Physiological Constraints on the Overwintering Potential of the

PHYSIOLOGICAL ECOLOGY

Physiological Constraints on the Overwintering Potential of the Soybean Aphid (Homoptera: Aphididae) B. P. MCCORNACK,1 M. A. CARRILLO, R. C. VENETTE,

AND

D. W. RAGSDALE

University of Minnesota, Department of Entomology, 219 Hodson Hall, 1980 Folwell Ave., St. Paul, MN 55108

Environ. Entomol. 34(2): 235Ð240 (2005)

ABSTRACT The soybean aphid, Aphis glycines Matsumura, has a heteroecious, holocyclic life cycle. Soybean aphids overwinter as eggs, hatch in the spring, reproduce asexually, and undergo three or more generations on buckthorn, Rhamnus spp., before migrating to a secondary host such as soybean, Glycine max (L.) Merr. The ability of different soybean aphid life stages to survive low temperatures potentially experienced during fall or winter is not known. The objectives of this study were to determine the supercooling point (SCP) of various soybean aphid life stages and to determine the annual probability that winter temperatures within the North Central region of the United States would equal or fall below the mean SCP of soybean aphid eggs. Aphid eggs are considered the most cold-hardy stage; therefore, their SCP can be used as a conservative estimate for aphid overwintering mortality. In our study, eggs had the lowest mean SCP (approximately ⫺34⬚C) among all life stages, whereas gynoparae and oviparae had the highest mean SCPs (approximately ⫺15⬚C). During the winter, extreme low air temperatures are likely to reach or exceed the mean SCP of soybean aphid eggs in portions of northern Minnesota, northern Wisconsin, and the upper peninsula of Michigan. Thus, widespread successful overwintering in the northern United States and southern Canada is less likely than in Illinois, Indiana, Ohio, Iowa, southern Minnesota, southern Wisconsin, and the lower peninsula of Michigan. KEY WORDS Aphis glycines, soybean aphid, supercooling point, overwintering, distribution

THE SOYBEAN APHID, Aphis glycines Matsumura, was Þrst detected in the United States in July 2000 (reviewed in Ragsdale et al. 2004). Soybean aphids have a heteroecious, holocyclic life cycle (i.e., alternates hosts and reproduces sexually during part of its life cycle) (Wu et al. 2004). When exposed to reduced photoperiods and cooler temperatures, winged soybean aphid males (androparae) and asexual females (gynoparae) are produced on a secondary host such as soybean, Glycine max (L.) Merr. These winged forms disperse to the primary overwintering host, buckthorn, Rhamnus spp. (Ragsdale et al. 2004). Voegtlin et al. (2004b) list two buckthorn species, R. cathartica L. and R. alnifolia LÕHe´ ritier, as suitable hosts for soybean aphid in North America. Nine other buckthorn species were not found suitable (Voegtlin et al. 2004b). Gynoparae deposit wingless oviparae on buckthorn that will mate with the androparae. Oviparae deposit overwintering eggs between or near buckthorn leaf buds rather than producing live young (Ragsdale et al. 2004). Soybean aphid eggs are thus relatively exposed and likely to experience winter air temperatures unless they are covered with snow. In the spring, soybean aphid eggs that survived the winter hatch into a stem mother (fundatrix) that produces 1

Corresponding author, e-mail: [email protected].

the Þrst parthenogenic generation of the year. Three or more generations occur on buckthorn with each subsequent generation producing an increasing proportion of winged adults (virginoparae) that leave buckthorn in search of secondary hosts, typically soybean (Voegtlin et al. 2004b). Once on soybean, high soybean aphid densities cause damage by reducing plant height, pod number, and total yield (Dai and Fan 1991, Lin et al. 1993, Wang et al. 1996) and by vectoring several viruses (Hill et al. 2001). Hirano et al. (1996) and McCornack et al. (2004) reported temperatures required for soybean aphid growth and reproduction. However, the degree to which various soybean aphid life stages can withstand cold temperatures is not known. Resident soybean aphid populations depend on successful overwintering of eggs (Ragsdale et al. 2004). Estimating the distribution of surviving soybean aphids may improve efforts to understand spring colonization of soybean Þelds and identify source populations for large-scale soybean aphid dispersal across the North Central region of the United States. These estimates should enable soybean growers and crop consultants to focus sampling efforts on areas where the probability of successful overwintering is greatest, thus increasing their chances for early detection of soybean aphid populations.

