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Assessing the Mating ‘Health’ of Commercial Honey Bee Queens

Author(s) :David R. Tarpy, Jennifer J. Keller, Joel R. Caren, and Deborah A.


Delaney
Source: Journal of Economic Entomology, 105(1):20-25. 2012.
Published By: Entomological Society of America
DOI:
URL: http://www.bioone.org/doi/full/10.1603/EC11276

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APICULTURE AND SOCIAL INSECTS

Assessing the Mating ‘Health’ of Commercial Honey Bee Queens


DAVID R. TARPY,1,2 JENNIFER J. KELLER,1 JOEL R. CAREN,1 AND DEBORAH A. DELANEY3

J. Econ. Entomol. 105(1): 20Ð25 (2012); DOI: http://dx.doi.org/10.1603/EC11276


ABSTRACT Honey bee queens mate with multiple males, which increases the total genetic diversity
within colonies and has been shown to confer numerous beneÞts for colony health and productivity.
Recent surveys of beekeepers have suggested that Ôpoor queensÕ are a top management concern, thus
investigating the reproductive quality and mating success of commercially produced honey bee
queens is warranted. We purchased 80 commercially produced queens from large queen breeders in
California and measured them for their physical size (fresh weigh and thorax width), insemination
success (stored sperm counts and sperm viability), and mating number (determined by patriline
genotyping of worker offspring). We found that queens had an average of 4.37 ⫾ 1.446 million stored
sperm in their spermathecae with an average viability of 83.7 ⫾ 13.33%. We also found that the tested
queens had mated with a high number of drones (average effective paternity frequency: 17.0 ⫾ 8.98).
Queen “quality” signiÞcantly varied among commercial sources for physical characters but not for
mating characters. These Þndings suggest that it may be more effective to improve overall queen
reproductive potential by culling lower-quality queens rather than systematically altering current
queen production practices.

KEY WORDS honey bee, queen, insemination success, mating number

Honey bees (Apis mellifera L.) are one of the most phenotypes. Numerous strains of honey bees have
beneÞcial insects in agriculture because they serve as been developed, each exhibiting an assortment of se-
invaluable pollinators of crops that are responsible for lectable traits, such as disease- or mite-tolerance, gen-
an estimated 35% of the western human diet (Klein et tleness, honey production, and hygienic behavior
al. 2007, National Research Council 2007). In recent (e.g., Harbo and Harris 1999, Rinderer et al. 2001,
years, much attention has focused on the overwinter- Spivak and Reuter 2001). The goal of these programs
ing lossesÑ both explained and unexplainedÑ of the has been to improve queen genetic quality, which, in
managed honey bee population in the United States turn, improves overall colony productivity.
and Europe (e.g., vanEngelsdorp et al. 2009, Pettis and Independent of genetic stock, a colony can be af-
Delaplane 2010, Potts et al. 2010, vanEngelsdorp fected by numerous factors that greatly diminish the
and Meixner 2010). Addressing overall colony health reproductive potential of its queen. First, a queen can
and productivity is therefore of primary importance be replaced by her worker offspring during superse-
for honey bee and apiculture research. dure events, typically when an aging queen depletes
Because of the central role that queens play within her sperm supply (Butler 1957). Supersedure queens
colonies, improving the productivity and health of are of lower quality compared with those raised under
honey bee colonies is often synonymous with improv- more favorable conditions. For example, Kostarelou-
ing the quality of queens. Indeed, many standard tech- Damianidou et al. (1995) showed that colonies headed
niques of bee management focus on maximizing the by replacement queens produced up to 34% less brood
reproductive potential of queens. For example, and 36% less honey. To maximize queen and colony
Ôswarm box cell-starterÕ colonies (Laidlaw and Page quality, the standard advice to beekeepers is to man-
1997), used to initiate queen rearing in commercial ually replace their queens every spring. Such proce-
operations, maximizes the number of nurse bees that dures can be expensive both directly (for the costs of
produce royal jelly to feed developing gyne brood. the queens themselves) and indirectly (accidental
However, most advances in improving queen quality queen and colony loss, increased labor, and dimin-
have been accomplished by various bee breeding pro- ished honey yields), thus minimizing how often col-
grams, which typically use the instrumental insemi- onies need to be restocked will beneÞt beekeepers.
nation technique to artiÞcially select certain colony Second, a queen may produce brood that is not highly
viable. This is evidenced by “shotgun brood patterns,”
1 Department of Entomology, Campus Box 7613, North Carolina
which are routinely observed by beekeepers and re-
State University, Raleigh, NC 27695-7613.
2 Corresponding author, e-mail: david_tarpy@ncsu.edu. sult in a colonyÕs inability to rapidly increase their
3 Department of Entomology and Wildlife Biology, University of worker force and thus decreases productivity (Woyke
Delaware, 252 Townsend Hall, Newark, DE 19716. 1980, Tarpy and Page 2002), usually as a result of local

