PLANTÐINSECT INTERACTIONS Nest Establishment, Pollination Efficiency, and Reproductive Success of rotundata (: Megachilidae) in Relation to Resource Availability in Field Enclosures

1 2 THERESA L. PITTS-SINGER AND JORDI BOSCH

Environ. Entomol. 39(1): 149Ð158 (2010); DOI: 10.1603/EN09077 ABSTRACT The alfalfa leafcutting , Megachile rotundata (Fabricius), is used to pollinate alfalfa, Medicago sativa L., for seed production in the United States and Canada. It is difÞcult to reliably sustain commercial M. rotundata populations in the United States because of problems with disease, parasites, predators, and unexplained mortality. One possible explanation for early immature mortality is that, relative to ßoral availability, superßuous numbers of are released in alfalfa Þelds where resources quickly become limited. Our objective was to determine how M. rotundata density affects bee nesting, pollination efÞciency, and reproductive success. Various numbers of bees were released into enclo- sures on an alfalfa Þeld, but only 10Ð90% of released female bees established nests. Therefore, a “bee density index” was derived for each enclosure from the number of established females and number of open ßowers over time. As the density index increased, signiÞcant reductions occurred in the number of pollinated ßowers, number of nests, and number of cells produced per bee, as well as the percentage of cells that produced viable prepupae by summerÕs end and the percentage that produced adult bees. The percentage of cells resulting in early brood mortality (i.e., pollen balls) signiÞcantly increased as the density index increased. We conclude that bee nest establishment, pollination efÞciency, and reproductive success are compromised when bee densities are high relative to ßoral resource availability. Open Þeld studies are needed to determine commercial bee densities that result in sustainable bee populations and adequate pollination for proÞtable alfalfa seed production.

KEY WORDS Medicago sativa, alfalfa leafcutting bee, bee density, reproductive success

The alfalfa leafcutting bee, Megachile rotundata the end of the summer, most bee larvae already have (Fabricius) (Hymenoptera: Megachilidae), is a very entered diapause in the prepupal stage and wait to effective pollinator of alfalfa, Medicago sativa L. resume development in the following summer. Other (Fabaceae). This bee has been used as a commercial individuals (so-called “second generation bees”) skip pollinator since the 1960s (Stephen 1961, Hobbs 1967, prepupal diapause and develop to adulthood to pro- Bohart 1971, Richards 1984, Frank 2003), especially for duce a new generation before the end of the summer. alfalfa seed production in northwestern North Amer- For commercial management, the diapausing prepu- ica. As a gregarious cavity-nesting bee, females build pal cells are stored at cold winter temperatures and are nests in holes in wood or polystyrene bee boards that artiÞcially incubated early in the following summer to are provided for them in domiciles placed in Þelds synchronize adult emergence with alfalfa bloom planted for alfalfa seed production. Female bees line (Richards 1984). nest cavities with leaf pieces cut from alfalfa or other The use of ϳ75,000Ð99,000 bees per hectare was plants creating a series of brood cells. By foraging for recommended in the early 1990s for seed production pollen and nectar to make nest provisions, bees pol- on established alfalfa Þelds in the United States (Peter- linate (i.e., “trip”) the alfalfa ßowers. Tripping occurs son et al. 1992). Currently, larger populations of M. when the action of the bee releases pressure on in- rotundata (100,000Ð150,000 bees/ha) are released in terlocking keel petals, allowing the ßowerÕs fused re- Þelds by U.S. alfalfa seed growers (Peterson et al. 1992, productive column to abruptly snap upward from unpublished data), and the seed yields obtained may within the keel (Frank 2003). M. rotundata popula- exceed 1,100 kg/ha (unpublished data). Although the tions are partially bivoltine (Krunic and Hinks 1972, highly efÞcient pollination ability of M. rotundata Tepedino and Parker 1986, Kemp and Bosch 2001). By leads to proÞtable seed yields, U.S. production of live bee progeny is seldom self-sustaining, with often 1 Corresponding author: USDAÐARS Bee Biology & Systematics Ͻ50% replacement of released bees (Pitts-Singer and Laboratory, Logan, UT 84326 (e-mail: Theresa.Pitts-Singer@ars. James 2008). Fewer bees (61,000Ð74,000 bees/ha) are usda.gov). 2 CREAF, Universitat Auto`noma de Barcelona, 08193 Bellaterra, used in Canada (Frank 2003), where bee populations Spain. are increased in alfalfa Þelds and sold to U.S. growers. 150 ENVIRONMENTAL ENTOMOLOGY Vol. 39, no. 1