0046-225X/05/0235Ð0240$04.00/0 䉷 2005 Entomological Society of America

236

ENVIRONMENTAL ENTOMOLOGY

The capacity for soybean aphid eggs to survive winter temperatures may depend on their ability to supercool and tolerate prefreeze temperatures (James and Luff 1982, Strathdee et al. 1995). Supercooling is a form of protection against low temperatures in which insects lower the freezing point of body ßuids to avoid formation of ice crystals (Lee 1991). No aphid species is known to survive ice crystallization; thus, all stages may be considered freeze intolerant. However, apterae and alatae are likely to die from chill injury before freezing occurs at temperatures ⬇15Ð18⬚C above the supercooling point (SCP) (Knight et al. 1986, Bale et al. 1988, Bale 1996). The SCP is the temperature at which spontaneous freezing occurs (Salt 1961, Sømme 1982), and for aphid eggs, is the lowest possible temperature reached before instantaneous death occurs (Salt 1961, Sømme 1982). Aphid eggs have very low SCPs (e.g., Sømme 1969, Parry 1979, James and Luff 1982, OÕDoherty 1986, Kaneko 1993, Strathdee et al. 1995) and can survive chilling from 3 to 6 mo at temperatures above the SCP (James and Luff 1982, Strathdee et al. 1995). Therefore, the SCP for eggs can be used as a conservative estimate for predicting aphid overwintering mortality. The objectives of this study were to determine the SCPs of various soybean aphid life stages under controlled laboratory conditions and to determine the annual probability that winter temperatures within the northcentral region of the United States would equal or fall below the mean SCP of soybean aphid eggs. Materials and Methods Insects and Plant Material. Laboratory colonies of soybean aphids were started in July 2002 with Þeldcollected apterae from soybean Þelds at the University of Minnesota Rosemount Research and Outreach Center (Rosemount, MN). Colonies were maintained on soybean seedlings (cultivar M96 Ð133151) at 20 ⫾ 1⬚C, 44 Ð 48% RH, and a photoperiod of 16:8 (L:D) h. Adult apterae and alatae voucher specimens were placed in the insect museum at the University of Minnesota (UMSP 000083760). Adult apterae were transferred from heavily infested plants to young, noninfested soybean plants every 2 wk (McCornack et al. 2004). Supercooling points for all life stages were measured for aphids from the source colony. Supercooling points were also measured for a separate cohort of apterous adults that were collected on 18 September 2002 from soybean plants grown on the St. Paul Agricultural Experiment Station (University of Minnesota). Aphids were collected by removing green foliage from plants. Aphids and plant material were brought to the laboratory and stored in an upright refrigerator at 3Ð5⬚C for ⬇3 h before testing. The soybean aphid colony was induced, through manipulation of the photoperiod, to produce eggs on buckthorn, R. cathartica, under greenhouse conditions. Nine buckthorn plants with a mean height of 0.76 ⫾ 0.09 m were collected from Rosemount, MN, and transplanted to 0.30-m-diameter pots in Septem-