0022-0493/12/0020Ð0025$04.00/0 䉷 2012 Entomological Society of America


February 2012 TARPY ET AL.: MATING HEALTH OF QUEEN BEES 21

inbreeding or low population-wide genetic diversity. face many problems that challenge the health and
Therefore, increasing the health and quality of a queenÕs longevity of their stocks. Annual colony losses, in-
brood has direct beneÞts for beekeepers. Finally, a col- creases in chronic infections, and poor queen perfor-
ony can host a litany of parasites and pathogens that mance have caused the apiculture industry to question
signiÞcantly diminish its productivity and overall health. the quality of commercially produced queen honey
Like most domesticated animals, honey bees may ac- bees. Several surveys of U.S. beekeepers (vanEngels-
quire any number of infectious agents as a consequence dorp et al. 2008, 2010, 2011) have consistently identi-
of spatial overcrowding and equipment sharing. The cost Þed “poor queens” as one of the top management
to prevent and treat disease is considerable, and a central concerns in the apiculture community. While symp-
focus of apicultural research has been to reduce the tomatic of many management problems, premature
impact of economically important pests. Such ap- supersedure and stored-sperm depletion are typical
proaches have relied heavily on chemotherapies, which complaints attributed to poor queens. Delaney et al.
can have negative effects on queens (Haarmann et al. (2011) recently found that commercial queens sam-
2002) and hive products (e.g., Tsigouri and Menkissoglu pled from Western and Southeastern queen breeders
2001). Moreover, there is mounting evidence that many were sufÞciently inseminated (⬇4 million sperm)
pests are becoming increasingly resistant to the more with a high average number of drones (⬇16 mates),
common treatments (Colin et al. 1997, Miyagi et al. 2000, but they only tested 24 queens (two from each sam-
Mozes et al. 2000), making them less reliable and, ulti- pled source) for their mating numbers. In this current
mately, ineffective. study, we concurrently examine the physical, insem-
Each of these concerns may be signiÞcantly impacted ination, and mating quality of a large sample of com-
by a single factor: how well a queen has mated. Multiple mercially produced queens.
mating by queen honey bees has been shown to 1)
mitigate the negative impacts of poor brood patterns as
Materials and Methods
a result of homozygosity at the sex locus and inbreeding
(Tarpy and Page 2002); 2) increase the number of stored Sampling. We purchased 10 queens each from
sperm (Schluns et al. 2005, Delaney et al. 2011), as well seven independent commercial queen producers lo-
as enhance queen attractiveness and pheromone proÞles cated in CaliforniaÕs central valley. All sources were
(Richard et al. 2007), thus prolonging longevity and re- generically coded to remain blind and retain anonym-
ducing the likelihood of supersedure; and 3) delay the ity. The genetic stock of all purchases was standard
spread of infectious diseases among nestmates (Palmer ÔItalianÕ (presumably A. m. ligustica L.) honey bees,
and Oldroyd 2003, Tarpy 2003, Seeley and Tarpy 2007). although we purchased an additional 10 ÔCarniolanÕ
As such, increasing the “mating health” of honey bee queens (presumably A. m. carnica) from Source C,
queens will likely beneÞt beekeepers by simultaneously resulting in a total of 80 experimental queens. The
increasing queen longevity, brood viability, and disease commercial operations were not informed as to
tolerance. the purchasers (North Carolina State University
A niche market within the apiculture industry is the [NCSU]) or the beesÕ use in a scientiÞc study (all
mass production and sale of open-mated queens to queens were ordered by an individual in our program
beekeepers. Commercial queens typically sell for $15 and shipped to his private residence). Therefore, this
to $25 each, and upwards of one million queens are represents a random sampling of queens that are com-
raised and sold every year in the United States (Cobey mercially available from each operation.
et al. 2011). However, there have been surprisingly Upon arrival at NCSU, we introduced each queen
few studies that have systematically and thoroughly into her own Þve-frame nucleus colony at the Lake
assessed mating frequency, insemination success, and Wheeler Honey Bee Research Facility following stan-
morphometric characters of queens produced by the dard methods. After their acceptance by the workers,
commercial queen-production industry in the United we permitted each queen to lay eggs for 3Ð5 wk before
States. Camazine et al. (1998) investigated the health sampling all queens within a source group. From each
of commercially purchased queens in the United colony, we caged a frame of capped brood and
States. In addition to bioassays of two adult parasites, emerged them in an incubator set at 34⬚C and ⬇50%
they estimated the stored sperm number of over 300 RH to sample newly emerged worker daughters from
queens from many different sources. In that sample, each queen, which were subsequently frozen at
almost one in Þve queens were “poorly mated,” de- ⫺80⬚C for paternity analysis (see below).
Þned as fewer than three million stored sperm Morphological Measures. We measured all queens
(Woyke 1962). While very telling, these Þndings pro- from a given commercial source concurrently once
vide a limited and somewhat outdated view of the their brood was sampled (see Delaney et al. 2011). We
mating health of commercial honey bee queens. immobilized each laying queen by temporarily placing
The physical and reproductive health of the queen her in a freezer and then recorded her fresh weight to
honey bee is a vital component to a colonyÕs well- the nearest 0.1 mg on a digital scale. We then measured
being, both genetically and socially. Commercial her thorax width and head width to within 0.1 mm
queen breeders supply most of the queens used by using digital calipers, then sacriÞced each queen by
migratory beekeepers who move thousands of honey decapitation, dissected the queenÕs abdomen, and re-
bee colonies along pollination routes across the coun- moved her spermatheca. We measured the diameter
try. The commercial and hobbyist beekeepers alike of the spermatheca using a calibrated ocular in a dis-
22 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 105, no. 1

