoptera: Noctuidae). Appl. Entomol. Zool. 34: 123Ð127. Coaker, T. H. 1959. Investigation on Heliothis armigera in Uganda. Bull. Entomol. Res. 50: 487Ð506. Coombs, M., A. P. Del Socorro, G. P. Fitt, and P. G. Gregg. 1993. The reproductive maturity and mating status of Helicoverpa armigera, H. punctigera and Mythimna convecta (Lepidoptera: Noctuidae) collected in towermounted light traps in northern New South Wales, Australia. Bull. Entomol. Res. 83: 529 Ð534. Cunningham, R. B., T. Lewis, and A.G.L. Wilson. 1981. Biothermal development: a model for predicting the distribution of emergence times of diapausing Heliothis armigera. Appl. Stat. 30: 132Ð140. Danks, H. V. 1987. Insect dormancy: an ecological perspective. Biological Survey of Canada, Ottawa, Canada. Drake, V. A., and A. G. Gatehouse. 1995. Insect migration: tracking resources through space and time. Cambridge University Press, Cambridge, United Kingdom. Duffield, S. J. 2004. Evaluation of the risk of overwintering Helicoverpa spp. pupae under irrigated summer crops in south-eastern Australia and the potential for area-wide management. Ann. Appl. Biol. 144: 17Ð26. Duffield, S. J., and M. L. Dillon. 2005. The emergence and control of overwintering Helicoverpa armigera pupae in southern New South Wales. Aust. J. Entomol. 44: 316 Ð320.

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[EPPO/CABI] European and Mediterranean Plant Protection Organization/CAB International. 1997. Helicoverpa armigera, pp. 289 Ð294. In I. M. Smith, D. G. McNamara, P.R. Scott, and M. Holderness (eds.), Quarantine Pests for Europe, 2nd ed. CAB International, Wallingford, United Kingdom. Feng, H. Q., K. M. Wu, Y. X. Ni, D. F. Cheng, and Y. Y. Guo. 2004. Northward migration of Helicoverpa armigera (Lepidoptera: Noctuidae) and other moths in early summer observed with radar in northern China. J. Econ. Entomol. 97: 1874 Ð1883. Feng, H. Q., K. M. Wu, Y. X. Ni, D. F. Cheng, and Y. Y. Guo. 2005. Return migration of Helicoverpa armigera (Lepidoptera: Noctuidae) during autumn in northern China. Bull. Entomol. Res. 95: 361Ð370. Fitt, G. P. 1989. The ecology of Heliothis in relation to agroecosystems. Annu. Rev. Entomol. 34: 17Ð52. Fitt, G. P., and J. C. Daly. 1990. Abundance of overwintering pupae and the spring generation of Helicoverpa spp. (Lepidoptera: Noctuidae) in northern New South Wales, Australia: Implication for pest management. J. Econ. Entomol. 83: 1827Ð1836. Foley, D. H. 1981. Pupal development rate of Heliothis armigera (Hu¨ bner) (Lepidoptera: Noctuidae) under constant and ßuctuating temperatures. J. Aust. Ent. Soc. 20: 13Ð20. Ge, F., F. Chen, M. N. Paralulee, and E. N. Yardim. 2005. QuantiÞcation of diapausing fourth generation and suicidal Þfth generation cotton bollworm, Helicoverpa armigera, in cotton and corn in northern China. Entomol. Exp. Appl. 116: 1Ð7. Gregg, P. C. 1993. Pollen as a marker for migration of Helicoverpa armigera and H. punctigera (Lepidoptera: Noctuidae) from Western Queensland. Aust. J. Ecol. 18: 209 Ð 219. Gregg, P. C., and A.G.L. Wilson. 1991. Trapping methods for adults, pp. 30 Ð 48. In M. P. Zalucki (ed.), Heliothis: research methods and prospects. Springer-Verlag, New York. Gregg, P. C., G. P. Fitt, M. P. Zalucki, and D.A.H. Murray. 1995. Insect migration in an arid continent. II Helicoverpa spp. in eastern Australia, pp. 151Ð172. In V. A. Drake and A. G. Gatehouse (eds.), Insect migration: tracking resources through space and time. Cambridge University Press, Cambridge, United Kingdom. Hackett, D., and A. G. Gatehouse. 1982. Diapause in Heliothis armigera (Hu¨ bner) and H. fletcheri (Hardwick) (Lepidoptera: Noctuidae) in the Sudan Gezira. Bull. Entomol. Res. 72: 409 Ð 422. Holtzer, T. O., J. R. Bradley, and R. L. Rabb. 1976. Effects of various temperature regimes on the time required for emergence of diapausing Heliothis zea. Ann. Entomol. Soc. Am. 69: 257Ð260. Isaakides, C. A. 1941. Insects interesting the Greek agriculture with some observations on them (in Greek). Proc. Acad. Athens 16: 238 Ð263. Izumi, Y., K. Anniwaer, H. Yoshida, S. Sonoda, K. Fujisaki, and H. Tsumuki. 