Vol. 34, no. 2

ber 2002. Pots were placed in a greenhouse at 24 ⫾ 4⬚C and 55 ⫾ 5% RH. Buckthorn plants were examined for aphid eggs, nymphs, and adults before transplanting. Supplemental light sources were not used, and buckthorn plants were exposed to natural day lengths. The photoperiod ranged from 13:11 (L:D) h at the beginning of the experiment to 9:15 (L:D) h on completion of the study. Crowded soybean aphid colonies on soybean containing large numbers of alatoid nymphs and winged alatae were placed 1 m from the potted buckthorn and were exposed to the same greenhouse conditions. Gynoparae and androparae leaving the infested soybean were allowed to colonize the potted buckthorn for the duration of the study. Gynoparae and oviparae were Þrst observed on the transplanted buckthorn plants in early October when the photoperiod was 11:13 (L:D) h. Subsequent soybean aphid egg production on buckthorn by oviparae was monitored weekly. Soybean aphid eggs were Þrst observed in early November and were allowed to accumulate on the buckthorn stems for 3 wk. Fertilized eggs were shiny and black in comparison with unfertilized eggs, which were dull and yellow/green in color (Blackman 1987). Only fertilized eggs were used to determine SCPs. SCP Determination. Soybean aphids at different stages of development were used for SCP determination as in Carrillo et al. (2004). Developmental stages were determined based on the number of antennal segments and caudal size and shape (Zhang 1988). Eggs and other developmental stages were attached individually to 40- and 36-gauge copper-constantan thermocouples, respectively, using a thin Þlm of high vacuum grease (Dow Corning, Midland, MI). The specimens were cooled at a rate of approximately ⫺1⬚C/min (Carrillo et al. 2004) to a target temperature of ⫺60 or ⫺40⬚C for eggs and other stages of development, respectively. Temperatures were recorded using a multi-channel data logger (Personal Daq/56 data acquisition system; Iotech Inc., Cleveland OH), which transferred data every 1 s through a USB cable directly into a computer. Data were downloaded into a spreadsheet and graphed, and the SCPs were determined as the lowest temperature reached before freezing. Freezing was visualized by the emission of an exotherm (i.e., a small peak indicating heat release during the phase change) (Lee 1991). Because SCPs for different developmental stages were measured in separate trials, statistical comparisons among life stages were not appropriate and are often not conducted for SCP studies (Ring 1972, Lee and Denlinger 1985, Rosales et al. 1994, Chauvin and Vannier 1997). However, the distributions of SCPs were tested for normality using the Royston (1992) modiÞcation to the ShapiroÐWilk W-test (Shapiro and Wilk 1965 and PROC UNIVARIATE; SAS Institute 2001). Median, mean, SE, and range were used to describe the distribution of SCPs for each stage. Temperature Probability Maps. We obtained historical records from the National Climatic Data Center (Midwest Regional Climate Center, Champaign, IL) for 395 weather stations on a ⬇50-km grid.

April 2005 Table 1.

MCCORNACK ET AL.: OVERWINTERING CONSTRAINTS OF SOYBEAN APHIDS

237

SCPs for soybean aphid, A. glycines, nymphs, adults, and eggs from greenhouse and field-established colonies

Life stage Apterae First instar Second instar Third instar Fourth instar Adult Adultc Oviparae Alatae Virginoparae Gynoparae Eggs

Host plant

Mean SCP ⫾ SE

Median SCP

n

SCP range (⬚C) (min, max)

Shapiro-Wilk (W-value)a

Pb

Soybean Soybean Soybean Soybean Soybean Soybean Buckthornd

⫺27.7 ⫾ 0.2 ⫺27.2 ⫾ 0.2 ⫺26.1 ⫾ 0.2 ⫺25.5 ⫾ 0.2 ⫺24.8 ⫾ 0.2 ⫺25.4 ⫾ 0.2 ⫺14.9 ⫾ 0.9

⫺27.7 ⫺27.0 ⫺26.4 ⫺25.6 ⫺24.9 ⫺25.8 ⫺13.2

13 13 15 14 18 15 18

(⫺28.6, ⫺26.5) (⫺28.3, ⫺26.3) (⫺26.9, ⫺24.8) (⫺26.3, ⫺24.5) (⫺25.8, ⫺23.1) (⫺26.8, ⫺23.4) (⫺21.9, ⫺11.1)

0.942 0.956 0.876 0.948 0.928 0.892 0.859

0.484 0.685 0.041 0.524 0.178 0.072 0.012

Soybean Buckthorn Buckthorn

⫺24.9 ⫾ 0.3 ⫺15.2 ⫾ 1.0 ⫺33.9 ⫾ 0.6

⫺25.4 ⫺15.2 ⫺33.9

17 18 10

(⫺26.4, ⫺22.7) (⫺22.2, ⫺9.0) (⫺37.4, ⫺31.9)

