Journal of Heredity 2011:102(5):562–566
doi:10.1093/jhered/esr075
Advance Access publication July 20, 2011
Ó The American Genetic Association. 2011. All rights reserved.
For permissions, please email: [email protected].
Asexually Produced Cape Honeybee
Queens (Apis mellifera capensis)
Reproduce Sexually
MADELEINE BEEKMAN, MICHAEL H. ALLSOPP, JULIANNE LIM, FRANCES GOUDIE,
AND
BENJAMIN P. OLDROYD
From the Behaviour and Genetics of Social Insects Lab, School of Biological Sciences A12, University of Sydney, Sydney,
NSW 2006, Australia (Beekman, Lim, Goudie, and Oldroyd); and the Honey Bee Research Section, ARC-Plant Protection
Research Institute, Stellenbosch, South Africa (Allsopp).
Address correspondence to M. Beekman at the address above, or e-mail: [email protected].
Abstract
Unmated workers of the Cape honeybee Apis mellifera capensis can produce female offspring including daughter queens. As
worker-laid queens are produced asexually, we wondered whether these asexually produced individuals reproduce asexually
or sexually. We sampled 11 colonies headed by queens known to be the clonal offspring of workers and genotyped 23
worker offspring from each queen at 5 microsatellite loci. Without exception, asexually produced queens produced female
worker offspring sexually. In addition, we report the replacement of a queen by her asexually produced granddaughter, with
this asexually produced queen also producing offspring sexually. Hence, once a female larva is raised as a queen, mating and
sexual reproduction appears to be obligatory in this subspecies, despite the fact that worker-laid queens are derived from
asexual lineages.
Key words: Apis mellifera capensis, asexual reproduction, Cape honeybee, mating, sexual reproduction
Most animals reproduce sexually. The near ubiquity of
sexual reproduction is intriguing because of the so-called 2fold cost of sex: Any lineage of females that produces
female offspring asexually will grow at twice the rate as
a lineage that reproduces sexually (Maynard Smith 1978).
The predominance of sexual reproduction is presumably the
outcome of selective tradeoffs between the benefits and
costs of sexual and asexual reproduction. What exactly these
tradeoffs are is as yet unclear, though most evolutionary
models focus on the benefits of recombination as a driving
force behind sexual reproduction (Hurst and Peck 1996;
Otto and Lenormand 2002).
In the social Hymenoptera (all ants, some bees, and
some wasps), females are produced sexually and are diploid,
whereas males develop from unfertilized eggs and are
haploid. Diploid fertilized eggs can develop into queens or
workers. The developmental switch is primarily under
environmental control, though in many species (but not
honeybees), there is a genetic component to caste determination as well (Schwander et al. 2010).
In 5 species of ant (Cagniant 1979; Itow et al. 1984;
Heinze and Hölldobler 1995; Tsuji and Yamauchi 1995;
Grasso et al. 2000) and the Cape honeybee from South
Africa (a subspecies of Apis mellifera) (Onions 1914),
562
unmated workers can produce female offspring via thelytoky. Furthermore, in at least 2 ant species, Cataglyphis cursor
(Pearcy et al. 2004) and Wasmannia auropunctata (Fournier
et al. 2005), queens produce new queens asexually, whereas
workers are produced sexually. Thus, queens effectively
prevent the genomes of their mates from being transferred
to the next generation of queens, thus avoiding the 2-fold
cost of sex in reproductive offspring while enjoying the
benefits of a genetically diverse sexually produced worker
population (Fournier et al. 2005; Oldroyd and Fewell 2007).
We have previously shown that workers of the Cape
honeybee (A. mellifera capensis) lay eggs in queen cells when
their colony is preparing for reproductive swarming (Jordan
et al. 2008; Allsopp et al. 2010) and when new queens are
produced in an emergency due to queen loss (Holmes et al.
2010). Moreover, laying eggs in queen cells appear to be
a successful parasitic strategy whereby nonnatal workers, in
genetical terms, become the next queen of a colony they
were not born in (Jordan et al. 2008; Allsopp et al. 2010;
Holmes et al. 2010).
These observations raise an interesting question about
the likely behavior of thelytokously produced queen
offspring of workers. Do such queens reproduce sexually
or asexually once they head a colony?
Beekman et al. Sexual Versus Asexual Reproduction in Honeybees
ter, that is, the thelytokously produced daughter of a sexually
produced natal worker. These samples were genotyped at 8
linked loci (Goudie et al. unpublished results).