2005. Comparison of cold hardiness and sugar content between diapausing and nondiapausing pupae of the cotton bollworm, Helicoverpa armigera (Lepidoptera: Noctuidae). Physiol. Entomol. 30: 36 Ð 41. Jiang, M., L. Xie, and X. Zhang. 1999. Characteristics of diapause induction of cotton bollworm. Chinese J. Appl. Ecol. 10: 60 Ð 62. Johnson, M. W., R. E. Stinner, and R. L. Rabb. 1975. Ovipositional response of Heliothis zea (Boddie) to its major hosts in North Carolina. Environ. Entomol. 4: 291Ð297.

August 2010


Kay, I. R. 1982. The incidence and duration of pupal diapause in Heliothis armigera (Hu¨ bner) (Lepideptera: Noctuidae) in southeast Queensland. J. Aust. Entomol. Soc. 21: 263Ð266. Kehat, M., and E. Dunkelblum. 1990. Behavioral analysis of male Heliothis armigera (Lepidoptera: Noctuidae) moths in a ßight tunnel to combinations of components identiÞed from female sex pheromone glands. J. Insect Behav. 3: 75Ð 83. Kehat, M., and E. Dunkelblum. 1993. Sex pheromones: achievements in monitoring and mating disruption of cotton pests in Israel. Arch. Ins. Biochem. Physiol. 22: 425Ð 431. Leather, S. R., K.F.A. Walters, and J. S. Bale. 1993. The ecology of insect overwintering. Cambridge University Press, New York. Lee, E. R. 1991. Principles of insect low temperature tolerance, pp. 17Ð 46. In E. R. Lee and D. L. Denlinger (eds.), Insects at low temperature. Chapman & Hall, New York and London. Logan, J. A., R. E. Stinner, R. L. Rabb, and J. S. Bacheler. 1979. A descriptive model for predicting spring emergence of Heliothis zea population in North Carolina. Environ. Entomol. 8: 141Ð146. Lopez, J. D. 1986. Thermal requirements for diapause termination in laboratory cultures of Heliothis zea and H. virescens (Lepidoptera: Noctuidae). Environ. Entomol. 15: 919 Ð923. Lopez, J. D., A. W. Hartstack, and R. Beach. 1984. Comparative pattern of emergence of Heliothis zea and H. virescens (Lepidoptera: Noctuidae) from overwintering pupae. J. Econ. Entomol. 77: 1421Ð1426. McCann, I. R., J. D. Lopez, Jr., and J. A. Witz. 1989. Emergence model for Þeld populations of overwintering Heliothis zea and H. virescens (Lepidoptera: Noctuidae). Environ. Entomol. 18: 618 Ð 624. Milonas, P. G., M. Savopoulou-Soultani, and D. G. Stavridis. 2001. Day-degree models for predicting the generation time and ßight activity of local populations of Lobesia botrana (Den. & Schiff) (Lep., Torticidae) in Greece. J. Appl. Entomol. 125: 515Ð518. Mironidis, G. K. 2009. Study of bio-ecology of Helicoverpa armigera (Lepidoptera: Noctuidae) and its parasitoid Hyposoter didymator (Hymenoptera: Ichneumonidae) in northern Greece. PhD thesis, Aristotle University of Thessaloniki, Thessaloniki, Greece. Mironidis, G. K., and M. Savopoulou-Soultani. 2008. Development, survivorship, and reproduction of Helicoverpa armigera (Lepidoptera: Noctuidae) under constant and alternating temperatures. Environ. Entomol. 37: 16 Ð28. Mourikis, P. A., and P. Vasilaina-Alexopoulou. 1969. Notes on the rearing on the synthetic diet of the cotton bollworm Heliothis armigera. Annls. Inst. Phytopath. Benaki. 9: 46 Ð53. Murray, D.A.H. 1991. Investigations into the development and survival of Heliothis spp. pupae in South-East Queensland. PhD thesis, University of Queensland, Brisbane, Australia. Murray, D.A.H., and A.G.L. Wilson. 1991. Methods for studying diapause, pp. 102Ð108. In M. P. Zalucki (ed.), Heliothis: Research Methods and Prospects. SpringerVerlag, New York, NY. Murray, D.A.H., and M. P. Zalucki. 1994. Spatial distribution and mortality of Helicoverpa spp. pupae (Lepidoptera: Noctuidae) under Þeld crops on the Darling Downs Queensland. J. Aust. Entomol. Soc. 33: 193Ð198.