0.934 0.893 0.920

0.249 0.043 0.353

0 ⬍ W-value ⱕ 1. Probability that the observed data came from a normal distribution. Collected from a soybean Þeld at the St. Paul Agricultural Experiment Station, University of Minnesota. d Rhamnus cathartica L. a

b c

Weather stations were selected if they were currently operational and had ⬎30 yr of historical weather data. The oldest observations dated to 1888. On average, each station had 73 ⫾ 26 (SD) yr of temperature data. We calculated the proportion of years in which the extreme low temperature was less than or equal to ⫺34⬚C. Spatially explicit proportions were imported into a geographic information system (ArcView 3.2; ESRI, Redlands, CA). As recommended for isotropic, regional temperature data by Collins and Bolstad (1996), interpolation between weather stations was performed by optimized inverse distance weighting. As per Bolstad et al. (1998), optimization was achieved by withholding a randomly selected station from the data set, interpolating a surface with a power parameter of 1Ð 4, 6, or 8, and comparing the predicted and known values at the withheld location (30 stations ⫻ 6 power parameters ⫽ 180 interpolated maps). The optimization process was restricted to weather stations in Minnesota, Wisconsin, and Iowa, where substantial spatial variation in the probabilities occurred. A power parameter of 1 gave the lowest mean absolute error (0.097). A power parameter of 2 had a similar mean absolute error (0.098) but did not obscure known geographic anomalies in temperature; therefore, this parameter was used for interpolation. Power parameters ⱖ3 had mean absolute errors of ⱖ0.103. Table 2.

Acyrthosiphon pisum (Harris)a Aphis fabae Scopoli Diuraphis noxia Mordvilko Elatobium abietinum (Walker) Megoura crassicauda Mordvilkoa Myzus persicae (Sulzer) Sitobion avenae (F.)

b

SCPs. Temperatures at which all life stages froze were normally distributed, except third-instar apterae, oviparae, and gynoparae (Table 1). Apterous (i.e., greenhouse and Þeld-collected) and alate adults had SCPs ⬇ ⫺25⬚C. In general, the SCP of nonsexual, laboratory-maintained, apterous nymphs increased (i.e., insects froze at warmer temperatures) at later stages of development. This trend in SCPs for various soybean aphid life stages has also been observed in other aphid species such as Acyrthosiphon pisum (Harris), Aphis fabae Scopoli, Diuraphis noxia Mordvilko, Elatobium abietinum (Walker), Megoura crassicauda Mordvilko, Myzus persicae (Sulzer), and Sitobion avenae (F.) (Table 2). Possible explanations for this trend in SCPs have been reported. Traditionally, samples with smaller volumes are thought to contain fewer impurities (e.g., gut content, bacteria), thus reducing sites for ice nucleation and lowering SCPs (Wilson et al. 2003). In contrast, Zachariassen et al. (2004) challenged this theory and attributed this trend to a positive correlation between temperature of ice nucleation and water volumes. In the case of the soybean aphid, the relationship between volume and/or impurities and SCP needs further investigation. Mean SCPs for reproductive soybean aphids collected from soybean (i.e., adult apterae and virgino-

Reported mean SCPs for different life stages of various aphid species Species

a

Results and Discussion

Apterous instar SCP (⬚C)

Adult SCP (⬚C)

First

Second

Third

Fourth

Apterae

Alatae

⫺26.3 ⫺26.9 ⫺26.8 ⫺20.6 ⫺26.6 ⫺26.2 ⫺27.0

⫺26.7 ⫺26.5 ⫺25.9 ⫺16.2 ⫺26.2 ⫺24.8 ⫺25.9

⫺25.9 ⫺23.8 ⫺25.5 ⫺16.0 ⫺25.4 ⫺23.8 ⫺25.2

⫺24.2 ⫺25.0 ⫺25.3 ⫺15.6 ⫺25.0 ⫺24.2 ⫺24.4

⫺23.7 ⫺23.6 ⫺24.9 ⫺15.7 ⫺24.5 ⫺24.2 ⫺24.2

Ñb ⫺23.9 Ñ Ñ Ñ ⫺23.9 ⫺23.3

Values were approximated from Fig. 1 in Asai et al. (2002). No value was reported.