Table 1. Comparisons of morphological characters of commercial queens

Fresh wt. Thorax width Head width Spermatheca


Source n
(mg) (mm) (mm) vol (mm3)
A 10 234.4 ⫾ 16.30a 4.43 ⫾ 0.295a 3.60 ⫾ 0.269a 0.90 ⫾ 0.142bc
B 8 221.0 ⫾ 15.27a 4.46 ⫾ 0.265a 3.59 ⫾ 0.232a 1.09 ⫾ 0.181ab
C(C) 7 233.0 ⫾ 15.42a 4.41 ⫾ 0.188ab 3.51 ⫾ 0.182ab 1.13 ⫾ 0.114a
C(I) 6 225.8 ⫾ 14.96a 4.45 ⫾ 0.067ab 3.44 ⫾ 0.190ab 0.93 ⫾ 0.050abc
D 7 220.1 ⫾ 16.37ab 4.36 ⫾ 0.221ab 3.38 ⫾ 0.211ab 0.90 ⫾ 0.176bc
E 7 221.2 ⫾ 20.58a 4.24 ⫾ 0.192ab 3.46 ⫾ 0.238ab 0.95 ⫾ 0.123ab
F 9 195.9 ⫾ 11.42bc 4.13 ⫾ 0.111b 3.24 ⫾ 0.161b 0.73 ⫾ 0.079c
G 5 191.1 ⫾ 7.90c 4.26 ⫾ 0.069ab 3.37 ⫾ 0.178ab 0.88 ⫾ 0.134abc
Avg. 218.7 ⫾ 20.66 4.34 ⫾ 0.226 3.45 ⫾ 0.233 0.94 ⫾ 0.175

Different commercial sources are coded for aninimity, both Italian (I), and Carniolan (C) queens were purchased from source C and were
analyzed separately. Different letters after each measure signify statistically signiÞcant differences using Tukey post hoc tests at ␣ ⫽ 0.05.