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Pearson, E. O. 1958. The insect pests of cotton in tropical Africa. Commonwealth Institute of Entomology, London, United Kingdom. Pedgley, D. E. 1985. Windborne migration of Heliothis armigera (Hu¨ bner) (Lepidoptera: Noctuidae) to the British Isles. Entomol. Gaz. 36: 15Ð20. Pedgley, D. E. 1986. Windborne migration in the Middle East by the moth Heliothis armigera (Hu¨ bner) (Lepidoptera: Noctuidae). Ecol. Entomol. 11: 467Ð 470. Pedgley, D. E., M. R. Tucker, and C. S. Pawar. 1987. Windborne migration of Heliothis armigera (Hu¨ bner) (Lepidoptera: Noctuidae) in India. Insect Sci. Applic. 8: 599 Ð 604. Pelekassis, C.E.D. 1962. A catalogue of the more important insects and other animals harmful to the agricultural crops of Greece during the last thirty year period. Ann. Inst. Phytopath. Benaki N. S. 5: 5Ð104. Potter, M. F., R. T. Huber, and T. F. Watson. 1981. Heat units requirements for emergence of overwintering tobacco budworm, Heliothis virescens (F.), in Arizona. Environ. Entomol. 10: 543Ð545. Qureshi, M. H., T. Murai, H. Yoshida, T. Shiraga, and H. Tsumuki. 1999. Effects of photoperiod and temperature on development and diapause induction in the Okayama population of Helicoverpa armigera (Hb.) (Lepideptera: Noctuidae). Appl. Entomol. Zool. 34: 327Ð331. Qureshi, M. H., T. Murai, H. Yoshida, T. Shiraga, and H. Tsumuki. 2000. Population variation in diapause induction and termination of Helicoverpa armigera (Lepideptera: Noctuidae). Appl. Entomol. Zool. 35: 357Ð360. Rafaeli, A., and V. Soroker. 1989. Inßuence of diel rhythm and brain hormone on pheromone production in two lepidopteran species. J. Chem. Ecol. 15: 447Ð 455. Reed, W. 1965. Heliothis armigera (Hb.) (Noctuidae) in Western Tanganyika. I: Biology, with special reference to the pupal stage. Bull. Entomol. Res. 56: 117Ð125. Re´gnie`re, J. 1984. A method of describing and using variability in development rates for the simulation of insect phenology. Can. Entomol. 116: 1367Ð1376. Roach, S. H., and P. L. Adkisson. 1971. Termination of pupal diapause in the bollworm. J. Econ. Entomol. 64: 1057Ð 60. Roome, R. E. 1979. Pupal diapause in Heliothis armigera (Hu¨ bner) (Lepidoptera: Noctuidae) in Botswana: its regulation by environmental factors. Bull. Entomol. Res. 69: 149 Ð160. Rummel, D. R., and J. L. Hatfield. 1988. Thermal-based emergence model for the bollworm (Lepidoptera: Noctuidae) in the Texas High Plains. J. Econ. Entomol. 81: 1620 Ð1623. Rummel, D. R., K. C. Neece, M. D. Arnold, and B. A. Lee. 1986. Overwintering survival and spring emergence of Heliothis zea (Boddie) in the Texas southern High Plains. Southwest. Entomol. 11: 1Ð9. Sevacherian, V., N. C. Toscano, R. A. Van Steenwyk, R. K. Sharma, and R. R. Sanders. 1977. Forecasting pink bollworm emergence by thermal summation. Environ. Entomol. 6: 545Ð546. Shimizu, K., and K. Fujisaki. 2002. Sexual differences in diapause induction of the cotton bollworm, Helicoverpa armigera (Hb.) (Lepidoptera: Noctuidae). Appl. Entomol. Zool. 37: 527Ð533. Shimizu, K., K. Shimizu, and K. Fujisaki. 2006. Timing of diapause induction and overwintering success in the cotton bollworm Helicoverpa armigera (Hb.) (Lepidoptera: Noctuidae) under outdoor condition in temperate Japan. Appl. Entomol. Zool. 41: 151Ð159. Shumakov, E. M., and L. A. Yakhimovich. 1955. Morphological and histological peculiarities of the metamorpho-