Reference Asai et al. 2002 OÕDoherty 1986 Butts 1992 Powell 1974 Asai et al. 2002 Bale et al. 1988 Knight et al. 1986

238 Table 3.


Vol. 34, no. 2

Reported ranges of mean SCPs for eggs of various aphid species Species

Range mean SCP (⬚C)

Reference

Acyrthosiphon brevicorne Hille Ris Lambers Acyrthosiphon svalbardicum Heikinheimo Hyalopterus pruni (Geoffroy)

Nonseasonal ranges ⫺38.7 to ⫺35.5a ⫺38.2 to ⫺36.2a ⫺36.8 to ⫺34.4b

Strathdee et al. 1995 Strathdee et al. 1995 Sømme 1969

Seasonal rangesc ⫺37.5 to ⫺29.1 ⫺39.9 to ⫺32.6 ⫺20.0 to ⫺36.0 ⫺35.7 to ⫺29.4 ⫺43.0 to ⫺25.0 ⫺40.0 to ⫺29.0

OÕDoherty 1986 Sømme 1969 Parry 1979 Kaneko 1993 James and Luff 1982 Strathdee et al. 1995

Aphis fabae Scopoli Aphis pomi De Geer Cinara pilicornis (Hartig) Myzus mumecola (Matsumura) Rhopalosiphum insertum (Walker) Rhopalosiphum padi (L.)

a Lowest mean SCPs represent eggs acclimated at ⫺30 and ⫺10⬚C for 1 mo for A. brevicorne and A. svalbardicum, respectively. Highest mean SCPs represent nonacclimated eggs. b Collected from Akershus, Norway. Lowest mean SCP represents eggs collected on 5 Nov. 1968. Highest mean SCP represents eggs collected on 16 Dec. 1968. c The reported ranges represent eggs exposed to seasonal changes during overwintering months (⬇4 Ð7 mo).

parae) were ⬇10⬚C lower than mean SCPs for aphids collected from buckthorn (i.e., oviparae and gynoparae; Table 1). Similar results were reported by OÕDoherty (1986) for A. fabae, where the mean SCP of virginoparae fed on a herbaceous host was ⬇10⬚C lower than oviparae fed on a woody host. In addition, OÕDoherty (1986) concluded that the supercooling capacity of A. fabae was dependent on host type and was reversible through hostÐplant transfer experiments. Therefore, our results suggest that the reduced supercooling capacity of soybean aphids fed on buckthorn could have been inßuenced by host type (i.e., woody versus herbaceous). However, further experiments are needed to understand the reasons for reduced ability of aphids to supercool when fed on buckthorn. Although aphid nymphs and adults have low SCP values, damage from cold is caused before SCPs are ever reached (Knight et al. 1986). As temperatures decrease below 0⬚C, more aphids are killed, and surviving individuals may incur cryo-injuries that could subsequently affect aphid population dynamics (Hutchinson and Bale 1994). For example, Hutchinson and Bale (1994) reported reduced development, fecundity, and survivorship of adult Rhopalosiphum padi L. that had been exposed to ⫺5 and ⫺7.5⬚C. Therefore, aphids should be classiÞed as chill-susceptible rather than freeze intolerant (Knight et al. 1986, Bale et al. 1988, Bale 1996). According to Sømme (1982), aphid eggs are the most cold-hardy stage of the aphid life cycle. In addition, no insect eggs are known to survive freezing (Sømme 1982). In our study, soybean aphid eggs had mean SCPs ⬇ ⫺34⬚C and were normally distributed (Table 1). Other researchers have reported similar SCPs for eggs of other aphid species (Table 3). Mean SCPs for soybean aphid eggs were within the range of mean SCPs reported for other aphid species in the same genus (e.g., Sømme 1969, OÕDoherty 1986; Table 3). The SCPs of aphid eggs can ßuctuate between ⫺43 and ⫺25⬚C depending on the time of year, with the lowest mean SCPs achieved during the coldest months (Sømme 1969, James and Luff 1982,