Therefore, one possible explanation for the inability to starting at Ϸ1400 hours MDT: (1) number of alfalfa sustain commercially viable populations of M. rotun- ßower racemes (from budding to withering); (2) data in the United States is the inundation of Þelds number of open tripped ßowers per raceme; and (3) with numbers of bees that are too high to be supported number of open untripped ßowers per raceme. By by the available ßoral resources (Strickler and Freitas averaging data from the 12 quadrats in each enclosure, 1999, Bosch and Kemp 2005, Maeta and Kitamura such counts revealed the estimated number of open 2007). ßowers and which of these ßowers had presumably For two Þeld seasons (2003 and 2004), we per- been pollinated (tripped) or remained available for formed an experiment to determine the effects of bee bees to visit for pollen (untripped). Starting 1 wk after density (relative to ßower availability) on M. rotun- the bees were released, an otoscope was used to look data nesting, pollination efÞciency, and reproductive inside each nest cavity of each board at twilight on the success. We predicted that nest establishment and same weekday on which ßowers were counted in the individual bee reproductive success would decline as afternoon after bees had had the opportunity to for- the number of foraging females increased relative to age. Both sexes of M. rotundata will rest in bee boards available ßoral resources but that pollination efÞ- at night, and physical characteristics make the sexes ciency would be greatest at higher densities before easy to distinguish from each other. Females do not reaching a plateau. Reductions in reproductive suc- share nests, but males sometimes do. From these cess could be realized as an increase in nonviable “night counts,” we scored the number of males and brood, a more pronounced male-bias in sex ratio, a females to determine the number of live bees and decrease in diapausing prepupae, or a failure of dia- presumably nesting females over time. pausing prepupae to emerge as adults in the next For evaluation of nesting success and bee repro- summer. An increase in pollination efÞciency could be duction per enclosure, we collected all complete and evaluated from the percentage of ßowers that are partial nests produced by the end of the study period tripped. by pulling out the straw inserts containing nest ma- terials. Nests were brought into the USDAÐARS Bee Biology & Systematics Laboratory, and the contents of Materials and Methods all straw inserts were examined using x-radiography In 2003 and 2004, eight screened enclosures (length (30-s exposure at 25 kvp in Faxitron 43804N; Faxitron by width by height ϭ 6.1 by 6.1 by 1.8 m) were X-Ray, Lincolnshire, IL) to determine percentages of established in an alfalfa hay Þeld (variety not speci- cells containing healthy prepupae or adults, “pollen Þed) that was allowed to ßower in North Logan, UT. balls” (provision with dead egg/young larva or no In the center of each enclosure, one small portion of detectable brood; Pitts-Singer 2004), and other imma- a commercial polystyrene bee board (range: length by ture mortality (Stephen and Undurraga 1976). All cells height by depth ϭ 10 by 4 by 9.2 cm to 34 by 15 by 9.2 containing live progeny were removed, placed into cm) was housed in a small plywood domicile sus- gelatin capsules (size 00; Eli Lilly and Co., Indianap- pended Ϸ1.5 m above the ground on a metal post. The olis, IN), and allowed to continue development in number of nesting cavities in the bee boards equaled darkness in an incubator (26ЊC). Any adult that four cavities for each of the females released, ensuring emerged from these cells by October was designated that nest availability was not a limiting factor for nest- as a “second-generation adult bee,” and its sex was ing or reproduction in the enclosure. Paper straws of recorded. Cells with live, diapausing prepupae were slightly smaller diameter than the nest cavities were stored for the winter at 4.5ЊC beginning 1 October and inserted into each hole to facilitate removal of nests at incubated for adult emergence at 29ЊC the following the end of the study for x-radiographic analysis (Ste- June for determination of adult number and sex. These phen and Undurraga 1976) and dissection of cells as bees were designated as “diapausing adult bees.” necessary. Bees that had been purchased from a Ca- For the bee foraging period to be as long as possible, nadian supplier (JWM Leafcutters, Nampa, ID) were bee release occurred after the onset of bloom but not incubated at 29ЊC in June, and they emerged from necessarily at peak bloom that often occurred later. their cocoons in the laboratory after ϳ3 wk. They The duration of bloom and timing for peak bloom were released from small containers into the enclo- within any enclosure could not be accurately pre- sures next to the bee boards. The number of bees dicted before assignment of bee density, although in released in each enclosure was varied to emulate bee 2004, ßowers were counted before bee release to as- densities between 9,100Ð95,300 bees/ha to reßect a sure that bloom was underway. As a result of using series of bee densities ranging from lower than would random assignment, high numbers of bees were some- be used commercially up to a modest commercial times paired with enclosures that ultimately contained level. Two male bees were released for every female relatively low numbers of ßowers. Therefore, because to attain a 2:1 sex ratio, which is near the average sex of uneven ßoral resources and failure of all released ratio for Canadian M. rotundata commercial popula- female bees to establish in the enclosures (see Re- tions (Pitts-Singer and James 2005). sults), a derived metric, “density index,” was used for Throughout the season at weekly intervals starting data analysis and interpretation. Throughout the rest a few days after bee release, the following data were of this manuscript, the density index is deÞned as “the recorded concerning ßower production from within area under a plotted curve of female bees by date 12 quadrats (area of