secting microscope and calculated its spherical vol- ance (ANOVA) with Tukey post hoc tests. We then
ume (see Hatch et al. 1999). used multivariate analyses using pairwise correlations
Sperm Counts and Viability. To quantify stored to quantify the relationships among all measures of
sperm number and viability, we followed the staining queen quality. We then conducted a Principal Com-
protocol of Collins and Donoghue (1999). Immediately ponent Analysis on the covariances of the measure-
after dissection, each spermatheca was ruptured in 0.5 ml ments and saved the top two PCs for comparisons
1.0 M HEPES. The ruptured spermatheca was removed among the different commercial sources. All means
and the remaining sperm and HEPES was transferred are reported with ⫾1 SD unless noted otherwise, and
into a 2 ml vial. We added 10 ␮l of Sybr 14 in DMSO to all statistical comparisons are two-tailed with a signif-
the vial and incubated the vial for 7 min in a 36⬚C water icance of ␣ ⫽ 0.05.
bath. We then added 5 ␮l of propidium iodide and again
incubated the vial for 7 min in a 36⬚C water bath
Results
(adapted from LIVE/DEAD Sperm Viability Kit, Mo-
lecular Probes, Eugene, OR). The stained sperm (⬇10 ␮l Several of the queens did not survive original ship-
of solution) was immediately loaded onto a hemacytom- ment, and several more were either not accepted by
eter. A Zeiss Axioskopepi ßuorescent microscope (Carl the workers in their introduced nucleus colonies or
Zeiss, Hanover, MD) with a digital camera and FITC and died before sampling, thus a total of 61 queens were
Rhodamine Þlters was used to visualize both live and analyzed. There was no signiÞcant pattern of queen
dead sperm. Live sperm ßuoresce at green wavelengths mortality with respect to their purchased source and
(Em 520Ð570 nm) while dead sperm ßuoresce at red thus were common losses experienced with overnight
wavelengths (Em 650 nm). We took Þve separate pic- shipping of queens. There were signiÞcant differences
tures (nonoverlapping Þelds of view) using Þrst the among the sources with respect to all morphological
FITC Þlter and then the Rhodamine Þlter. The numbers measurements of the queens, including fresh weight
of live and dead sperm were counted for each picture (F ⫽ 7.74; df ⫽ 7, 51; P ⬍ 0.0001), thorax width (F ⫽
under the differing Þlters and the percent of viable sperm 2.79; df ⫽ 7, 51; P ⬍ 0.05), head width (F ⫽ 2.64; df ⫽
present from each spermatheca was calculated. The av- 7, 48; P ⬍ 0.05), and spermatheca volume (F ⫽ 7.19;
erage sperm count from the Þve Þelds of view was cor- df ⫽ 7, 51; P ⬍ 0.0001; Table 1).
rected for dilution to calculate the total number of stored When comparing measurements of queen insemi-
sperm. nation, however, there were less pronounced differ-
Mating Numbers. We calculated the observed mat- ences among queen producers (Table 2). Overall, the
ing number and effective paternity frequency for each average number of stored sperm in the 59 queens
queen following Tarpy et al. (2010). Brießy, we ex- examined was 4.37 ⫾ 1.446 million, and there were no
tracted whole genomic DNA from the legs of up to 48 signiÞcant differences in sperm counts among the
workers from each colony using Chelex 100 resin sources (F ⫽ 1.18; df ⫽ 7, 51; P ⫽ 0.33). While one
(Walsh et al. 1991) and subjected it to two multiplexed queen from Source E was shown to still be a virgin (no
polymerase chain reactions (PCRs) for eight microsat- detectable stored sperm), only eight queens (13.6%)
ellite loci (Am010, Am043, Am052, Am059, Am061, were Ôpoorly inseminatedÕ (i.e., had fewer than three
Am098, Am125, and Am553). We analyzed the PCR million stored sperm). Given the calculated volume of
products on a 3730 ABI sequencer through the NCSU the spermatheca (see above) and the average size of
Genomic Science Laboratory, and we inferred the pa- an individual spermatozoon, we also calculated the
ternity of each worker using COLONY (Wang 2004) percentage of the maximum Þlled capacity of each
from which we were able to calculate the observed queen following (Tarpy et al. 2011). The average for
mating number (No) and the effective paternity fre- all queens was 47.0 ⫾ 20.05%, and there were no
quency (me) of each queen following Nielsen et al. signiÞcant differences among queen sources (F ⫽ 1.39;
(2003). df ⫽ 7, 51; P ⫽ 0.23). There were, however, highly
Statistical Analyses. We compared the different signiÞcant differences in the percentage of viable
commercial sources using one-way analysis of vari- sperm across sources (F ⫽ 7.24; df ⫽ 7, 49; P ⬍ 0.0001),
February 2012 TARPY ET AL.: MATING HEALTH OF QUEEN BEES 23