OÕDoherty 1986, Kaneko 1993, Strathdee et al. 1995). In some aphid species, this seasonal change in cold hardiness is often attributed to shifts in glycerol and mannitol levels and is inßuenced by previous experience of low temperatures and the time of year (Sømme 1964, Dixon 1985). Eggs used in our study were not acclimated to cold temperatures before SCPs were measured. If acclimation inßuences the SCP, soybean aphid eggs collected from the Þeld during the coldest winter months would conceivably freeze at colder temperatures than we have reported. We did not use Þeld-collected eggs of soybean aphid because they have proven extremely difÞcult to locate in Minnesota (BPM, RCV, and DWR, unpublished data) and currently cannot be distinguished from eggs of buckthorn aphid, Aphis nasturtii Kaltenbach, that may also occur on buckthorn (Voegtlin et al. 2004a). In general, aphid eggs are better adapted to surviving lower temperatures than active stages (OÕDoherty 1986, Strathdee et al. 1995). Emerging holocyclic populations must undergo one or two full generations on the primary host before dispersing to its secondary host (Dixon 1973). Even if temperatures never reach the SCP, active stages in late fall and early spring are subject to subzero temperatures (i.e., hard frosts) and may die as a result of chill injury. According to Bale (2002), winter temperatures give a good estimation of insect abundance the following season. Therefore, the ability of soybean aphid to overwinter not only depends on the proportion of surviving eggs but also on the sexual forms and spring populations withstanding exposure to cold temperatures. Temperature Probability Maps. Soybean aphid eggs cannot survive temperatures lower than the SCP, and where such temperatures occur, eggs cannot overwinter. Extreme low air temperatures are likely to reach or exceed the mean SCP of soybean aphid eggs (approximately ⫺34⬚C) in portions of northern Minnesota, northern Wisconsin, and the upper peninsula of Michigan (Fig. 1). This probability is ⬎50% in 56% of Minnesota, 20% of Wisconsin, and 6% of Michigan. Because soybean aphids deposit eggs along leaf buds, eggs are unlikely to beneÞt from insulating effects of

April 2005

MCCORNACK ET AL.: OVERWINTERING CONSTRAINTS OF SOYBEAN APHIDS

239

Fig. 1. Probability of having an extreme low annual temperature event less or equal to the mean supercooling point of eggs (⫺34⬚C) of the soybean aphid, A. glycines.

a snow pack. Consequently, overwintering is not likely in these areas. Winter temperatures are more favorable for overwintering in Missouri, Kentucky, Illinois, Indiana, Ohio, Iowa, southern Minnesota, southern Wisconsin, and the lower peninsula of Michigan. Therefore, soybean aphid populations present in northern United States and southern Canada are more likely to be the result of soybean aphid migration than of localized overwintering. In some of these areas, the limited availability of overwintering hosts may further restrict chances for successful overwintering (Voegtlin et al. 2004b). Proximity to successful overwintering sites is likely to inßuence which soybean Þelds are Þrst colonized by soybean aphid. Results in Fig. 1 do not account for possible small-scale (⬍50 km), local microclimates (i.e., river valleys, snow pack), where temperatures might permit overwintering; conversely, if prefreeze mortality occurs, as with other species, the area of successful overwintering would be smaller than predicted. Therefore, the map presented in this study only provides a regional prediction of soybean aphid overwintering sites based on the probability of these areas having temperature events at or below the SCP of eggs. Future research on cold hardiness of soybean aphid should include the survival of eggs at various subzero temperatures (e.g., long-term exposure experiments) and the long-term seasonal effects (i.e., changes in photoperiod and ßuctuating temperatures) on aphid reproduction when they are exposed to temperatures

above the SCP. However, our study provides a basis for estimating the potential distribution of surviving soybean aphid populations and improves our understanding of spring colonization across the northcentral region of the United States. Acknowledgments We thank E. Hodgson (University of Minnesota) and three anonymous reviewers for helpful comments on an earlier version of this manuscript. This study was supported in part by the North Central Soybean Research Program, the University of Minnesota Rapid Agricultural Response Fund, and a University of Minnesota Doctoral Dissertation Fellowship awarded to M.A.C.