each ϭ 0.3 m2) in each enclosure calculated from the number of female bees counted at February 2010 PITTS-SINGER AND BOSCH: M. rotundata DENSITY AFFECTS SUSTAINABILITY 151 night once a week” per “the area under a plotted curve low (Fig. 1; Table 1). Correlation analysis showed a of estimated open ßowers by date in the enclosure signiÞcant positive relationship between the percent- harboring those females for the same time period.” age of ßowers that were tripped and the density index This density index summarizes bee density relative to (R ϭ 0.53; P ϭ 0.04; Fig. 2A), but a negative relation- ßower availability over the season, given the dynamic ship between the number of ßowers tripped per interaction of female bees and ßower production. female and the density index (R ϭϪ0.83; P Ͻ 0.01; Spearman rank partial correlation analysis of pooled Fig. 2B). data from both years, but controlling for year, was used Production of bee nests and also bee cells per female to determine if the highest number of female bees were negatively correlated with the density index found in the cavities correlated with the highest num- (nests: R ϭϪ0.855; P Ͻ 0.01; bee cells: R ϭϪ0.861; P Ͻ ber of ßowers counted during the study period and to 0.01; Fig. 3A and B). However, bee cell production did evaluate the relationships between the density index not translate directly to reproductive success because and several dependent variables: (1) the percentage many of the cells produced (20Ð100%) were pollen of open ßowers that were tripped, (2) the number of ball cells. The correlation between the percentage of ßowers tripped per highest count of female bees, (3) cells that were pollen balls and the density index was the number of nests and cells produced per the highest signiÞcant and positive (R ϭ 0.62; P ϭ 0.01; Fig. 3C). count of females, (4) the percentage of all cells pro- No death caused by other known M. rotundata mo- duced that became pollen ball cells, (5) diapausing rality factors, such as hymenopteran parasitoids and prepupae, (6) adult progeny (male and female), and chalkbrood (a fungal disease of larvae), was observed; (7) the sex ratios of progeny produced. Additionally, however, some large larvae were found dead from the lengths and widths of all prepupal cocoons formed unknown causes. in cells from all enclosures that ultimately resulted in The density index was not signiÞcantly correlated second generation or diapausing adults were mea- with the percentage of diapausing prepupae (R ϭ sured from x-ray images to estimate prepupal masses Ϫ0.32; P ϭ 0.24; Fig. 4A). However, a signiÞcant neg- (mg) according to the Bosch and Kemp (2005) model ative correlation existed between the density index for this estimation: prepupal weight ϭ 1.42 (cocoon and percentage of cells that survived to adulthood length ϫ width) Ð 10; R2 ϭ 0.67. A three-way factorial (total of second generation and diapausing adults of mixed model analysis of variance (ANOVA) was per- both sexes; R ϭϪ0.64; P Ͻ 0.01; Fig. 4B). Both the formed to evaluate whether prepupal mass (mg) was percentage of all cells that resulted in live female affected by year, sex, or diapause condition (i.e., progeny (R ϭϪ0.55; P ϭ 0.03; Fig. 4C) and male whether prepupae were destined to develop into sec- progeny (R ϭϪ0.66; P Ͻ 0.01) were signiÞcantly and ond-generation or diapausing adults). Correlation negatively correlated with the density index. The den- analyses were computed using the CORR procedure sity index did not correlate with sex ratio (R ϭϪ0.37; and analysis of variance was computed using the P ϭ 0.18; Table 2). MIXED procedure in the SAS System for Windows, Fewer second generation bees were produced in Release 9.2. (SAS Institute, Cary, NC). 2004 than in 2003. Analysis of pooled data from all enclosures in each year revealed differences in pre- papal mass between years and according to sex and Results diapause status (Table 3). Prepupal mass was signiÞ- The number of ßowers was inconsistent between cantly greater for females than males (F ϭ 40.05; df ϭ enclosures because of variability in the plant stand, 1, 331; P Ͻ 0.01), was greater in 2003 than 2004 (F ϭ incidence of plant pests, and yearly differences in 14.24; df ϭ 1, 331; P Ͻ 0.01), and was greater for blooming time in relation to bee release (Fig. 1). In diapausing than second generation individuals (F ϭ both 2003 and 2004, the total number of open ßowers 5.82; df ϭ 1, 331; P Ͻ 0.02; Table 3). No signiÞcant declined by the second week after bee release. Com- interaction was found between any of the indepen- pared with 2003, there were many fewer ßowers in dent variables. 2004, and the number of untripped ßowers (that served as pollen and nectar sources for the bees) Discussion represented a much smaller proportion of open ßow- ers throughout the study (Fig. 1). Clearly, bee reproductive success is limited by the The number of bees counted in cavities at night availability of ßoral resources, yet resource availability within enclosures was always lower than the number is hard to both predict and measure because of the released, but a higher proportion of released bees complex interplay between foraging bees and alfalfa established nests in 2003 when more ßowers were ßower production. By monitoring the numbers of present (Fig. 1; Table 1). However, the highest num- nesting bees (rather than released bees) as well as ber of female bees found in the cavities was not sig- open ßowers throughout our study, we were able to niÞcantly correlated with the highest number of ßow- determine the contribution of only the surviving bees ers counted during study periods (R ϭϪ0.20; P ϭ 0.48; to pollination and reproduction. In our alfalfa Þeld Table 1). enclosures, relatively high numbers of foraging M. Of the available open ßowers in the enclosures in rotundata could not be sustained, and the number of 2003 and 2004, a much higher percentage of them were established females was never as high as the number tripped in 2004, when overall ßower abundance was of females released even in low numbers. Because 152 ENVIRONMENTAL ENTOMOLOGY Vol. 39, no. 1