Table 2. Comparisons of insemination and mating characters of commercial queens

Stored sperm Sperm Percent Observed Effective paternity


Source n
(⫻106) viability (%) Þlled mating no. (No) frequency (me)
A 9 4.96 ⫾ 1.648 92.6 ⫾ 4.60ab 54.1 ⫾ 28.26 20.3 ⫾ 5.20 21.4 ⫾ 9.40
B 8 4.37 ⫾ 1.582 78.0 ⫾ 7.44b 39.0 ⫾ 12.79 18.1 ⫾ 4.94 16.4 ⫾ 7.94
C(C) 7 4.98 ⫾ 1.113 85.4 ⫾ 8.24ab 42.6 ⫾ 9.94 20.2 ⫾ 5.42 21.8 ⫾ 11.77
C(I) 6 3.54 ⫾ 1.452 86.0 ⫾ 8.86ab 36.4 ⫾ 14.39 16.7 ⫾ 5.32 14.6 ⫾ 9.20
D 7 4.43 ⫾ 1.018 82.8 ⫾ 9.38ab 49.9 ⫾ 18.02 16.3 ⫾ 2.87 11.4 ⫾ 6.32
E 7 3.67 ⫾ 1.748 79.6 ⫾ 5.96ab 38.4 ⫾ 18.22 20.0 ⫾ 4.84 21.0 ⫾ 8.58
F 9 3.95 ⫾ 1.005 94.0 ⫾ 1.73a 52.8 ⫾ 14.06 15.8 ⫾ 4.66 12.2 ⫾ 5.24
G 6 4.96 ⫾ 1.710 59.4 ⫾ 26.47c 60.5 ⫾ 31.34 17.6 ⫾ 4.04 15.1 ⫾ 8.59
Avg. 4.37 ⫾ 1.446 83.7 ⫾ 13.33 47.0 ⫾ 20.05 18.2 ⫾ 4.84 17.0 ⫾ 8.98

Coded sources are the same as in Table 1. Different letters after each measure signify statistically signiÞcant differences using Tukey post
hoc tests at ␣ ⫽ 0.05, which were only signiÞcant for sperm viability and none of the other variables.

where the average sperm viability was 83.7 ⫾ 13.33 but for PC1 were all positive (except for percentage
with some queens having as low as 20.5% viable sperm. Þlled), whereas PC2 was negatively associated with all
There was a signiÞcant effect of processing date on morphometric characters (listed in Table 1) and pos-
sperm viability (F ⫽ 4.09; df ⫽ 5, 51; P ⬍ 0.005), but itively associated with all “mating” variables (listed in
there was no consistent downward trend over time so Table 2). Overall, there was no signiÞcant correlation
that temporal effects on sperm longevity (e.g., Burley between the two principal components (r2 ⫽ 0.049;
et al. 2008) was likely not a factor. P ⫽ 0.66), although there was one commercial source
We genotyped a total of 2,798 individual workers where there was a strong negative relationship
(45.9 ⫾ 1.97 per colony) and assigned them to their (Source G: r2 ⫽ ⫺0.807; P ⬍ 0.005). A one-way
respective patriline within their colonies to infer ANOVA on PC1 demonstrates signiÞcant differences
queen mating numbers. Overall, the mating numbers among queen sources for this single composite of
of the tested queens were higher than expected (Ta- queen reproductive quality (F ⫽ 7.28, df ⫽ 7, 46, P ⬍
ble 2). The average observed mating number was 0.0001; Fig. 1), but there were no differences for PC2
18.2 ⫾ 4.84 and the average effective paternity fre- (F ⫽ 1.07; df ⫽ 7, 46; P ⫽ 0.40).
quency was 17.0 ⫾ 8.98, both of which are numerically
(but not statistically) higher than their global averages
Discussion
for the species (Tarpy and Nielsen 2002). As with the
insemination measures, there were no signiÞcant dif- The intent of this study was to determine if commer-
ferences in mating frequency across commercial cially produced honey bee queens are of low reproduc-
sources (observed: F ⫽ 1.22; df ⫽ 7, 53; P ⫽ 0.31; tive potential and may therefore explain their diminished
effective: F ⫽ 1.89; df ⫽ 7, 53; P ⫽ 0.09). quality perceived by the industry. However, our data
There were many statistically signiÞcant correla- shows very little, if any, evidence that this is the case for
tions among the measured variables, particularly newly mated queens. These Þndings corroborate Dela-
among the morphological measurements (data not ney et al. (2011), who also suggest that commercially
shown). Similar to Schluns et al. (2005) and Delaney
et al. (2011), we found a signiÞcant nonlinear (loga-
rithmic) relationship between effective paternity fre-
quency and stored sperm number (F ⫽ 7.97; df ⫽ 1, 55;
P ⬍ 0.01). We did not, however, verify the observed
relationships between thorax width and sperm num-
ber (F ⫽ 2.97; df ⫽ 1, 57; P ⫽ 0.09) or mating frequency
(F ⫽ 1.03; df ⫽ 1, 56; P ⫽ 0.31) that were reported in
our previous study (Delaney et al. 2011).
No singular variable can adequately capture queen
reproductive potential in its entirety, and thus many
different measurements can each reßect a different
facet of this complex quantitative trait. A proxy of
these measures can be generated in multi-dimensional
space using Principle Component Analysis (PCA), in
essence to collapse multiple correlated variables into Fig. 1. Plot of the two principal components (PC1 and
one or a few composite variables. PCA on the cova- PC2) grouped by commercial source. No one variable ade-
riances of the nine measured variables yielded two quately measures queen reproductive potential, but a compos-
ite of many different measures can be generated in multi-di-
prominent principal components that accounted for mensional space using PCA. PCs were generated from a
99.2% of the total variance in queen reproductive principal component analysis on the covariances of all nine
potential, the Þrst (PC1) accounting for 81.6% of the measured variables (Tables 1 and 2) and together explain over
total variance and the second (PC2) accounting for an 99% of the overall variation in queen quality. Ellipses signify 50th
additional 17.6%. The loading variables (eigenvectors) percentiles on both axes for each identiÞed source.
24 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 105, no. 1