Results
In all our 11 colonies, we found more than 2 alleles at each
locus, except for A29, colony 8 (Table 1). Therefore, we can
conclude that all 11 queens had mated and produced
workers sexually. Similarly, the thelytokous granddaughter
of the original queen in our colony in which the queen was
superseded had also mated and was actively producing
worker offspring sexually at the time the colony was
sampled (Table 2).
Discussion
Figure 1. Hypothetical genotypes of workers produced
asexually (left panel) and sexually (right panel). For simplicity,
only one haploid male is shown in the right panel, but because
honeybee queens mate with multiple males, sexually produced
workers will show a diversity of paternal alleles if they are sired
by different fathers. We therefore expect to find multiple
paternal alleles among our worker population if the queen has
mated. If, on the other hand, the queen is unmated and
produced offspring asexually, we only expect to find maternal
alleles in the worker population.
Materials and Methods
We collected preemergent worker-brood from 11 A.
m. capensis colonies from the Stellenbosch area (lat
33°56#S, long 18°51#E), Western Cape, South Africa. These
colonies were headed by marked queens that were known to
be the offspring of workers (Allsopp et al. 2010; Holmes
et al. 2010). Samples were collected during the summers of
2008 and 2009. For each colony, we genotyped 23
preemergent workers at 5 microsatellite loci (A113, A29,
A88, B124, and A7; Solignac et al. 2003). (See Holmes et al.
(2010) for details of the genetic analysis.) We then identified
the maternal alleles for each locus for each colony (see
Figure 7.6 in Oldroyd and Wongsiri (2006)) and then
considered, for each worker, whether they carried a paternally derived allele at any locus thus indicating that they had
been produced sexually (Figure 1). Because none of the
colonies were raising new queens at the time of sampling,
we did not expect workers to contribute significantly to
worker brood (Beekman et al. 2009). However, because
workers produce offspring clonally, even if our samples did
include some worker-produced workers, this would not
affect our conclusion with respect to the mode of their
mother queen’s reproduction: An asexually produced
worker-offspring of a sexually produced worker will also
carry paternal alleles.
We also included a colony in which the original queen
was superseded by her thelytokously produced granddaugh-
Our data show that Cape honeybee queens that are the
thelytokous daughters of workers mate and produce
workers sexually. Our previous studies also suggest that
surviving queen-laid larvae in queen cells are also sexually
produced (Jordan et al. 2008; Allsopp et al. 2010). Thus,
clonally produced queens revert to sexual reproduction in
one generation. This is interesting because asexual lineages
of workers can persist as reproductive parasites for at least
a decade (Oldroyd et al. 2011).
The mechanism of thelytoky in A. m. capensis (automixis
with central fusion, Verma and Ruttner 1983) results in
increased levels of homozygozity for each generation of
asexual reproduction for all loci that are free to recombine
(Baudry et al. 2004; Pearcy et al. 2006; Oldroyd et al. 2008).
In addition to the disadvantages of loss of complementation
in general (Archetti 2004), increased levels of homozygozity
have a severe fitness cost in honeybees due to the method of
sex determination. The expression of the female sex requires
heterozygosity at the complementary sex determining locus or csd
(Beye et al. 2003). Individuals that are homozygous at the
sex locus are inviable diploid males (Woyke 1963; Beye et al.
2003; Gempe et al. 2009). Because the sex locus is distant
from the centromere on chromosome 3 (Solignac et al.
2007), and therefore free to recombine, a few generations of
thelytokous reproduction should inevitably result in loss of
heterozygosity at the sex locus and the production
of inviable diploid males instead of workers. Sexual
reproduction and polyandry allow queens to maintain high
brood viability and put their colonies at a competitive
advantage over colonies where workers (and diploid males)
are produced asexually (Page 1980). Furthermore, a genetically diverse worker population provides colony level fitness
benefits in terms of disease resistance (Seeley and Tarpy
2007), task allocation (Mattila and Seeley 2007), and nest
homeostasis (Jones et al. 2004).
The combined effect of homozygosity at the sex locus
and the fitness benefits of a genetically diverse worker
population make it unsurprising that Cape honeybee queens
mate and produce worker offspring sexually, even when
563
1
2
3
4
5
6
7
8
9
10
11
Inferred queen alleles and paternally derived alleles found in worker offspring produced by 11 worker-produced queens
Colony
A113
A29
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
211,
213,
223,
211,
133,
125,
125,
127,
133
127, 131, 135
133
128, 131, 135, 137
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
?
211, 229
213, 215, 217
129,
121,
129,
127,
137
131
137
133, 159
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
Queen alleles
Paternal alleles
215,
207,
223,
217,
225,
215,
215,
229
211,
213,
209,
215,
221,
215,
217
211, 223, 225, 227
225
221, 227
?