Fig. 1. By date of data collection for 2003 and 2004, the left y-axis gives the number of M. rotundata female bees released (Þrst data point) and then found nesting in Þeld enclosures and the right y-axis gives estimated numbers of total open (tripped ϩ untripped) alfalfa ßowers and only untripped ßowers. Graphs represent data for enclosures with relatively low, moderate, and high numbers of female bees released in each year. bees were trapped with limited ßoral resources in the can be compared with data from a 2.2-ha alfalfa Þeld enclosures, we surmised that bees found at the highest planted for seed production that had a maximum night counts represented the number of bees that ßower count of 68.7 mil ßowers/ha during the nesting could survive (at least at one point in time) with the season (unpublished data). In each year of this study, amount of resources in the enclosures and that the the alfalfa stand endured various and different prob- missing bees had died. These Þndings further suggest lems that reduced bloom in some of the enclosures, that, in commercial open-Þeld environments, high especially in 2004. Such problems included pests (al- densities of bees may not be sustained; bees in open- falfa weevil, lygus bugs, grasshoppers, mites, meadow Þeld situations may die or simply leave the release site voles, and weeds) that damaged plants, ßowers, or (Bosch and Kemp 2005). seeds. Overall, the small Þeld (Ϸ0.25 ha) used for this trial Using the number of bees that were known to be was spatially heterogeneous and did not offer consis- alive in the enclosures, a derived measurement of bee tently ideal conditions for survival and nesting of many density, the density index, was used to encapsulate the female bees in some enclosures. Extrapolation of dynamic interaction of bees and ßowers over time. ßower counts in our enclosures showed a range of This metric allowed for analysis and interpretation of 23.2Ð50.8 mil ßowers/ha while bees were nesting. This a complex system, despite the variability between en- February 2010 PITTS-SINGER AND BOSCH: M. rotundata DENSITY AFFECTS SUSTAINABILITY 153