produced queens are generally high in their reproduc- Therefore, it is likely that if queen quality is an important
tive potential. The current population of queens had very factor in honey bee management, then testing the nu-
low levels of the gut microsporidian Nosema, as only merous environmental factors that can inßuence queen
three of the 59 queens tested had detectable levels as productivity after mating should take priority. Future
determined by PCR analysis following (Webster et al. research should emphasize the potential roles of factors
2008, data not shown). However, because the queens that affect queen quality before (e.g., genetic bottle-
were sampled 3Ð5 wk after being in nucleus colonies, and necks, pedigree relationships among breeding popula-
we did not sample for Nosema among the workers in tions) and subsequent to mating (e.g., mechanisms of
those colonies, we cannot determine if they were in- supersedure, capacity to store live sperm) to address
fected prior or subsequent to colony introduction. Nev- these concerns.
ertheless, it is quite clear that contemporary commercial
queens have signiÞcantly lower parasite loads than in the
Acknowledgments
past (see Farrar 1947, Furgala 1962, Liu et al. 1987, Ca-
mazine et al. 1998). We thank John Harman, Winnie Lee, Flora Lee, Mithun
We did not consistently observe strong correlations Patel, and Matt Mayer for their help in DNA extractions and
between body size and mating success. Delaney et al. PCR analyses. We also express our appreciation and gratitude
(2011) found signiÞcant relationships between thorax to Marla Spivak, the CA Bee Breeders, and the NC State
width and both stored sperm and effective paternity Social Insect Working Group for their cooperation and many
helpful discussions subsequent to the collection of queens
frequency, but fresh weight did not show a similar used in this study. Special thanks goes to Laura Mathies for
correlation (see also Tarpy et al. 2011). In the current use of her ßuorescent microscope for the sperm analyses
study, we found a marginally signiÞcant relationship without which those measures would have been impossible.
between weight and mating number (No: r2 ⫽ 0.260, This study was supported by the North Carolina Department
P ⬍ 0.05; me: r2 ⫽ 0.245, P ⫽ 0.06), but no such of Agriculture and Consumer Services and grant number
relationships with thorax width. It is possible that 2007-02281 from the National Research Initiative of the
sample size may explain these differences; the current USDA Cooperative State Research, Education and Extension
study compared a total of 61 queens for all morpho- Service awarded to DRT.
logical and mating variables, whereas Delaney et al.
(2011) tested a total of 117 queens but only 22 of them References Cited
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