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sis of the cotton bollworm (Choridea obsoleta F.) in connection with the phenomenon of diapause. Doklady Akademy Sci. USSR 101: 779 Ð782. Sokal, R. R., and F. J. Rohlf. 1995. Biometry, 3rd ed. Freeman, New York. Sømme, L., and W. Block. 1991. Adaptations to alpine and polar environments in insects and other terrestrial arthropods, pp. 318 Ð359. In E. R. Lee and D. L. Denlinger (eds.), Insects at low temperature. Chapman & Hall, New York and London. Son, Y., J. H. Lee, and Y. J. Chung. 2007. Temperaturedependent post-diapause development and prediction of spring emergence of the pine needle gall midge (Diptera: Cecidomyiidae). J. Appl. Entomol. 131: 674 Ð 683. Southwood, T.R.E. 1977. Habitat, the templet for ecological strategies? J. Anim. Ecol. 46: 337Ð365. SPSS. 2006. SPSS 14 for Windows, userÕs guide. SPSS, Chicago, IL. Tauber, M. J., and C. A. Tauber. 1976. Insect seasonality: diapause maintenance, termination, and postdiapause development. Annu. Rev. Entomol. 21: 81Ð107. Tauber, M. J., C. A. Tauber, and S. Masaki. 1986. Seasonal adaptations in insects. Oxford University Press, New York. Wagner, T. L., H. I. Wu, P.J.H. Sharpe, R. M. Schoolfield, and R. N. Coulson. 1984. Modeling insect development

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rates: a literature review and application of a biophysical model. Ann. Entomol. Soc. Am. 77: 208 Ð225. Williams, D. W. 1977. Effects of temperature and moisture on overwintering pupae of Heliothis zea (Boddie). MS thesis, North Carolina State University, Raleigh, NC. Wilson, A.G.L. 1983. Abundance and mortality of overwintering Heliothis spp. J. Aust. Entomol. Soc. 22: 191Ð199. Wilson, A.G.L., T. Lewis, and R. B. Cunningham. 1979. Overwintering and emergence of Heliothis armigera (Hu¨ bner) (Lepidoptera: Noctuidae) in Namoi Valley, New South Wales. Bull. Entomol. Res. 69: 97Ð109. Zalucki, M. P., G. Daglish, S. Firempong, and P. H. Twine. 1986. The biology and ecology of Heliothis armigera (Hu¨ bner) and H. punctigera Wallengren (Lepidoptera: Noctuidae) in Australia: what do we know? Aust. J. Zool. 34: 779 Ð 814. Zhou, X., S. W. Applebaum, and M. Coll. 2000a. Overwintering and spring migration in the bollworm Helicoverpa armigera (Lepidoptera: Noctuidae) in Israel. Environ. Entomol. 29: 1289 Ð1294. Zhou, X., M. Coll, and S. W. Applebaum. 2000b. Effect of temperature and photoperiod on juvenile hormone biosynthesis and sexual maturation in the cotton bollworm, Helicoverpa armigera: implications for life history traits. Insect Biochem. Mol. Biol. 30: 863Ð 868. Received 17 May 2009; accepted 6 April 2010.