Table 1. Number of M. rotundata females and open flowers

Females released versus highest count Range and mean count of open ßowers (% released) (ϫ1,000) 2003 2004 2003 2004 Range Mean Range Mean 10 versus 5 (50%) 16 versus 14 (88%) 18.3Ð168.1 82.5 2.5Ð22.6 9.4 20 versus 17 (85%) 32 versus 23 (72%) 6.8Ð111.3 59.2 2.8Ð27.3 14.7 30 versus 25 (83%) 48 versus 39 (81%) 0.0Ð46.2 18.3 1.8Ð39.7 15.1 40 versus 24 (60%) 64 versus 49 (77%) 0.4Ð27.7 9.6 3.4Ð39.7 18.3 50 versus 45 (90%) 80 versus 42 (53%) 1.3Ð61.8 26.0 6.8Ð32.9 15.9 60 versus 47 (78%) 96 versus 22 (23%) 0.4Ð44.2 17.1 3.4Ð60.5 20.1 90 versus 75 (83%) 112 versus 14 (13%) 3.0Ð33.2 15.0 2.3Ð13.3 6.9 105 versus 80 (76%) 128 versus 13 (10%) 6.4Ð20.9 13.2 0.7Ð32.3 11.3

In a 2-yr study with bees in enclosures on an alfalfa Þeld, the no. M. rotundata females released into Þeld enclosures versus the highest night count of female bees (i.e., surviving females) in nest cavities in enclosures, and the range and mean of open ßowers while bees were present. The ßower data shown here were collected while bees were present in the enclosures. closures in ßoral abundance. Results supported our increase in M. rotundata density. There was a signif- hypothesis that pollination efÞciency and individual icant, but not steep, increase in the percentage of bee reproductive success are impaired by a relative ßowers tripped as bee density index increased. How-

Fig. 2. In alfalfa Þeld enclosures in 2003 and 2004 studies, according to density index, (A) the percentage of the ßowers (estimated for each enclosure) that were tripped and (B) the number of ßowers tripped per the highest count of M. rotundata female bees. 154 ENVIRONMENTAL ENTOMOLOGY Vol. 39, no. 1

Fig. 3. In alfalfa Þeld enclosures in 2003 and 2004 studies, according to density index, (A) the number of nests made per M. rotundata female, (B) the number of cells made per female, and (C) the percentage of all cells that contained pollen balls. February 2010 PITTS-SINGER AND BOSCH: M. rotundata DENSITY AFFECTS SUSTAINABILITY 155

Fig. 4. In alfalfa Þeld enclosures in 2003 and 2004 studies, according to density index, (A) the percentage of all M. rotundata cells that contained diapausing prepupae, (B) the percentage of all cells that produced adult M. rotundata progeny (second generation ϩ diapausing), and (C) the percentage of all cells that produced female adults (second generation ϩ diapausing). 156 ENVIRONMENTAL ENTOMOLOGY Vol. 39, no. 1