References Cited Asai, M., H. Yoshida, K. Honda, and H. Tsumuki. 2002. Cold hardiness of three aphid species, Acyrthosiphon pisum, Megoura crassicauda and Aulacorthum solani (Hemiptera: Aphididae). Appl. Entomol. Zool. 37: 341Ð346. Bale, J. S. 1996. Insect cold hardiness: a matter of life and death. Eur. J. Entomol. 93: 369 Ð382. Bale, J. S. 2002. Insects and low temperatures: from molecular biology to distributions and abundance. Phil. Trans. R. Soc. Lond. B. 357: 849 Ð 862. Bale, J. S., R. Harrington, and M. S. Clough. 1988. Low temperature mortality of the peach-potato aphid Myzus persicae. Ecol. Entomol. 13: 121Ð129. Blackman, R. L. 1987. Reproduction, cytogenetics, and development, pp. 163Ð207. In A. K. Minks and P. Harrewijn

240


(eds.), vol. 2A. Aphids: their biology, natural enemies and control. Elsevier, Amsterdam, The Netherlands. Bolstad, P. V., L. Swift, F. Collins, and J. Re´gnie`re. 1998. Measured and predicted air temperatures at basin to regional scales in the southern Appalachian mountains. Agric. For. Meteorol. 91: 161Ð176. Butts, R. A. 1992. Cold hardiness and its relationship to overwintering of the Russian wheat aphid (Homoptera: Aphididae) in southern Alberta. J. Econ. Entomol. 85: 1140 Ð1145. Carrillo, M. A., N. Kaliyan, C. A. Cannon, R. V. Morey, and W. F. Wilcke. 2004. A simple method to adjust cooling rates for supercooling point determination. CryoLetters. 25: 155Ð160. Chauvin, G., and G. Vannier. 1997. Supercooling capacity of Tineola bisselliella (Hummel) (Lepidoptera: Tineidae): its implication for disinfestation. J. Stored Prod. Res. 33: 283Ð287. Collins, F. C., Jr., and P. V. Bolstad. 1996. A comparison of spatial interpolation techniques in temperature estimation. Available online at http://www.ncgia.ucsb. edu/conf/SANTA_FE_CD-ROM/sf_papers/collins_fred/ collins.html. Accessed November 18, 2004. Dai, Z., and J. Fan. 1991. Effects of aphid population dynamics and damage period on soybean yield. J. Shen. Agric. Univ. 22: 135Ð139. Dixon, A.F.G. 1973. Biology of aphids. Arnold, London, UK. Dixon, A.F.G. 1985. Aphid ecology. Chapman & Hall, New York. Hill, J. H., R. Alleman, D. B. Hogg, and C. R. Grau. 2001. First report of transmission of Soybean mosaic virus and Alfalfa mosaic virus in the New World. Plant Dis. 85: 561. Hirano, K., K. Honda, and S. Miyai. 1996. Effects of temperature on development, longevity, and reproduction of the soybean aphid, Aphis glycines (Homoptera: Aphididae). Appl. Entomol. Zool. 31: 178 Ð180. Hutchinson, L. A., and J. S. Bale. 1994. Effects of sublethal cold stress on the aphid Rhopalosiphum padi. J. Appl. Ecol. 31: 102Ð108. James, B. D., and M. L. Luff. 1982. Cold hardiness and development of eggs of Rhopalosiphum insertum. Ecol. Entomol. 7: 277Ð282. Kaneko, J. 1993. Change in supercooling point of eggs of aphid, Myzus mumecola (Matsumura), through overwintering period. Ann. Rep. Plant Prot. North Japan. 44: 167Ð168. Knight, J. D., J. S. Bale, F. Franks, S. F. Mathias, and J. G. Baust. 1986. Insect cold hardiness: supercooling points and pre-freeze mortality. CryoLetters. 7: 194 Ð203. Lee, R. E., Jr. 1991. Principles of insect low temperature tolerance, pp. 17Ð 46. In R. E. Lee Jr. and D. L. Denlinger (eds.), Insects at low temperature, 1st ed. Chapman & Hall, New York. Lee, R. E., Jr., and D. L. Denlinger. 1985. Cold tolerance in diapausing and non-diapausing stages of the ßesh ßy, Sarcophaga crassipalpis. Physiol. Entomol. 10: 309 Ð315. Lin, C., L. Li, Y. Wang, Z. Xun, G. Zhang, and S. Li. 1993. Effects of aphid population on the major economic parameters of soybean. Soybean Sci. 12: 252Ð254. McCornack, B. P., Ragsdale, D. W., and R. C. Venette. 2004. Demography of soybean aphid (Homoptera: Aphididae) at summer temperatures. J. Econ. Entomol. 97: 854 Ð 861. O’Doherty, R. 1986. Cold hardiness of laboratory-maintained and seasonally-collected populations of the black