217, 223, 227
215
131,
124,
129,
125,
133,
129,
129,
133
129, 137
137
133, 135
137
141
135
225
215, 219, 221, 223
223
217
223
217, 219, 227, 233
128,
135,
129,
137,
128,
123,
159,
133
139
131
141, 159
128
127, 131, 133,
161
219
217,223, 225, 227
225
217, 221, 227
‘‘?’’ denotes uncertainty regarding the queen allele(s).
A88
138,
152,
144
131,
142,
138,
144,
142,
139,
154,
143,
138,
138
142,
139,
142,
142,
138,
139,
142,
139,
132,
138,
139,
149
154, 155
133,
148,
155
146,
151
143,
155
148
139,
135, 137, 138, 139,
151
144,
139
143,
144
139,
139
143,
148
138,
144
143,
148, 151, 152, 153, 154
148, 149, 151, 152
144, 146, 148, 153,
142, 152, 155
148, 149
148, 151, 154
144, 149, 152, 154
142, 143, 146, 152, 153
148, 151, 152, 154, 156
B124
A7
215,
217,
217,
211,
107, 112
96, 99, 102, 109, 116
104, 112
100, 108
228
220, 224, 226, 234, 236, 238
220
215, 219, 222, 229, 234
232, 234
213, 219
234
213, 215, 217, 219, 222, 228, 230
100, 107
96, 98, 105, 110, 113, 116
96, 110
101, 104, 105, 108, 111, 114, 116
220,
213,
213,
215,
213,
220,
220,
211,
217,
215,
213,
215,
219,
211,
109, 112
96, 99, 107, 116, 159
109, 112
96, 99, 107, 159
101, 110
100, 104, 105, 106, 107
108, 110
99, 114
105, ?
98, 99, 102, 113, 116, 117, 169
96, 105
99, 102, 104, 107, 109, 110, 114
99, 110
101, 102, 105, 107, 111, 112
243
215,
217
219,
219
230,
?
213,
219
220,
230
219,
230
215,
217, 230, 232, 234, 238
232, 241
236, 243
217, 222, 226, 228, 230
232, 240, 241
222, 232, 238
217, 241
Journal of Heredity 2011:102(5)
564
Table 1
227, 229, 231,
233, 235
174, 180, 182,
184, 188, 190
126, 133, 138,
142, 144
187, 188, 198, 190,
192, 194, 195, 196, 205
159, 207
194, 195,
196, 202, 208
222, 226, 228, 229,
230, 231, 232, 234
238, 239, 240, 242,
244, 245, 248, 257
224, 224
243, 243
140, 140
186, 193
191, 195
198, 200
224, 227
186, 186
235, 237
174, 184
240, 246
186, 192
159, 195
198, 200
224, 227
126, 140
The3
The2
UN283
KO105
KO101
BI326
Sex3
Original queen
alleles
Thelytokous
queen alleles
Paternal alleles
Table 2
Genotype of the original queen, her thelytokously produced granddaughter, and paternal alleles found in worker offspring of the asexually produced queen
The4
Beekman et al. Sexual Versus Asexual Reproduction in Honeybees
they themselves were produced thelytokously. What is
surprising is that thelytokously produced Cape queens do
not produce eggs thelytokously when producing queen
offspring (Jordan et al. 2008), as is observed in the ants C.
cursor (Pearcy et al. 2004) and W. auropunctata (Fournier et al.
2005). Unmated Cape queens can produce eggs both
thelytokously and arrhenotokously and have at least partial
control over which kind of meiosis their eggs undergo
(Oldroyd et al. 2008). Thus, it seems likely that selection
should favor asexual reproduction in Cape queens when
they lay eggs in queen cells. The homozygous queen–
produced larvae found in queen cells by Jordan et al. (2008)
may be evidence of unsuccessful attempts by queens to lay
diploid eggs in queen cells without using her mates’ sperm.
However, such homozygous individuals have not been
found beyond the third larval instar, suggesting that they are
inviable (Beekman et al. 2009).
The revision to sexual reproduction in asexually produced queens also argues against thelytoky in Cape honeybees being solely genetically determined (Lattorff et al. 2005,
2007). Should this be the case, it would seem that the
modified meiosis required for thelytokous reproduction
(Verma and Ruttner 1983) should persist in the queen caste
and be incompatible with sexual reproduction.
Funding
Australian Research Council grants to M.B. and B.P.O.
(DP0878924 and DP0985189).
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Corresponding Editor: Rob DeSalle
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