Table 2. Number and sex ratio of M. rotundata progeny resources; however, they did not see a signiÞcant dif- ference in the number of cells that failed to produce Density Total no. Adult progeny Year adult bees. indexa live progeny sex ratio Population density and its effects on resource avail- 2003 0.03 15 0.67:1 ability play a role in determining progeny investment 0.12 40 1.35:1 0.47 52 1.00:1 and sex ratio. A study of Osmia cornifrons (Radosz- 1.05 97 1.94:1 kowski) (Megachilidae) showed that an increase in 1.48 14 0.27:1 bee density resulted in a decrease in number of eggs 1.60 29 0.71:1 laid and number of female offspring produced per 3.27 4 0.33:1 3.95 1 No males female (Maeta et al. 2005). The proportion of female 2004 0.63 37 1.10:1 bee progeny was reduced as density increased, but 0.73 26 0.44:1 density had no clear effect on cocoon weights. For O. 0.92 21 0.75:1 cornuta, progeny sex ratio was more male-biased, and 1.03 31 0.24:1 1.27 None No males production of undersized offspring increased when 1.32 2 No males resources were scarce (Bosch 2008). Kim (1999) 2.03 15 0.25:1 found that Megachile apicalis increased maternal in- 2.45 6 2.00:1 vestment in daughters, as well as in total investment per offspring, when more ßoral resources were avail- In a 2-yr study with bees in enclosures on an alfalfa Þeld, according to density index, the total M. rotundata progeny (both sexes of second able. For , Philanthus triangulum F. (Hy- generation ϩ diapausing) produced that survived to adulthood and menoptera: Sphecidae), low food availability resulted the sex ratios (M:F) of those progeny. in a severely male-biased sex ratio (Strohm and Lin- a Area of female bees/area of open ßowers over study period; see senmair 1997). In their resource allocation study in detailed description in Materials and Methods. small tents, Peterson and Roitberg (2006) found that the sex ratio of M. rotundata diapausing progeny (M: ϭ ever, per female contribution to the number of tripped F 2.4:1) was unaffected by resource allocation. ßowers was negatively and dramatically affected by an Their results suggest that all resource allocation treat- increase in the bee density index, thus showing a ments in their tents were somewhat limiting, and thus pronounced reduction in individual bee pollination they concluded that the production of the “cheaper” efÞciency. (smaller) male progeny occurred across treatments As bee density indices increased, reproductive suc- because of those resource limitations. However, our cess decreased. At the higher indices, each bee pro- sex ratios that included both summer-emerging and duced fewer nests and cells and generated a larger diapausing progeny were less male-biased (range, 0.2: proportion of cells that were pollen balls. Additionally, 1Ð2:1) than in the aforementioned study under our the percentage of live adult progeny of both sexes low resource conditions. The sex ratios of only the (including adults that emerged at the end of summer diapausing progeny (range, 0.3:1Ð1.65:1) were even or after winter) was negatively correlated with in- less male-biased than the sex ratio of the summer- creases in bee density index, although the index did emerging bees. Our low sex ratios may be explained by not correlate with the percentage of diapausing pre- the production of female cells Þrst in the nesting cycle pupae (that would be available for pollinating in the when there were enough resources in our enclosures subsequent growing season). In a study of the effects for making these cells, but a lull in the subsequent of limited ßoral resources on M. rotundata reproduc- production of male cells because of the later paucity tion, Peterson and Roitberg (2006) also found in- of resources. creased numbers of bee cells, nests, and progeny made Progeny investment was also shown in prepupal by bees released in tents with relatively high ßoral mass as well as sex ratio. Although we lacked enough resources compared with those with low or medium progeny from each enclosure for a comparison by bee density index, we could compare the estimated weights of nondiapausing and diapausing prepupae by Table 3. Mass of M. rotundata prepupae pooling samples across enclosures in each year. A reduction in M. rotundata offspring investment from 1 Females Males yr to the next is shown by the 2003 prepupae being Year Second Second Diapausing Diapausing heavier than the 2004 prepupae, possibly because of generation generation the fact that the Þeld in 2003 had about twice the 2003 47.48 Ϯ 0.92 49.76 Ϯ 0.93 41.29 Ϯ 0.61 43.33 Ϯ 0.76 number of ßowers as the Þeld in 2004. We also found n ϭ 47 n ϭ 55 n ϭ 65 n ϭ 52 in both years that prepupae destined to remain in Ϯ Ϯ Ϯ Ϯ 2004 42.57 1.07 46.81 0.68 39.04 2.24 39.44 1.27 diapause through the winter were signiÞcantly n ϭ 15 n ϭ 61 n ϭ 5 n ϭ 39 heavier than second-generation prepupae, as we may In a 2-yr study with bees in enclosures on an alfalfa Þeld, the have predicted because M. rotundata adults that de- estimated mass (mg; mean Ϯ SE) of M. rotundata prepupae that were velop from diapausing prepupae are heavier than sum- produced and developed to adults from second generation or dia- mer-emerging adults (Tepedino and Parker 1988). A Ͻ pausing prepupae. Prepupal mass differed signiÞcantly (P 0.05) positive relationship between body size and propen- between years, sexes, and destinies (whether development to adult occurred in summer ϭ second generation, or after winter ϭ diapaus- sity to diapause has been reported in other ing). (Nesin 1985, Danforth 1999, Menu and Desouhant February 2010 PITTS-SINGER AND BOSCH: M. rotundata DENSITY AFFECTS SUSTAINABILITY 157