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bean aphid, Aphis fabae Scopoli (Hemiptera: Aphididae). Bull. Entomol. Res. 76: 367Ð374. Parry, W. H. 1979. Factors affecting low temperature survival of Cinara pilicornis eggs on Sitka spruce. Int. J. Biometeor. 23: 185Ð193. Powell, W. 1974. Supercooling and the low-temperature survival of the green spruce aphid Elatobium abietinum. Ann. Appl. Biol. 78: 27Ð37. Ring, R. A. 1972. Relationship between diapause and supercooling in the blowßy, Lucilia sericata (Mg.) (Diptera: Calliphoridae). Can. J. Zool. 50: 1601Ð1605. Ragsdale, D. W., D. J. Voegtlin, and R. J. O’Neil. 2004. Soybean aphid biology in North America. Ann. Entomol. Soc. Am. 97: 204 Ð208. Rosales, A. L., E. S. Krafsur, and Y. Kim. 1994. Cryobiology of the face ßy and house ßy (Diptera: Muscidae). J. Med. Entomol. 31: 671Ð 680. Royston, J. P. 1992. Approximating the Shapiro-WilkÕs W-Test for non-normality. Statistics Computing. 2: 117Ð 119. Salt, R. W. 1961. Principles of insect cold-hardiness. Annu. Rev. Entomol. 6: 55Ð74. SAS Institute. 2001. PROC userÕs manual, 6th ed. SAS Institute, Cary, NC. Shapiro, S. S., and M. B. Wilk. 1965. An analysis of variance test for normality (complete samples). Biometrika. 52: 591Ð 611. Sømme, L. 1964. Effects of glycerol on cold hardiness in insects. Can. J. Zool. 43: 87Ð101. Sømme, L. 1969. Manitol and glycerol in overwintering aphid eggs. Norsk Entomol. Tidsskr. 16: 107Ð111. Sømme, L. 1982. Supercooling and winter survival in terrestrial arthropods. Comp. Biochem. Physiol. 73A: 519 Ð 543. Strathdee, A. T., G. G. Howling, and J. S. Bale. 1995. Cold hardiness of overwintering aphid eggs. J. Insect. Physiol. 41: 653Ð 657. Voegtlin, D. J., S. E. Halbert, and G. Qiao. 2004a. A guide to separating Aphis glycines Matsumura and morphologically similar species that share its hosts. Ann. Entomol. Soc. Am. 97: 227Ð232. Voegtlin, D. J., R. J. O’Neil, and W. R. Graves. 2004b. Tests of suitability of overwintering hosts of Aphis glycines: identiÞcation of a new host association with Rhamnus alnifolia LÕHe´ ritier. Ann. Entomol. Soc. Am. 97: 233Ð234. Wang, S., X. Bao, Y. Sun, R. Chen, and B. Zhai. 1996. Effects of soybean aphid, Aphis glycines, on soybean growth and yield. Soybean Sci. 15: 243Ð247. Wilson, P. W., A. F. Heneghan, and A.D.J. Haymet. 2003. Ice nucleation in nature: supercooling point (SCP) measurements and the role of heterogeneous nucleation. Cryobiology. 46: 88 Ð98. Wu, Z., D. Schenk-Hamlin, W. Zhan, D. W. Ragsdale, and G. E. Heimpel. 2004. The soybean aphid in China: a historical review. Ann. Entomol. Soc. Am. 97: 209 Ð218. Zachariassen, K. E., E. Kristiansen, S. A. Pedersen, and H. T. Hammel. 2004. Ice nucleation in solutions and freezeavoiding insectsÑ homogenous or heterogeneous? Cryobiology. 48: 309 Ð321. Zhang, X. 1988. Study on the insects of soybean aphid, Aphis glycines Matsumura, in Jilin Province. Acta Agric. Univer. Jilinensis. 10: 15Ð17. Received 2 August 2004; accepted 30 November 2004.