For the first time we demonstrate that brother-sister mating in B. terrestris exacts high costs under field conditions through the production of diploid males. A number of fitness parameters were negatively affected by diploid male production, including colony growth rate, total offspring production and colony survival, but no significant effects of inbreeding in the absence of diploid male production were detected.
The costs of diploid male production were first evident whilst colonies were growing in the laboratory, where the number of days between the emergence of the 1st
offspring was considerably greater for the diploid male colonies. This slower growth rate presumably occurs because colony resources are diverted away from the production of industrious female workers; diploid males are idle within the colony and so the workforce is approximately halved, resulting in less brood care and slower growth. These findings augment the study by Plowright & Pallett [11
] who found that DMP colonies in Bombus atratus
had a considerably slower rate of growth than all-worker-producing inbred colonies in laboratory conditions.
Overall colony fitness was gauged by the total number of offspring produced by the end of the experiment, as the number of reproductives reared by a colony is highly correlated with the number of workers [29
]. The mean number of offspring produced by the diploid male colonies was significantly lower than in the other treatments. In fact, the mean was only 15.8, which is barely greater than the colony size of 15 when nests were placed in the field. The low number of offspring in these colonies would result in fewer foraging workers and hence a lower food intake. This would have initially impeded growth and subsequently led to colony starvation. This is reflected in lower survival of DMP colonies; the queens survived approximately a third of the time of the outbred colonies, and died presumably due to starvation due to the lack of foraging workers. A similar outcome has been found in the fire ant Solenopsis invicta
, where DMP colonies had lower brood weight, fewer adult workers and higher queen mortality compared to all-worker-producing colonies [12
]. This was explained by the queen having to cope on her own for longer before there were sufficient workers to take over foraging duties.
As well as reducing colony survival, bumblebee diploid males impose a genetic load on populations as they yield no reproductive return for the resources invested in them. Bombus terrestris
diploid males produce diploid (rather than haploid) sperm. They also have smaller testes and fewer spermatozoa than haploid males, and hence have reduced fertility [32
]. Although they develop normally in other respects and are capable of mating, Duchateau & Marien [32
] found that the queens mated to diploid males did not produce colonies. It has since been found that such queens are capable of producing a viable colony containing triploid offspring, but the triploid queens produced are inviable or infertile [33
]. Therefore, as in other species such as the sawfly Athalia rosae ruficornis
, the costs of diploid males are not all immediately apparent, but become so a generation later [10
Diploid males have been found to be sensitive indicators of the loss of genetic diversity in Hymenoptera. For example, an apparently abundant species of orchid bee Euglossa imperiali
was found to have large numbers of diploid males, ranging from 12% to 100% of the total population. This turned out to be the result of an extremely low effective population size [34
]. In a further study of more orchid bee species, the highest diploid male frequency and the lowest genetic variability was found in the rarest species [35
]. Diploid males have also been found in rare and localised bumblebee species. In the Japanese bumblebee Bombus florilegus
, diploid males were found in 28% of colonies produced in the laboratory from wild caught mated queens, a figure thought to be due to matched matings resulting from notably low genetic diversity and small population size. Additionally, the frequency of triploid females was found to be 2.7% in natural populations [22
]. Diploid males were detected at a frequency of 5% in the wild (with respect to haploid males) in the threatened bumblebee Bombus muscorum
, again probably due to reduced genetic diversity brought about by population fragmentation and isolation [20
]. As diploid males are produced from the first brood, they will be found on the wing, even if the colony from which they have been produced dies prematurely, as our results suggest is highly likely. Because of the significant costs diploid males represent for bumblebee fitness, their frequency could potentially be used as an indicator of the genetic health of the population and hence its sustainability and conservation requirements [34
]. Where the production of diploid males is high, translocations from other populations might be considered as a means of increasing genetic diversity. However, given that DMP colonies are short-lived under field conditions, their apparent absence will not always indicate a genetically healthy population. A method of directly assessing CSD allele diversity would therefore be of great value.
In this experiment the only apparent cost of inbreeding was the production of diploid males, as the non-DMP inbred colonies did not differ significantly from the outbred colonies in all the variables measured. It should be noted, however, that this lack of difference could be due to the inbred colonies resulting from only one generation of brother-sister mating, which would not substantially decrease their level of heterozygosity relative to the outbred colonies. Indeed, one study has demonstrated decreased fecundity and colony size when B. terrestris
queens are sibmated for several generations (Beekman et al.
, 1999). Despite the fact that some evidence indicates that Hymenoptera, including bumblebees [29
], suffer from inbreeding depression, a meta-analysis has shown that the magnitude of fitness loss on inbreeding is less than that experienced by diploid insects [4
]. This supports ideas that deleterious recessive alleles are expressed and thus purged in haploid males [3
]. Our data show that the high costs of DMP following inbreeding far outweigh any apparently small effects of conventional inbreeding depression. Thus, whilst Hymenoptera may be spared some costs of inbreeding by virtue of their haplodiploidy, their sex determination system imposes unique costs through diploid male production. Due to these negative fitness effects, selection should act strongly on haplodiploids to avoid incestuous matings and the production of diploid males, a theory that has been supported by a recent study [36
]. There is some evidence to suggest that this avoidance behaviour occurs through a kin recognition system [37
Hibernation survival and colony growth in the laboratory were significantly influenced by maternal family line. This among-family variation has been found in a number of different fitness traits in bumblebees [28
] and is evidently an important aspect of their evolutionary ecology. The factors that maintain this variation in wild populations remain to be established. Mating date was another significant predictor of the variation in hibernation survival observed; queens that were mated first were more likely to survive than those mated at a later date, despite standardised hibernation duration and conditions. This substantiates the idea that individuals that are born and reproduce early in the season have a higher survival rate and fitness [29