2002). Studies of the adult-wintering bee O. cornuta References Cited (Megachilidae) provided further support of this idea: Bohart, G. E. 1971. Management of habitats for wild bees. larger O. cornuta were more likely to survive the win- Proc. Tall Timbers Conf. Ecol. Anim. Control Habitat ter and live longer than smaller bees (Bosch and Kemp Manage. 3: 253Ð256. 2004, Bosch and Vicens 2005, Bosch 2008). It is inter- Bosch, J. 2008. Production of undersized offspring in a sol- esting, however, that the diapausing prepupae in 2004 itary bee. Anim. Behav. 75: 809Ð816. weighed less on average than the 2003 second-gener- Bosch, J., and W. P. Kemp. 2004. Effect of pre-wintering and ation prepupae. Therefore, other factors besides pre- wintering temperature regimes on weight loss, survival, pupal weight must be important for the determination and emergence time in the mason bee Osmia cornuta of whether prepupae will enter diapause. Previous (Hymenoptera: Megachilidae). Apidologie 35: 469Ð479. Bosch, J., and W. P. Kemp. 2005. Alfalfa leafcutting bee studies have shown that such factors as photoperiod, population dynamics, ßower availability, and pollination weather conditions throughout immature develop- rates in two Oregon alfalfa Þelds. J. Econ. Entomol. 98: ment, and maternal inheritance signiÞcantly affect the 1077Ð1086. ratio of diapausing and nondiapausing individuals in Bosch, J., and N. Vicens. 2005. Sex allocation in the solitary M. rotundata (Parker and Tepedino 1982, Tepedino bee Osmia cornuta: do females behave in agreement with and Parker 1986, Kemp and Bosch 2000). Overall, our FisherÕs theory? Behav. Ecol. Sociobiol. 59: 124Ð132. study conforms to, but is not completely explained by, Danforth, B. N. 1999. Emergence dynamics and bet hedging in a desert bee, Perdita portalis. Proc. Roy. Soc. Lond. B the idea that nondiapausing prepupae lack the fat 266: 1985Ð1994. stores to survive the winter because their larval pro- Frank, G. 2003. Alfalfa seed & leafcutter bee: productions visions were inadequate because of the low ßoral re- and marketing manual. Irrigated Alfalfa Seed Producers source availability and the resulting competition be- Association, Brooks, Alberta, Canada. tween foraging bees for those sparse resources Hobbs, G. A. 1967. Domestication of alfalfa leaf-cutter bees. (Strickler and Freitas 1999, Bosch and Kemp 2005). Canada Department of Agriculture, Ottawa, Canada. By conÞning bees in enclosures, we eliminated dis- Kemp, W. P., and J. Bosch. 2000. Development and emer- persal as a survival and reproduction strategy for M. gence of the alfalfa pollinator Megachile rotundata (Hy- menoptera: Megachilidae). Ann. Entomol. Soc. Am. 93: rotundata females. Although we unexpectedly found 904Ð911. that the caged situation presented a dilemma for as- Kemp, W. P., and J. Bosch. 2001. Postcocooning tempera- signing bee density, our results using a density index tures and diapause in the alfalfa pollinator Megachile ro- support the idea that the high numbers of bees re- tundata (Hymenoptera: Megachilidae). Ann. Entomol. leased for pollination on U.S. alfalfa seed Þelds could Soc. Am. 94: 244Ð250. negatively affect each beeÕs pollination ability and Kim, J. 1999. Inßuence of resource level on maternal in- reproductive success, which in turn results in poor vestment in a leaf-cutter bee (Hymenoptera: Megachili- population sustainability. For further understanding dae). Behav. Ecol. 10: 522Ð556. Krunic, M. D., and C. F. Hinks. 1972. The effect of tem- of M. rotundata population sustainability in the United perature and of temperature pretreatment on diapause States, an open-Þeld study currently is underway to and on the synchronization of adult emergence in examine the effects of releasing low, moderate, and Megachile rotundata (Hymenoptera: Megachilidae). Can. standard bee densities (range, 37,000Ð124,000 bees/ Entomol. 104: 889Ð893. ha) in open alfalfa seed Þelds. Again, it is imperative Maeta, Y., and K. Kitamura. 2007. EfÞciency of seed pro- to examine the number of nesting bees (including duction in breeding of ladino clover by three species of second-generation adults) and ßoral resources avail- wild bees, Ceratina flavipes Smith, Megachile rotundata able throughout the nesting season. Along with the (Fabricius) and Megachile spissula Cockerell (Hymenop- tera, Apidae and Megachilidae). Chugoku Kontyu 21: basic density-dependent relationships for bee repro- 35Ð53. ductive success found in this study, an open-Þeld study Maeta, Y., S. Yoshida, S. Gotoˆ, and K. Kitamura. 2005. Re- performed over several years will allow for a com- lationship between seed yield of Chinese milk vetch and mercial-scale assessment of both M. rotundata repro- density of female bees, Osmia cornifrons (Radoszkowski) duction and alfalfa seed production. (Hymenoptera, Megachilidae). Chugoku Kontyu 19: 45Ð61. Menu, F., and E. Desouhant. 2002. Bet-hedging for variabil- ity in life cycle duration: bigger and later-emerging chest- Acknowledgments nut weevils have increased probability of prolonged dia- pause. Oecologia 132: 167Ð174. We thank ARS technicians S. Kalaskar, E. Klomps, and Nesin, A. P. 1985. Contribution to the knowledge of the G. Trostle and Utah State University undergraduates M. diapause of some pests of cones and seeds of conifers. Barker, J. Beattie, N. Boehme, C. Gorrell, E. Sharp, and Entomol. Rev. 68: 38Ð43. M. J. Watterson for their combined efforts over the years Parker, F. D., and V. J. Tepedino. 1982. Maternal inßuence in setting up and executing experiments. We also thank on diapause in the alfalfa leafcutting bee (Hymenoptera: statisticians M. West and especially S. Durham for advice Megachilidae). Ann. Entomol. Soc. Am. 75: 407Ð410. in project planning and analysis. We appreciate critical Peterson, J. H., and B. D. Roitberg. 2006. Impact of resource manuscript reviews by S. Durham, W. Goerzen, S. Peterson, and levels on sex ratio and resource allocation in the solitary three anonymous reviewers whose comments and suggestions bee, Megachile rotundata. Environ. Entomol. 35: 1404Ð were very helpful. 1410. 158 ENVIRONMENTAL ENTOMOLOGY Vol. 39, no. 1

Peterson, S. S., C. R. Baird, and R. M. Bitner. 1992. Current Stephen, W. P., and J. M. Undurraga. 1976. X-radiography, status of the alfalfa leafcutting bee, Megachile rotundata, an analytical tool in population studies of the leafcutter as a pollinator of alfalfa seed. Bee Sci. 2: 135Ð142. bee, Megachile pacifica. J. Apic. Res. 15: 81Ð87. Pitts-Singer, T. L. 2004. Examination of “pollen balls” in Strickler, K., and S. Freitas. 1999. Interactions between ßo- nests of the alfalfa leafcutting bee, Megachile rotundata. J. ral resources and bees (Hymenoptera: Megachilidae) in Apic. Res. 43: 40Ð46. commercial alfalfa seed Þelds. Environ. Entomol. 28: 178Ð Pitts-Singer, T. L., and R. R. James. 2005. Emergence suc- 187. cess and sex ratio of commercial alfalfa leafcutting bees, Strohm, E., and K. E. Linsenmair. 1997. Low resource avail- Megachile rotundata Say, from the United States and Can- ability causes extremely male-biased investment ratios in the European , Philanthus triangulm F. (Hyme- ada. J. Econ. Entomol. 98: 1785Ð1790. noptera, Sphecidae). Proc. R. Soc. Lond. B. 264: 423Ð Pitts-Singer, T. L., and R. R. James. 2008. Do weather con- 429. ditions correlate with Þndings in failed, provision-Þlled Tepedino, V. J., and F. D. Parker. 1986. Effect of rearing nest cells of Megachile rotundata (Hymenoptera: temperature on mortality, second-generation emer- Megachilidae) in western North America? J. Econ. En- gence, and size of adult in Megachile rotundata (Hyme- tomol. 101: 674Ð685. noptera: Megachilidae). J. Econ. Entomol. 79: 974Ð977. Richards, K. W. 1984. Alfalfa leafcutter bee management in Tepedino, V. J., and F. D. Parker. 1988. Alternation of sex Western Canada. Agriculture Canada, Ottawa, Canada. ratio in a partially bivoltine bee, Megachile rotundata Stephen, W. P. 1961. ArtiÞcial nesting sites for the propa- (Hymenoptera: Megachilidae). Ann. Entomol. Soc. Am. gation of the leaf-cutter bee, Megachile (Eutricharaea) 81: 467Ð476. rotundata, for alfalfa production. J. Econ. Entomol. 54: 989Ð993. Received 9 March 2009; accepted 28 September 2009.